Detection of polyomavirus simian virus 40
tumor antigen DNA in AIDS-related systemic non-Hodgkin lymphoma.
Vilchez RA, Lednicky JA, Halvorson SJ, White ZS, Kozinetz CA, Butel JS.
Department of Medicine, Baylor College of Medicine, Houston, Texas 77030, USA.
rvilchez@bcm.tmc.edu
Systemic non-Hodgkin lymphoma (S-NHL) is a common malignancy during HIV
infection, and it is hypothesized that infectious agents may be involved in the
etiology. Epstein-Barr virus DNA is found in <40% of patients with AIDS-related
S-NHL, suggesting that other oncogenic viruses, such as polyomaviruses, may play
a role in pathogenesis. We analyzed AIDS-related S-NHL samples, NHL samples from
HIV-negative patients, peripheral blood leukocytes from HIV-infected and
-uninfected patients without NHL, and lymph nodes without tumors from
HIV-infected patients. Specimens were examined by polymerase chain reaction
analysis with use of primers specific for an N-terminal region of the
oncoprotein large tumor antigen ( T-ag ) gene conserved among all three
polyomaviruses (simian virus 40 [SV40], JC virus, and BK virus).
Polyomavirus T-ag DNA sequences, proven to be
SV40-specific, were detected more frequently in AIDS-related S-NHL samples (6 of
26) than in peripheral blood leukocytes from HIV-infected patients (6 of 26 vs.
0 of 69; p =.0001), NHL samples from HIV-negative patients (6 of 26 vs. 0 of 10;
p =.09), or lymph nodes (6 of 26 vs. 0 of 7; p =.16).Sequences of C-terminal T-ag DNA from SV40 were
amplified from two AIDS-related S-NHL samples. Epstein-Barr virus DNA
sequences were detected in 38% (10 of 26) AIDS-related S-NHL samples, 50% (5 of
10) HIV-negative S-NHL samples, and 57% (4 of 7) lymph nodes. None of the S-NHL
samples were positive for both Epstein-Barr virus DNA and SV40 DNA. Further
studies of the possible role of SV40 in the pathogenesis of S-NHL are warranted.
SV40 Tag-specific cytotoxic T lymphocytes
generated from the peripheral blood of malignant pleural mesothelioma patients.
Bright RK, Kimchi ET, Shearer MH, Kennedy RC, Pass HI.
Laboratory of Prostate Cancer Biology, Robert W. Franz Cancer Research Center,
Earle A. Chiles Research Institute, and the Oregon Cancer Center, 4805 NE Glisan
Street, Portland, OR 97213, USA, rbright@providence.org
Malignant pleural mesothelioma (MPM) is an aggressive cancer, with survival of
less than one year following diagnosis and treatment with current protocols.
Recent studies have demonstrated the presence
of the simian virus 40 (SV40)-like, large tumor antigen (Tag) in nearly 60% of
MPMs. SV40 Tag is a viral-encoded tumor-specific antigen, and thus a
potential target for the induction of anti-tumor immunity and the development of
therapeutic vaccines. We describe here evidence for the existence of SV40
Tag-specific immune responses in patients with MPM whose tumors express Tag.
Humoral immunity was demonstrated by the detection of IgG titers against Tag in
serum samples from 1/3 of patients examined. CTLs were generated from the
peripheral blood of an HLA-A2(+) MPM patient with a synthetic peptide
representing an HLA-A2 binding epitope in SV40 Tag. The CTLs demonstrated
epitope fine specificity, in that other peptides from SV40 Tag and a peptide
from influenza virus were not recognized in the context of HLA-A2. Moreover, the
CTLs were capable of recognizing mesothelioma tumor cells that expressed SV40
Tag, in an MHC class I restricted manner.
SV40 infection induces telomerase activity in
human mesothelial cells.
Foddis R, De Rienzo A, Broccoli D, Bocchetta M, Stekala E, Rizzo P, Tosolini
A, Grobelny JV, Jhanwar SC, Pass HI, Testa JR, Carbone M.
Cancer Immunology Program, Department of Pathology, Cardinal Bernardin Cancer
Center, Loyola University Chicago, Maywood, Illinois, IL 60153, USA.
Mesotheliomas are malignant tumors of the pleural and peritoneal membranes which
are often associated with asbestos exposure and with Simian virus 40 (SV40)
infection. Telomerase activity is repressed in somatic cells and tissues but is
activated in immortal and malignant cells. We evaluated telomerase activity in
seven primary malignant mesothelioma biopsies and matched lung specimens and 20
mesothelioma cell lines and eight corresponding primary tumor cultures. All the
tumor biopsies, and nearly all primary cell mesothelioma cultures and cell lines
were telomerase positive. The findings in cell lines paralleled those observed
in primary cultures in cases where paired samples were available.
Next, we found that SV40, a DNA tumor virus
present in approximately 50% of mesothelioma biopsies in the USA, induced
telomerase activity in primary human mesothelial cells, but not in primary
fibroblasts. Telomerase activity became detectable as early as 72 h following
wild-type (strain 776) SV40 infection, and a clear DNA ladder was detectable 1
week after infection. The amount of telomerase activity increased during passage
in cell culture and appeared to parallel increases in the cellular amounts of
the SV40 large T-antigen. Thus, SV40 infection leads to telomerase activity
before the infected mesothelial cells become transformed and immortalized.
SV40 infection of human fibroblasts did not cause detectable telomerase
activity. We also determined that the SV40 small t-antigen (tag) plays an
important role in inducing telomerase activity because this activity was
undetectable or minimal in mesothelial cells infected and/or transformed by SV40
tag mutants. Asbestos alone did not induce telomerase activity, and asbestos did
not influence telomerase activity in mesothelial cells infected with SV40.
Induction of telomerase activity by SV40 may be related to the very high rate of
mesothelial cell immortalization that is characteristically associated with SV40
infection of mesothelial cells.
Simian virus 40 large tumor antigen forms
specific complexes with p53 and pRb in human brain tumors.
Zhen H, Zhang X, Zhang Z, Fei Z, He X, Liang J, Huang W, Liu X, Zhang P.
Department of Neurosurgery, Xijing Hospital, Institute of Neurosurgery of PLA,
Fourth Military Medical University, Xi'an 710032, China. zh_ha@263.net
OBJECTIVE: To study the role of simian virus 40 (SV40) early region gene coding
product large tumor antigen (Tag) expression and the interaction between Tag and
tumor suppressors p53 and pRb in human brain tumorigenesis. METHODS: Tag was
investigated by immunoprecipitation followed by silver staining and Western blot
in 65 cases of human brain tumors and 8 cases of normal brain tissues. Tag-p53
and Tag-pRb complexes were screened in 18 and 15 Tag positive tumor tissues,
respectively. RESULTS: Tag was found in all 8 ependymomas and 2 choroid plexus
papillomas, 90% of pituitary adenomas (9/10), 73% of astrocytomas (11/15), 70%
of meningiomas (7/10), 50% of glioblastomas multiforme (4/8), 33% of
medulloblastomas (2/6). 5 oligodendrogliomas, 1 pineocytoma, and 8 normal brain
tissues were negative for Tag. Tag-p53 complex was detected in all 18 Tag
positive tumors. Tag-pRb complex was found in all 15 Tag positive tumors.
CONCLUSION: SV40 Tag is expressed in human
brain tumors and can form specific complexes with tumor suppressors p53 and pRb.
The inactivation of p53 and pRb due to the formation of Tag-p53 and Tag-pRb
complexes may be an important mechanism in the etiopathogenesis of human brain
tumors.
Increasing evidence for involvement of SV40
in human cancer.
Butel JS.
Department of Molecular Virology and Microbiology, Baylor College of Medicine,
One Baylor Plaza, Houston, TX 77030, USA. jbutel@bcm.tmc.edu
SV40, a small DNA virus, is known to possess
strong oncogenic potential. Millions of people were exposed to SV40 as an
unknown contaminant of some early poliovaccines. This article briefly summarizes
the increasing evidence of the association of SV40 with certain types of human
cancer, including mesotheliomas and brain tumors. Unanswered questions
pertaining to the pathogenesis of human infections by SV40 and the functional
role of the virus in tumor development are noted. It is concluded that SV40
should be considered a candidate human tumor virus and that vigorous efforts to
clarify the role of the virus in human disease should be supported.
Cellular transformation by SV40 large T
antigen: interaction with host proteins.
Ali SH, DeCaprio JA.
Department of Adult Oncology, Dana-Farber Cancer Institute and Harvard Medical
School, Boston, MA 02115, USA.
SV40 large T antigen (TAg) is a powerful
oncoprotein capable of transforming a variety of cell types. The
transforming activity of TAg is due in large part to its perturbation of the
retinoblastoma (pRB) and p53 tumor suppressor proteins. In addition, TAg binds
to several other cellular factors, including the transcriptional co-activators
p300 and CBP, which may contribute to its transformation function. Several other
features of TAg that appear to contribute to its full transformation potential
are yet to be completely understood. Study of TAg therefore continues to provide
new insights into the mechanism of cellular transformation. Copyright 2001
Academic Press.
Role of T antigen interactions with p53 in
tumorigenesis.
Pipas JM, Levine AJ.
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA
15260, USA.
SV40 induces neoplastic transformation by
disabling several key cellular growth regulatory circuits. Among these
are the Rb- and p53-families of tumor suppressors. The multifunctional,
virus-encoded large T antigen blocks the function of both Rb and p53. Large T
antigen uses multiple mechanisms to block p53 activity, and this action
contributes to tumorigenesis, in part, by blocking p53-mediated growth
suppression and apoptosis. Since the p53
pathway is inactivated in most human tumors, T antigen/p53 interactions offer a
possible mechanism by which SV40 contributes to human cancer. Copyright
2001 Academic Press.
SV40 and cell cycle perturbations in
malignant mesothelioma.
Testa JR, Giordano A.
Human Genetics Program, Fox Chase Cancer Center, Philadelphia, PA 19111, USA.
JR.Testa@fccc.edu
Although epidemiological findings have
established that exposure to asbestos fibers is the major cause of malignant
mesothelioma (MM), recent studies have implicated simian virus 40 (SV40) in the
etiology of some of these tumors. Cytogenetic and molecular genetic
evidence suggests that multiple somatic genetic events are required for
tumorigenic conversion of a mesothelial cell. As with many other types of
cancer, in MM critical oncogenic events exert their action via perturbations of
the cell cycle. Interactions between the
retinoblastoma (Rb) family of proteins and oncoproteins encoded by SV40 lead to
cell cycle alterations. Likewise, inhibition of the p53 tumor suppressor by SV40
can inactivate a crucial cell cycle checkpoint, thereby permitting cells to
undergo mitosis regardless of the presence of DNA damage. Many MMs
exhibit loss and/or inactivation of the tumor suppressors p16(INK4a)and
p14(ARF), components of the pRb and p53 cell cycle regulatory pathways,
respectively. Recent investigations have demonstrated that SV40 large T antigen,
isolated from frozen biopsies of human MM specimens, binds to and inactivates
various tumor suppressor gene products such as pRb and p53.
In this review, we discuss how SV40-oncosuppressor interactions can lead to
functional alterations of the pRb- and p53-dependent cell cycle regulatory
pathways and thereby contribute to neoplastic transformation of human
mesothelial cells. Copyright 2001 Academic Press.
Publication Types:
·Review
·Review, Tutorial
PMID: 11243897 [PubMed - indexed for MEDLINE]
Simian virus 40 regulatory region structural
diversity and the association of viral archetypal regulatory regions with human
brain tumors.
Lednicky JA, Butel JS.
Department of Molecular Virology and Microbiology, Baylor College of Medicine,
Houston, TX 77030, USA.
The regulatory region (RR) of simian virus 40 (SV40) contains enhancer/promoter
elements and an origin of DNA replication. Natural SV40 isolates from simian
brain or kidney tissues typically have an archetypal RR arrangement with a
single 72-basepair enhancer element. A rare simpler, shorter SV40 RR exists that
lacks a duplicated sequence in the G/C-rich region and is termed protoarchetypal.
Occasionally, SV40 strain variants arise de novo that have complex RRs, which
typically contain sequence reiterations, rearrangements, and/or deletions. These
variants replicate faster and to higher titers in tissue culture; we speculate
that such faster-growing variants were selected when laboratory strains of SV40
were initially recovered. SV40 strains with
archetypal RRs have been found in some human brain tumors. The possible
implications of these findings and a brief review of the SV40 RR structure are
presented. Copyright 2001 Academic Press.
Detection of simian virus 40 DNA sequence in
human primary glioblastomas multiforme.
Kouhata T, Fukuyama K, Hagihara N, Tabuchi K.
Department of Neurosurgery, Saga Medical School, Japan.
OBJECT: Deoxyribonucleic acid oncoviruses can induce neoplastic transformation
of cells because their viral proteins interfere with antiproliferative cellular
proteins. Simian virus 40 (SV40) is a DNA virus
that induces the emergence of ependymomas, choroid plexus tumors, mesotheliomas,
osteosarcomas, sarcomas, and various tumors when injected into newborn hamsters.
Recently, approximately 60% of human ependymomas, choroid plexus tumors, and
mesotheliomas were reported to contain and express SV40 DNA sequences. In this
study the presence of SV40 DNA sequences was investigated in human brain tumors.
METHODS: Three of 32 glioblastomas mutiforme (GBMs), but none of two ependymomas
and five medulloblastomas, were found to possess SV40 DNA sequences when
examined using polymerase chain reaction (PCR). The DNA sequence analysis of PCR-amplified
fragments disclosed that the samples were identical to the regulatory region of
SV40. All three GBMs, which arose in elderly patients with wild-type p53, were
considered to be primary (de novo) tumors. Although each of the three tumors was
immunohistochemically negative for SV40 T antigen, in situ hybridization
successfully demonstrated the messenger RNA for SV40 T antigen. CONCLUSIONS:
The results of this study indicate that latent
infection of SV40 in elderly people may be implicated in the tumorigenesis of
certain primary GBMs.
Jasani B, Cristaudo A, Emri SA, Gazdar AF, Gibbs A, Krynska B, Miller C,
Mutti L, Radu C, Tognon M, Procopio A.
Immunocytochemistry and Molecular Pathology Unit, Department of Pathology,
University of Wales College of Medicine, CF14 4XN, Cardiff, UK.
SV40 was discovered as a contaminant of poliovirus vaccines that were
inadvertently administered to millions of people in Europe and the United States
between 1955 and 1963. Shortly afterwards, SV40 was proven to be oncogenic in
rodents and capable of transforming human and animal cells in vitro. The
possibility that SV40 might cause tumours in humans thus became a subject of
scientific and public interest and scrutiny. However, largely due to a lack of
significant epidemiological evidence, interest in assessing SV40's potential
carcinogenic role in humans diminished.
Recently, many laboratories have reported the presence of SV40-like DNA in a
high proportion of human mesotheliomas, ependymomas and osteosarcoma (the three
main types of tumours caused by virus in hamsters), renewing the question
whether SV40 might be a human tumour virus. Molecular data from these studies
are reviewed to re-evaluate the potential role of SV40 as a human carcinogen.
Copyright 2001 Academic Press.
Included in this reference list is some of the epidemiological evidence in favor
of there being a relationship between SV40 and cancer. To the extent to which
there is not more, to what extent is it due to the question not being adequately
studied? (To adequately assess the risk, comparisons between those vaccinated
with polio vaccine, other vaccines, and NO vaccines would have had to have been
made.)
The role of the SV40 ST antigen in cell
growth promotion and transformation.
Rundell K, Parakati R.
Department of Microbiology-Immunology, Northwestern University, and The Robert
H. Lurie Comprehensive Cancer Center, Chicago, IL 60611, USA. krundell@northwestern.edu
The simian virus 40 small-t (ST) antigen plays
a key role in permissive and nonpermissive infections, increasing virus yields
in lytic cycles of primate cells and enhancing the ability of large-T (LT) to
transform rodent or even human cells. In the absence of ST, tumors in
rodent model systems appear primarily in lymphoid and other proliferative
tissues and transformation is reduced in several in vitro systems. The functions
of ST largely reflect its binding and inhibition of protein phosphatase 2A,
although a recently described dnaJ domain also contributes to its biology. The
dnaJ domain is present in LT and a third early gene product, the 17kT protein,
for which a potential role in transformation deserves further evaluation.
Copyright 2001 Academic Press.
Rizzo P, Bocchetta M, Powers A, Foddis R, Stekala E, Pass HI, Carbone M.
Cancer Immunology Program, Cardinal Bernardin Cancer Center, Department of
Pathology, Loyola University Chicago, Maywood, IL 60153, USA.
Malignant mesothelioma, a tumor of the pleura,
pericardium, and peritoneum, is presently a worldwide problem. Current therapy
is ineffective in slowing the course of the disease, and median survival from
the time of diagnosis is rarely greater than 1 year. While the tumor was almost
unknown prior to the second half of the twentieth century, it is presently
responsible for more than 2000 deaths per year in the US alone. Mesothelioma is
frequently associated with exposure to asbestos, but the incidence of cases
involving individuals with low levels of asbestos exposure is increasing. For
this reason, there has been much interest in studying whether there are
alternative factors that act alone or in conjunction with asbestos in producing
this malignancy. In the last decade, simian virus 40 (SV40) has become the most
notable suspected agent. Copyright 2001 Academic Press.
Strategies to circumvent SV40 oncoprotein
expression in malignant pleural mesotheliomas.
Schrump DS, Waheed I.
Thoracic Oncology Section, Surgery Branch, National Cancer Institute, Bethesda,
MD, 20892, USA. schrump@pop.nci.nih.gov
Although nearly 60% of mesotheliomas contain
SV40 early region DNA sequences, the role of T/t antigens in initiating and
maintaining the transformed state of mesothelioma cells remains unclear.
The majority of mesothelioma cells which contain SV40 early region sequences
exhibit extremely low basal expression of SV40 oncoproteins; however, T/t
antigen expression can be induced under conditions of cellular stress.
Abrogation of SV40 T/t expression by antisense techniques induces apoptosis in
part via restoration of p53 function, and enhances chemosensitivity in SV40 (+)
MPM cells by mechanisms which have not been fully elucidated.
This review briefly summarizes our ongoing
efforts to define the role of SV40 oncoproteins in modulating the malignant
phenotype of mesothelioma cells, and highlights strategies which may prove
efficacious in vivo for circumventing SV40 T/t antigen expression in
mesotheliomas. Copyright 2001 Academic Press.
Simian virus 40 detection in human
mesothelioma: reliability and significance of the available molecular evidence.
Jasani B, Jones CJ, Radu C, Wynford-Thomas D, Navabi H, Mason M, Adams M,
Gibbs A.
Department of Pathology, University of Wales College of Medicine, Heath Park,
Cardiff, CF14 4XN, Wales, UK. Jasani@cf.ac.uk
Simian virus 40 was discovered as a contaminant of early poliovirus vaccines
that were inadvertently administered to millions of people in Europe and the
United States between 1955 and 1963. Although
SV40 was proven to be oncogenic in rodents and capable of transforming human and
animal cells in vitro, its role in human cancer could not be proven
epidemiologically. The matter was forgotten until 1993 when SV40 was
accidentally found to cause
mesotheliomas in hamsters injected intra-cardially. Subsequently, DNA
sequences associated with its powerful oncogenic principal, the large T antigen,
were found with high frequency in human pleural mesothelioma using the
polymerase chain reaction (PCR). Since then many laboratories have confirmed the
human findings. However, a few laboratories have failed to reproduce these data
and the authors of the studies have claimed that the detection of SV40 DNA may
simply represent PCR contamination artefacts. The controversy raised by this
viewpoint is reviewed in this article together with a critical appraisal of the
reliability of the molecular techniques used to detect SV40 DNA, in order to
evaluate the potential aetiopathogenic role of SV40 in human mesothelioma.
If SV40
was “proven to be oncogenic in rodents
and capable of transforming human and animal cells in vitro”,
why wasn’t it studied more aggressively? Why did it take an accident to
initiate more interest?
A genetically tractable model of human glioma
formation.
Rich JN, Guo C, McLendon RE, Bigner DD, Wang XF, Counter CM.
Department of Medicine, Duke University Medical Center, Durham, North Carolina
27710, USA. rich0001@mc.duke.edu
Gliomas remain one of the deadliest forms of cancer. Improved therapeutics will
require a better understanding of the molecular nature of these tumors. We,
therefore, mimicked the most common genetic changes found in grade III-IV
gliomas, disruption of the p53 and RB pathways and activation of telomere
maintenance and independence from growth factors,
through the ectopic expression of the SV40
T/t-Ag oncogene, an oncogenic form of H-ras (H-ras(V12G)), and the human
telomerase catalytic subunit hTERT in normal human astrocytes. The resulting
cells displayed many of the hallmarks of grade III-IV gliomas, including greatly
expanded life span and growth in soft agar and, most importantly, were
tumorigenic with pathology consistent with grade III-IV neuroectodermal tumors
in mice. This model system will, for the first time, allow the biological
significance of selected genetic alterations to be studied in human gliomas.
Does SV40 infection contribute to the
development of human cancers?
Shah KV.
Department of Molecular Microbiology and Immunology, Johns Hopkins University
School of Public Health, 615 N. Wolfe Street, Baltimore, MD 21205, USA.
A large number of recent studies have reported
the detection of simian virus 40 (SV40) nucleotide sequences in a number of
unrelated human cancers which include paediatric and adult brain tumours,
pleural mesotheliomas, bronchopulmonary carcinomas and osteosarcomas. Most of
the data have been gathered by the use of PCR assays. These observations imply
that SV40 is circulating in human communities by person-to-person transmission
and that the virus is capable of wide dissemination in the infected individual.
These claims must be regarded with skepticism for a variety of reasons. There
are several inconsistencies in the reported data; for example, there is a wide
variation (0-23%) in the frequency of SV40 sequences in normal tissues. The
results of PCR assays have been difficult to confirm by less error-prone and
independent assays and the relationship of the virus to the tumour cell is not
characterised. The SV40 sequences reported from humans are essentially identical
to those of wild-type SV40; it is difficult to conceive that a highly
species-specific polyomavirus like SV40 would cross the species barrier, become
a human infection, and acquire pathogenic potential for the new host without
significant adaptive change in its genome. The available evidence, although not
extensive, does not indicate that SV40 is circulating in human communities. It
is premature to discuss or speculate on the potential role of SV40 in the
development of human cancer until the presence of SV40 in cancers and in human
communities is established unequivocally, the risk factors for SV40 infection
are elucidated, and the effect of SV40 exposure on the development of specific
cancers is examined in the context of other known risk factors for those cancers.
Copyright 2000 John Wiley & Sons, Ltd.
Low frequency of SV40, JC and BK polyomavirus
sequences in human medulloblastomas, meningiomas and ependymomas.
Weggen S, Bayer TA, von Deimling A, Reifenberger G, von Schweinitz D,
Wiestler OD, Pietsch T.
Department of Neuropathology, University of Bonn Medical Center, Germany.
Several reports have suggested a role for polyomaviruses in the pathogenesis of
human brain tumors. This potential involvement is not conclusively resolved. For
the present study, a highly sensitive PCR-assay with fluorescence-labelled
primers was developed to search for polyomavirus sequences in human brain tumor
and control DNA samples. The assay was shown to detect approximately one viral
large T-antigen (TAg) gene per 250 cells. We identified simian virus 40
(SV40)-like sequences in 2/116 medulloblastomas, in 1/131 meningiomas, in 1/25
ependymomas and in 1/2 subependymomas. A single case of ependymoma contained
SV40 VP-1 late gene sequences. Moreover, one of the meningioma samples showed JC
virus sequences. In contrast, 60 hepatoblastoma samples and 31 brain samples
from schizophrenic patients were consistently negative. BK virus sequences were
not detectable in any of our samples. Immunohistochemical analysis of two SV40
positive tumor biopsies failed to detect large TAg in the tumor cells. In the JC
positive meningioma, immunoreactivity for the viral late gene product (VP-1) was
not observed. Our data do not entirely rule out
SV40 and JC virus as an initiative agent with a hit-and-run mechanism. However
the low frequency of virus sequences and the absence of TAg protein expression
argue against a major role of these viruses in the pathogenesis of human
medulloblastomas, meningiomas and ependymomas.
Overexpression of simian virus 40 small-T
antigen blocks centrosome function and mitotic progression in human fibroblasts.
Gaillard S, Fahrbach KM, Parkati R, Rundell K.
Department of Microbiology-Immunology and the Robert H. Lurie Comprehensive
Cancer Center, Northwestern University, Chicago, Illinois 60611-3010, USA.
Recombinant adenoviruses that express high levels of the simian virus 40 (SV40)
small-t (ST) antigen have been used to study the requirement for ST to drive
cell cycle proliferation of confluent human diploid fibroblasts. This occurs
when either large-T (LT) antigen or serum is added to provide a second signal.
While cells readily completed S phase in these experiments, they were found to
accumulate with 4N DNA content. Cellular and nuclear morphology, as well as the
biochemical status of cyclin B complexes, showed that these cells entered
mitosis but were blocked prior to mitotic metaphase. The defect appears to
reflect an inability of cells overexpressing ST to form organized centrosomes
that duplicate and separate normally during the cell cycle and, therefore, the
absence of a mitotic spindle. The ability of ST to bind protein phosphatase 2A
was required for this pattern, suggesting that altered phosphorylation of key
centrosomal components may occur when ST is overexpressed.
Although the possible significance of ST
effects on the centrosome cycle is not fully understood, these findings suggest
that ST could influence chromosomal instability patterns that are a hallmark of
SV40-transformed cells and LT expression.
[A study of simian virus 40 infection and its
origin in human brain tumors]
[Article in Chinese]
Bu X, Zhang X, Zhang X, et Al.
Department of Neurosurgery, Henan Provincial People's Hospital, Zhengzhou
450003, China.
OBJECTIVE: To study the status of simian virus 40 (SV40) infection and its
origin in human brain tumors. METHODS: Polymerase chain reaction (PCR) and Dot
blot hybridization were used to detect SV40 DNA sequences in 516 human brain
tumor tissues, 80 peripheral blood cells and 50 sperm fluids from healthy
individuals, 100 human embryo tissues from artificial abortion, 30 normal brain
tissues and two human glioma cell lines: SHG44 and BT325. RESULTS:
SV40 DNA sequences were found in 36.4% of human
brain tumors (188/516), 16.3% of healthy peripheral blood cells (13/80), 22.0%
of healthy semen (11/50), 8.0% of human embryo tissues (8/100) and 6.7% of
normal brain tissues (2/30). SV40 DNA sequences were also detected in
SHG44 and BT325 cell lines. The positive rates
of SV40 DNA in human brain tumors, peripheral blood cells and semen from healthy
donors were significantly higher than those in human normal brain and embryo
tissues (P < 0.05). CONCLUSIONS: (1)
SV40 has a higher infection rate in human brain tumors, (2) SV40 is closely
related to the etiopathogenesis of human brain tumors, (3) the ways of SV40
spread may due to both horizontal and diaplacental infections in human
population.
Simian virus 40, poliovirus vaccines, and
human cancer: research progress versus media and public interests.
Butel JS.
Department of Molecular Virology and Microbiology, Baylor College of Medicine,
Houston, TX 77030-3498, USA.
From 1955 through early 1963, millions of people were inadvertently exposed to
simian virus 40 (SV40) as a contaminant of poliovirus vaccines; the virus had
been present in the monkey kidney cultures used to prepare the vaccines and had
escaped detection. SV40 was discovered in 1960 and subsequently eliminated from
poliovirus vaccines. This article reviews current knowledge about SV40 and
considers public responses to reports in the media.
SV40 is a potent tumour virus with broad tissue
tropism that induces tumours in rodents and transforms cultured cells from many
species. It is also an important laboratory model for basic studies of molecular
processes in eukaryotic cells and mechanisms of neoplastic transformation. SV40
neutralizing antibodies have been detected in individuals not exposed to
contaminated poliovirus vaccines. There have been many reports of detection of
SV40 DNA in human tumours, especially mesotheliomas, brain tumours and
osteosarcomas; and DNA sequence analyses have ruled out the possibility that the
viral DNA in tumours was due to laboratory contamination or that the virus had
been misidentified. However, additional studies are necessary to prove
that SV40 is the cause of certain human cancers. A recently published review
article evaluated the status of the field and received much media attention. The
public response emphasized that there is great interest in the possibility of
health risks today from vaccinations received in the past.
Presence and expression of the simian
virus-40 genome in human giant cell tumors of bone.
Gamberi G, Benassi MS, Pompetti F, Ferrari C, Ragazzini P, Sollazzo MR,
Molendini L, Merli M, Magagnoli G, Chiesa F, Gobbi AG, Powers A, Picci P.
Laboratory of Oncologic Research, Rizzoli Institute, Bologna, Italy.
gabriella.gamberi@ior.it
SV40 DNA sequences have been found in human tumors, such as mesotheliomas,
ependymomas, and bone tumors, suggesting that SV40 may be involved in their
etiology. The FOS oncogene could play an important role in bone development
because SV40 is able to induce FOS in cell culture. In this study, the presence
of SV40 sequences, large T antigen (Tag), and FOS protein expression were
investigated in 120 giant cell tumors (GCTs), moderately benign bone tumors that
in some cases can progress to a malignant phenotype. Polymerase chain reaction (PCR),
using primers that amplify the RB1 pocket binding domain and the intron of Tag,
was used to analyze GCT for the presence of SV40 DNA. Tag and FOS protein
expression was evaluated by immunohistochemistry. SV40 sequences were found in
30/107 GCTs, and of these, 22/30 samples expressed Tag protein (73%) and 15/30
overexpressed the FOS oncogene (50%). FOS was undetectable in 77 SV40-negative
GCTs. Sequence analysis of the amplified DNAs confirmed that the amplified
sequences corresponded to SV40 DNA. The correlation between FOS overexpression
and SV40-positive GCTs was highly statistically significant (P < 0.001).
These results show that SV40 DNA sequences and
SV40 Tag are present in GCTs and might induce FOS activity. These data suggest
that SV40 might play a role in the development and progression of some GCTs.
Copyright 2000 Wiley-Liss, Inc.
High incidence of SV40-like sequences
detection in tumour and peripheral blood cells of Japanese osteosarcoma
patients.
Yamamoto H, Nakayama T, Murakami H, Hosaka T, Nakamata T, Tsuboyama T, Oka M,
Nakamura T, Toguchida J.
Institute for Frontier Medical Sciences, and Department of Orthopaedic Surgery,
Faculty of Medicine, Kyoto University, Japan.
Recent studies have revealed the evidence for
the significance of SV40 genome in human malignancies. In this paper, the
presence of SV40-like sequences was investigated in 54 Japanese osteosarcomas in
which mutations of the retinoblastoma (Rb), p53, MDM2, and CDK4 genes had been
already analysed. Using polymerase chain reaction and Southern hybridization,
SV40-like sequences were detected in 25 cases (46.3%). In most cases, only a
part of SV40 genome was detected, and the regulatory region containing enhancer
sequences was most frequently found (21/54, 38.9%). There was no apparent
relationship between the presence of SV40-like sequences and tumour suppressor
genes mutations in each tumour. The SV40-like sequences were also detected in
peripheral blood cells of substantial proportion of the patients (43.3%),
whereas the incidence was much lower (4.7%) in normal healthy controls.
This difference is statistically highly
significant (P < 0.0001), suggesting that the presence of SV40-like sequences,
even if only a part, may play some roles to predispose individuals to
osteosarcoma.
Simian virus 40: the link with human
malignant mesothelioma is well established.
Carbone M, Rizzo P, Pass H.
Cardinal Bernardin Cancer Center, Loyola University Medical Center, Maywood, IL
60153, USA.
Mesotheliomas are malignancies of the pleural, pericardial, and peritoneal
surfaces with a mean survival of less than 1 year from the time of diagnosis
(1). While mesotheliomas were extremely rare in the first half of this century,
the incidence of these tumors has increased enormously in the last several
decades. Presently, 2-3 thousand people in the US develop and die of
mesothelioma each year (1). It is estimated that approximately 80% of
mesotheliomas develop in people with a history of occupational asbestos exposure
or in individuals with family member(s) professionally exposed to asbestos that
brought home fibers on their clothing (1).
Although conventional wisdom dictates that asbestos is the most commonly
associated "environmental" factor with mesothelioma, asbestos does not transform
human mesothelioma cells in tissue culture (2). This suggests that additional
carcinogens act in concert with asbestos to cause mesothelioma. Recent evidence
indicated that Simian Virus 40 (SV40) preferentially causes mesotheliomas in
hamsters, and that SV40 is present in up to 80% of human mesotheliomas in the US
and in Europe (reviewed in ref. 3 and 4).
The nuclear matrix as a target for viral and
cellular oncogenes.
Deppert W.
Heinrich-Pette-Institut fur Experimentelle Virologie und Immunologie an der
Universitat Hamburg, Germany.
As the key integrator of nuclear structure and function, the nuclear matrix is
likely to be an important target for structural and functional alterations
during the process of neoplastic transformation. Here I summarize and discuss
data demonstrating that the major transforming protein of the small DNA tumor
virus simian virus 40 (SV40), the SV40 large tumor antigen (large T),
specifically targets the chromatin and the nuclear matrix during viral
transformation. I then turn to recent evidence endorsing the concept that mutant
p53--the most commonly expressed oncogene in human cancer--might exert its
oncogenic activities by specifically interacting with the nuclear matrix.
The data suggest that SV40 large T and mutant
p53 might be members of a new family of oncogenes that exert their oncogenic
functions by directly modulating nuclear structure and function.
Recent advances in the molecular analysis of
human malignant mesothelioma.
De Rienzo A, Testa JR.
Human Genetics Program, Fox Chase Cancer Center, Philadelphia, PA 19111, USA.
PURPOSE: To evaluate the role of asbestos, somatic genetic alterations, and
simian virus 40 (SV40) in the formation of malignant mesothelioma (MM). DESIGN:
To review recent cytogenetic and molecular genetic advances in MM. RESULTS:
Exposure to asbestos is a major factor contributing to the development of most
MMs. The accumulation of recurrent cytogenetic deletions in most MMs suggests a
multistep process in this malignancy characterized by the loss and/or
inactivation of multiple tumor suppressor genes (TSGs). Karyotypic, comparative
genomic hybridization (CGH), and loss of heterozygosity (LOH) analyses of MMs
have demonstrated frequent deletions of specific chromosomal regions within 1p,
3p, 6q, 9p, 13q, 15q, and 22q. Positional candidate gene approaches have
identified TSGs within two of these regions, i.e., CDKN2A at 9p21 and NF2 at
22q12, which are frequently altered in MMs. In
addition, recent studies have demonstrated the presence and expression of SV40
in many MMs. Proposed mechanisms by which asbestos and SV40 contribute to the
development of MM are reviewed. CONCLUSIONS: The identification of new
TSGs involved in MM and understanding the role of these genes and of SV40 in the
pathogenesis of this malignancy may lead to design of more effective therapeutic
strategies.
Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot,
Israel. lioren@wiccmail.weizmann.ac.il
The p53 protein was discovered 20 years ago, as a cellular protein tightly bound
to the large T oncoprotein of the SV40 DNA tumour virus. Since then, research on
p53 has developed in many exciting and sometimes unexpected directions. p53 is
now known to be the product of a major tumour suppressor gene that is the most
common target for genetic alterations in human cancer. The nonmutated wild-type
p53 protein (wtp53) is often found within cells in a latent state and is
activated in response to various intracellular and extracellular signals.
Activation involves an increase in overall p53 protein levels, as well as
qualitative changes in the protein. Upon activation, wtp53 can induce a variety
of cellular responses, most notable among which are cell cycle arrest and
apoptosis. To a great extent, these effects are mediated by the ability of p53
to activate specific target genes. In addition, the p53 protein itself possesses
biochemical functions which may facilitate DNA repair as well as apoptosis. The
role of p53 in normal development and particularly in carcinogenesis has been
elucidated in depth through the use of mouse model systems. The insights
provided by p53 research over the years are now beginning to be utilized towards
better diagnosis, prognosis and treatment of cancer.
Identification in human brain tumors of DNA
sequences specific for SV40 large T antigen.
Huang H, Reis R, Yonekawa Y, Lopes JM, Kleihues P, Ohgaki H.
International Agency for Research on Cancer, Lyon, France.
Simian virus 40 (SV40) sequences have recently
been identified in a variety of human neoplasms, including mesothelioma,
osteosarcoma, and brain tumors, but significant discrepancies exist regarding
the frequency at which this occurs. The SV40 genome is 70% homologous to
JC and BK, two related polyomaviruses that are highly prevalent in humans and
which may cause in immune-compromised patients progressive multifocal
leukoencephalopathy (PML) and cystitis, respectively. We have established a
specific and sensitive method to identify SV40 sequence in DNA extracted from
histological sections, using PCR followed by Southern hybridization to probes
specific to the large T region. We found SV40 large T antigen sequences in all
brain tumor types investigated. High frequencies were found in low-grade
astrocytomas, anaplastic astrocytomas and secondary glioblastomas derived
thereof (13/22, 59%) while somewhat lower frequencies were found in gemistocytic
astrocytomas (9/28, 32%) and oligodendrogliomas (3/12, 25%). Primary
glioblastomas, giant cell glioblastomas, and gliosarcomas, which clinically
develop de novo, contained SV40 sequences in 11-25% of cases. Presence of viral
DNA was also observed in pediatric brain tumors, including ependymomas (9/16,
56%), choroid plexus papillomas (6/16, 38%), and medulloblastomas (5/17, 29%).
In 8 tumor biopsies with SV40 sequences, the adjacent normal brain tissue was
also analyzed but was devoid of viral DNA in all but one case. BK and JC virus
sequences were rarely detected, the overall frequencies being 3% and 2%,
respectively. It remains to be shown whether
the presence of SV40 contributes significantly to malignant transformation or
whether certain human neoplasms provide a microenvironment that favors viral
replication in humans with latent SV40 infection.
The simian virus 40 small-t and large-T
antigens jointly regulate cell cycle reentry in human fibroblasts.
Porras A, Gaillard S, Rundell K.
Department of Microbiology-Immunology and Robert H. Lurie Comprehensive Cancer
Center, Northwestern University, Chicago, Illinois 60611, USA.
Focus formation in human diploid fibroblasts (HDF cells) is known to require
both the simian virus 40 (SV40) large-T and small-t antigens. Similarly, both
SV40 proteins were required to stimulate confluent, density-arrested HDF cells
to reenter the cell cycle. This study used defective recombinant adenoviruses to
examine the roles of the individual SV40 proteins in altering specific steps in
the cell cycle. Small-t antigen and, to a lesser extent, large-T antigen
increased the level of the S phase cyclin cyclin A but without increasing the
activity of associated cyclin kinases unless the two SV40 proteins were
coexpressed. The absence of kinase activity reflected the presence in
density-arrested cells of high levels of the cyclin-dependent kinase inhibitors
p21(WAF1) and p27(KIP1). We report here that expression of SV40 large-T antigen
reduced levels of p21(WAF1), while expression of small-t antigen was required to
decrease p27(KIP1). The separate effects of
large-T and small-t antigens on these two inhibitors may explain the joint
requirement for the two proteins to drive cell cycle reentry of HDF cells and
ultimately transform these cells.
The influence of SV40 immortalization of
human fibroblasts on p53-dependent radiation responses.
Kohli M, Jorgensen TJ.
Division of Radiation Research, Vincent T. Lombardi Cancer Center, Georgetown
University Medical Center, 3970 Reservoir Road, N.W., Washington, DC 20007-2197,
USA.
The simian virus 40 large tumor antigen (SV40 Tag) has been ascribed many
functions critical to viral propagation, including binding to the mammalian
tumor suppressor p53. Recent studies have demonstrated that SV40-transformed
murine cells have functional p53. The status of p53 in SV40-immortalized human
cells, however, has not been characterized. We have found that in response to
ionizing radiation, p53-dependent p21 transactivation activity is present,
albeit reduced, in SV40-immortalized cells and that this activity can be further
reduced with either dominant negative p53 expression or higher SV40 Tag
expression. Furthermore, overexpression of p53 in SV40-immortalized ataxia-telangiectasia
(A-T) cells restores p53-dependent p21 induction to typical A-T levels. All
SV40-immortalized cell lines exhibited an absence of G1 arrest. Moreover, all
SV40-immortalized cell lines exhibited increased apoptosis relative to primary
cells in response to ionizing radiation,
suggesting that SV40 immortalization results in a unique phenotype with regard
to DNA damage responses. Copyright 1999 Academic Press.
Increased oncogenicity of subclones of SV40
large T-induced neuroectodermal tumor cell lines after loss of large T
expression and concomitant mutation in p53.
Salewski H, Bayer TA, Eidhoff U, Preuss U, Weggen S, Scheidtmann KH.
Institut fur Genetik der Universitat Bonn, Germany.
A model for medulloblastoma-like primitive
neuroectodermal tumors was established in rat using retrovirally transduced SV40
large T antigen (LT) as an inducing agent (O. D. Wiestler et al., Brain
Pathol., 2: 47-59, 1992). A cell line isolated
from such a tumor and clonal derivatives thereof were biologically and
molecularly characterized. In the parental tumor cell line, TZ870, which
had been selected for G418 resistance, virtually all cells expressed LT and
wild-type p53, which were complexed to each other. When plated in soft agar,
these cells grew relatively slowly and formed disperse colonies.
However, when grown without selection pressure,
these cells reproducibly gave rise to LT-negative and G418-sensitive
derivatives, LT-0 cells. Surprisingly, these latter cells exhibited a higher
degree of malignancy both in vitro, growing readily to large colonies in soft
agar, and in vivo, where they gave rise to a rapidly growing malignant tumor.
Clonal selection from TZ870 cells revealed two types of clones: in one type, LT
expression was stably maintained, even without selection pressure, whereas the
other type lost the LT coding sequences. All LT-negative clones exhibited the
same phenotype as the LT-0 cells. Reexpression of LT had no effect.
However, LT no longer formed complexes with
p53, and p53 was metabolically stable, suggesting that it had been mutated.
Sequence analyses and diagnostic restriction digests of the p53 gene revealed
that (a) both the parental LT-transformed cells and their derivatives contained
only one complete p53 allele and (b) all LT-positive clones expressed wild-type
p53, whereas all LT-negative clones expressed a mutant allele with a common
mutation at Cys-174-->Tyr, indicating their clonal origin.
We assume that the loss of LT coding sequences
is the consequence of the p53 mutation, perhaps by inducing genomic instability,
and that both the p53 mutation and additional genetic alterations that accompany
the loss of LT coding sequences might contribute to enhanced malignancy.
Asbestos, chromosomal deletions, and tumor
suppressor gene alterations in human malignant mesothelioma.
Murthy SS, Testa JR.
Program of Molecular Oncology, Fox Chase Cancer Center, Philadelphia,
Pennsylvania 19111, USA.
Exposure to the carcinogen asbestos is considered to be a major factor
contributing to the development of most malignant mesotheliomas (MMs). We
highlight the role of asbestos in MM and summarize cytogenetic and molecular
genetic findings in this malignancy. The accumulation of numerous clonal
chromosomal deletions in most MMs suggests a multistep process of tumorigenesis,
characterized by the loss and/or inactivation of multiple tumor suppressor genes
(TSGs). Cytogenetic and loss of heterozygosity (LOH) analyses of MMs have
demonstrated frequent deletions of specific sites within chromosome arms 1p, 3p,
6q, 9p, 13q, 15q, and 22q. Furthermore, TSGs within two of these regions, i.e.,
p16/CDKN2A-p14ARF at 9p21 and NF2 at 22q12, are frequently altered in MMs.
Homozygous deletion appears to be the major mechanism affecting
p16/CDKN2A-p14ARF, whereas inactivating mutations coupled with allelic loss
occur at the NF2 locus. Finally, recent studies have demonstrated the presence
and expression of simian virus 40 (SV40) in many MMs.
SV40 large T antigen has been shown to
inactivate the TSG products Rb and p53, suggesting the possibility that asbestos
and SV40 could act as cocarcinogens in MM. The frequent occurrence of homozygous
deletions of p16/CDKN2A-p14ARF and the ability of SV40 Tag to bind TSG products
suggest that perturbations of both Rb- and p53-dependent growth-regulatory
pathways are critically involved in the pathogenesis of MM.
The role of the J domain of SV40 large T in
cellular transformation.
DeCaprio JA.
Dana-Farber Cancer Institute, Harvard Medical School, 44 Binney Street, Boston,
Massachusetts 02115, USA.
SV40 large T antigen (TAg)-mediated transformation is dependent on binding to
p53 and the retinoblastoma tumor suppressor protein (pRB) and inactivating their
growth suppressive functions. Transformation minimally requires three regions of
TAg: a C-terminal domain that mediates binding to p53; the LXCXE motif (residues
103-107), necessary for binding to pRB and the related proteins p107 and p130;
and an N-terminal domain (residues 1-82) that contains homology to the J domain
found in cellular DnaJ/Hsp40 molecular chaperone proteins.
We have found that the N-terminal J domain of T
Ag cooperates with the LXCXE motif to inactivate the growth suppressive
functions of the pRB-related proteins. Copyright 1999 The International
Association for Biologicals.
Cancer risk associated with simian virus 40
contaminated polio vaccine.
Fisher SG, Weber L, Carbone M.
Cancer Cause and Prevention Program, Loyola University Medical Center, Maywood,
Illinois 60153, USA.
BACKGROUND: The presence of SV40 in monkey cell
cultures used in the preparation of the polio vaccine from 1955 through 1961 is
well documented. Investigations have consistently demonstrated the oncogenic
behavior of SV40 in animal models. Early epidemiologic studies were inadequate
in demonstrating an increase in cancer incidence associated with contaminated
vaccine. Recently, investigators have provided persuasive evidence that SV40 is
present in human ependymomas, choroid plexus tumors, bone tumors, and
mesotheliomas, however, the etiologic role of the virus in tumorigenesis has not
been established. MATERIALS AND METHODS: Using data from SEER, we
analyzed the incidence of brain tumors, bone tumors, and mesotheliomas from
1973-1993 and the possible relationship of these tumors with the administration
of the SV40 contaminated vaccine. RESULTS: Our
analysis indicates increased rates of ependymomas (37%), osteogenic sarcomas
(26%), other bone tumors (34%) and mesothelioma (90%) among those in the exposed
as compared to the unexposed birth cohort. CONCLUSIONS: These data suggest that
there may be an increased incidence of certain cancers among the 98 million
persons exposed to contaminated polio vaccine in the U.S.; further
investigations are clearly justified.
PMID: 10472327 [PubMed - indexed for MEDLINE]
AN: 99401547
The IOM
cited this article in the references, but said there was “no evidence”.
Expression of the simian virus 40 large tumor
antigen (Tag) and formation of Tag-p53 and Tag-pRb complexes in human brain
tumors.
Zhen HN, Zhang X, Bu XY, Zhang ZW, Huang WJ, Zhang P, Liang JW, Wang XL.
Department of Neurosurgery, Xijing Hospital, The Fourth Military Medical
University, Xi'an, The People's Republic of China.
BACKGROUND: The presence of simian virus 40 (SV40) in human brain tumors remains
a controversial issue. Even if SV40 does exist in brain tumors, the questions of
whether it is associated with brain tumorigenesis and by what mechanisms are
unknown. METHODS: SV40 large tumor antigen (Tag) was investigated by
immunoprecipitation, silver staining, and Western blot analysis in 65 brain
tumor cases and 8 cases of normal brain tissue. Tag-p53 and Tag-pRb complexes
were screened by immunoprecipitation and Western blot analysis in 18 and 15 Tag
positive tumor tissues, respectively. RESULTS: Tag was found in all 8 cases of
ependymoma and 2 cases of choroid plexus papilloma, 90% of pituitary adenoma
cases (9 of 10), 73% of astrocytoma cases (11 of 15), 70% of meningioma cases (7
of 10), 50% of glioblastoma multiforme cases (4 of 8), and 33% of
medulloblastoma cases (2 of 6). Five oligodendroglioma cases, 1 pineocytoma
case, and 8 cases of normal brain tissue were negative for Tag. The Tag-p53
complex was detected in all 18 Tag positive tumors tested and the Tag-pRb
complex was detected in all 15 Tag positive tumors tested. CONCLUSIONS:
SV40 Tag not only is expressed in brain tumors;
it also can form specific complexes with tumor suppressors p53 and pRb. SV40 is
correlated with brain tumorigenesis. The inactivation of p53 and pRb due to the
formation of Tag-p53 and Tag-pRb complexes possibly is a significant mechanism
in the etiopathogenesis of brain tumors. Copyright 1999 American Cancer
Society.
Antisense to SV40 early gene region induces
growth arrest and apoptosis in T-antigen-positive human pleural mesothelioma
cells.
Waheed I, Guo ZS, Chen GA, Weiser TS, Nguyen DM, Schrump DS.
Thoracic Oncology Section, Surgery Branch, National Cancer Institute, NIH,
Bethesda, Maryland 20892, USA.
Although SV40 oncoproteins have been detected
in malignant pleural mesotheliomas (MPMs), their role in the pathogenesis and
clinical behavior of these neoplasms remains controversial. In the
present study, we sought to define the relevance of SV40 T/t antigen expression
in established human mesothelioma cell lines deficient for p16INK4a as well as
ARF expression. SV40 early region sequences were readily detected in genomic DNA
isolated from pleural mesothelioma lines; however, levels of SV40 T/t antigen
expression were highly variable in these cells. An adenoviral vector expressing
an antisense transcript to SV40 early region inhibited T antigen expression and
mediated significant growth inhibition and apoptosis in T-antigen-positive
mesothelioma cells and SV40-transformed COS-7 cells. Abrogation of T/t antigen
expression coincided with enhanced p21/WAF-1 expression, suggesting that
restoration of p53-mediated pathways may have contributed to the growth
inhibition and apoptosis induced by the antisense construct. These effects were
not observed after similar treatment of mesothelioma or lung cancer cells
containing no SV40 DNA sequences. Collectively,
these data suggest that SV40 oncoproteins contribute to the malignant phenotype
of pleural mesotheliomas and indicate that interventions designed to
abrogate their expression may be efficacious in the treatment of individuals
with these neoplasms.
Unique strains of SV40 in commercial
poliovaccines from 1955 not readily identifiable with current testing for SV40
infection.
Rizzo P, Di Resta I, Powers A, Ratner H, Carbone M.
Loyola University Medical Center, Cardinal Bernardin Cancer Center, Department
of Pathology, Maywood, Illinois 60153, USA.
SV40 was first identified as a contaminant of poliovaccines used from 1955 until
1963. Recently, SV40 has been detected in several human tumors. The virus
detected in human tumors often contained only one 72-bp enhancer in the
regulatory region, in contrast to the SV40 originally isolated from
poliovaccines, which contained two 72-bp enhancers. The origin of viruses with
one 72-bp enhancer in humans was unknown, because it was thought that these
viruses were not present in poliovaccines. It was also thought that all
poliovaccine vials produced from 1955 until 1963 had been discarded, thus the
possibility that one 72-bp virions contaminated those vials could not be tested.
We unexpectedly obtained what appear to be the last available vials of
poliovaccine produced in 1955. In these vials, we detected and sequenced SV40
containing only one 72-bp enhancer in the regulatory region. The tissue culture
cytopathic test currently used in the United States to screen oral poliovaccines
was designed to detect rapidly proliferating SV40 virions containing two 72-bp
enhancers. We found that this test is not sensitive enough to detect low amounts
of the slow-replicating SV40 virions containing one 72-bp enhancer. This virus
was easily detected in the same cells by immunostaining and PCR. Twelve current
vials of poliovaccines tested uniformly negative for SV40, suggesting that the
precaution of preparing poliovaccines from kidneys obtained from monkeys bred in
isolated colonies prevented SV40 contamination.
Our data demonstrate that humans were exposed to SV40 viruses with both one
72-bp enhancer and two 72-bp enhancers SV40 through contaminated vaccines. Our
data also suggest that instead of cytopathic tests, immunohistochemical and/or
molecular studies should be used to screen poliovaccines for SV40 to completely
eliminate the risk of occasional contamination.
Simian virus 40 and human tumors: It is time
to study mechanisms.
Carbone M.
Loyola University Chicago, Cancer Immunology Program, Department of Pathology,
Cardinal Bernardin Cancer Center, Maywood, Illinois 60153, USA.
Several studies found simian virus 40 (SV40) in 47% to 83% of human
mesotheliomas. Mesotheliomas are malignant tumors of the pleura and peritoneum,
firmly associated with asbestos exposure. In this issue, Gazdar and colleagues
[inverted question mark]Shivapurkar et al., 1999 found that SV40 is present only
in the malignant cells and not in the surrounding stromal cells. Using the
microdissection technique, they found SV40 in 54% of 93 mesotheliomas of the
epithelial type. The surrounding reactive stromal cells, (20 lung cancers and 14
mesotheliomas of the sarcomatoid/fibrous type) did not contain SV40, confirming
the specificity of their positive findings. Furthermore, SV40 was found in 14%
of 14 non-malignant reactive mesothelial cell proliferations. In 12 cases of
mesothelioma a noninvasive (or in situ) component was also identified. In all
four cases in which SV40 sequences were present in the invasive component,
sequences were also present in the accompanying noninvasive component.
These data suggest that the virus resides in
the mesothelial cells prior to tumor development. The data address the
remaining concerns raised at an International Meeting organized by the NIH, FDA,
and CDC in 1997 to definitively associate SV40 with human mesothelioma.
It is time now to investigate the pathogenic
mechanisms of this association, and if SV40-infected mesothelial cells are more
susceptible to other carcinogens, such as asbestos. Furthermore, we must
investigate the interaction between the host immune system and SV40-infected
mesothelial cells, and study if the immunosuppressive activity of asbestos
interferes with tumor rejection. These studies should lead to a better
understanding of mesothelioma pathogenesis, and possibly to new therapeutic
approaches aimed at interfering with the expression of the SV40 genome and/or at
eliciting a strong immune response against SV40 infected mesothelial cells.
Copyright 1999 Wiley-Liss, Inc.
From the article: “Nevertheless,
the work of Huang et al. suggests that SV40 may now be endemic in the human
population, and that the viral T antigen may be at least a co[factor in the
induction of a wide variety of astrocytic and non-astrocytic brain
tumors…..While the authors are properly cautious in not claiming an etiologic
role for SV40 at this time, the results clearly imply that this widely studied
virus of high oncogenic potential in animal models is more than a mere brain
tumor passenger.”
Gibbs AR, Jasani B, Pepper C, Navabi H, Wynford-Thomas D.
Department of Pathology, Llandough Hospital NHS Trust, Penarth, South Glamorgan,
UK.
To investigate the presence of SV40 DNA sequences in human British mesotheliomas,
PCR analysis using PYV and SV primers which amplify a 172 bp fragment of
SV40LTAg and a 105 bp fragment unique to the SV40LTAg respectively was performed
on archival and frozen tissues. Nine pleural mesotheliomas, nine adenocarcinomas
metastatic to the pleura and three inflammatory disorders of the pleura were
studied. PCR positivity with the SV primer set was restricted to four of the
nine cases of mesothelioma with concordance between paraffin embedded and frozen
tissues. Positivity with the PYV primer set was observed in six mesotheliomas,
two adenocarcinomas and one of the reactive pleurae.
This study indicates that SV40 DNA sequences
are present in a substantial proportion of British mesotheliomas.
Simian virus 40-like DNA sequences in human
papillary thyroid carcinomas.
Pacini F, Vivaldi A, Santoro M, Fedele M, Fusco A, Romei C, Basolo F,
Pinchera A.
Istituto di Endocrinologia, Metodologia Clinica e Medicina del Lavoro, Pisa,
Italia.
Sequences of the SV40 virus, a virus of Asian macaques, have been found in human
tumors, such as pleural mesotheliomas, ependimomas and choroid plexus tumors.
Transgenic mice carrying the SV40 large T gene under the transcriptional control
of the thyroglobulin gene promoter, develop thyroid dedifferentiation and
follicular thyroid cell proliferation, leading to thyroid hyperplasia and
adenocarcinomas. On these bases we investigated the presence of SV40 DNA
sequences in 69 samples of papillary thyroid carcinomas (PTC) and in other
thyroid and non-thyroid carcinomas, as well as in benign thyroid diseases. By
Southern blot and PCR amplification followed by sequence analysis, we found the
presence of SV40-related sequences integrated in the tumoral DNA of three cases
of PTC. At least the 203 bp fragment of the aminoterminus of large T antigen,
the 294 bp fragment of the VP1 gene and the 483 bp entire regulatory region were
present in the tumoral DNA of these patients. SV40 sequences were not found in
tissues other than PTC. Our results demonstrate
that, in addition to previous findings in mesotheliomas and brain tumors, SV40
is somehow linked to papillary thyroid carcinoma. Although our data do not
demonstrate a causative role in the development of PTC, this possibility must be
considered and requires further studies.
Discovery of simian virus 40 (SV40) and its
relationship to poliomyelitis virus vaccines.
Hilleman MR.
Merck Institute for Therapeutic Research, Merck Research Laboratories, West
Point, PA 19486, USA.
Simian Virus 40 (SV40) was discovered in 1959 as a covert contaminant of
poliovirus vaccines prepared using Macacus monkey renal cell cultures. This
inapparent polyoma virus of monkeys was detected using Cercopithecus renal cell
cultures and was eliminated from poliovaccines.
There has been no evidence to implicate SV40 virus of vaccine origin in long- or
short-term consequences in human subjects. Of importance, SV40 virus
provided a new model for basic studies of viral pathogenesis and for cell
transformation and neoplasia. Neoplastic transformation is fixed on the
promiscuous binding of SV40 large T antigen to anti-oncogene cellular protein
elements. SV40 also served as a valuable model for defining the immunology of
virus-induced cancer and in its prevention and cure. Further, it has been a
prime tool for elucidating the molecular details of eukaryotic cell processes.
Numerous techniques now used in molecular biology were pioneered in the SV40
system. The SV40 promoter is commonly used in vector expression constructs and
it has continued to be a model to develop new tools for site-specific
mutagenesis. The virus has been critically important to studies in modern
genetics and in molecular biology.
No
evidence? Notice who did the study? (The IOM showed good sense in not citing
THIS study.)
Detection of authentic SV40 DNA sequences in
human brain and bone tumours.
Butel JS, Jafar S, Stewart AR, Lednicky JA.
Division of Molecular Virology, Baylor College of Medicine, Houston, Texas
77030-3498, USA.
This report summarizes our follow-up studies of SV40 DNA sequences in human
brain tumors of early childhood and our confirmation of the presence of SV40 DNA
in human osteosarcomas. We examined brain tumors and osteosarcoma samples by the
polymerase chain reaction (PCR) using primers from four separated regions of the
SV40 genome. Sequence analysis confirmed that authentic SV40 DNA was present.
The regulatory region of each tumor-associated viral DNA was of archetypal
length (non-duplicated enhancer); sequence variation was noted at the extreme
C-terminus of the large T-antigen (T-ag) genes. Infectious SV40 was recovered
from one brain tumor. We sequenced the entire early genomic region from three
human isolates of SV40 and two laboratory strains originally recovered from
monkeys. The predicted amino acid sequence of the large T-ags showed remarkable
sequence conservation among isolates, except for a small variable region
identified at the C-terminus of the protein. There were no
human-isolate-specific changes detected that could serve to distinguish a human
variant of SV40 nor were any tumor-type-specific viral markers observed.
Based on these data, we conclude that authentic
SV40 is associated with some human brain and bone tumors and that multiple SV40
strains can infect humans.
The biological activities of simian virus 40
large-T antigen and its possible oncogenic effects in humans.
Matker CM, Rizzo P, Pass HI, Di Resta I, Powers A, Mutti L, Kast WM, Carbone
M.
Cardinal Bernardin Cancer Center, Loyola University of Chicago, Maywood, IL
60153, USA.
Simian virus 40 (SV40) is an oncogenic virus
which induces tumors in hamsters and transforms human cells in tissue culture.
Between 1955 and 1963, polio vaccines and adenovaccines were contaminated with
SV40; therefore, millions of people were exposed to this oncogenic virus.
The SV40 proteins responsible for in vivo oncogenesis and in vitro cell
transformation are encoded by the early region of the virus. These proteins are
called T (tumor) antigens (Tags), because animals with tumors induced by SV40
have antibodies against these viral proteins.
Recently, we and other research laboratories have found SV40 in specific types
of human tumors: mesothelioma, ependymoma and choroid plexus tumors,
osteosarcoma and sarcoma. The same tumor types will develop in hamsters which
have been injected systemically with SV40. SV40 causes cell transformation in
tissue culture and tumors in animals, because SV40 Tag binds and inactivates the
cellular tumor suppressor gene products, Rb and p53. We found that SV40 Tag
binds p53 and Rb in human mesotheliomas, possibly contributing to the malignant
phenotype.
S. Maugeri Foundation, IRCCS, Rehabilitation Institute of Veruno/Varallo S.,
Italy.
Deoxyribonucleic acid (DNA) oncoviruses can induce neoplastic transformation by
interfering with proliferative proteins. Simian virus 40 (SV40) has been shown
to induce brain tumors, osteosarcoma, lymphoid tumors and malignant mesothelioma
in hamsters and SV40-like DNA sequences corresponding to the Rb-pocket binding
domain of SV40 T-antigen (Tag) have been detected in the same human tumors.
Since only a small percentage of people exposed to asbestos fibers develop a
malignant mesothelioma, SV40 has been suspected to co-operate with the fibers in
the neoplastic transformation or even to itself induce the onset of malignant
mesothelioma in patients without expositive history.
The mechanism that seems to be involved in the
SV40-induced carcinogenesis process is mediated by interaction of Tag, both with
p53 and Rb proteins, leading to their functional inactivation that is
responsible for the removal of their inhibitory cell cycle effect which
determines the increase of the number of cells entering the G1-S phase.
Up to now the source of SV40 human infections has not yet been completely
identified even though administration from 1957-1965 of SV40 contaminated polio
vaccines is highly suspected. Horizontal infection by sexual transmission has
been also hypothesized. Due to the important public health implications further
investigations are required in order to establish both the source and the
carcinogenetic role of simian virus 40 in humans.
The detection of simian virus 40 in human
tumors by polymerase chain reaction.
Rizzo P, Di Resta I, Powers A, Matker CM, Zhang A, Mutti L, Kast WM, Pass H,
Carbone M.
Loyola University of Chicago, Cardinal Bernardin Cancer Center, Maywood, IL,
USA.
Simian virus (SV) 40 is a deoxyribonucleic acid (DNA) virus that induces
mesotheliomas, ependymomas, bone tumors, and lymphomas in hamsters. In recent
years SV40 sequences have been detected in approximately 60% of mesotheliomas
and ependymomas, in 33% of bone tumors and sarcomas, and in 13% of lymphomas.
Because the amount of human specimens available for molecular studies is usually
minimal, the method most commonly used to demonstrate SV40 in human specimens is
the polymerase chain reaction (PCR). PCR is a highly sensitive and useful
technique. In the PCR reaction, different sets of primers are used for targeting
different regions of DNA. The regions of the SV40 genome targeted by PCR include
the large T-antigen, the small t-antigen, the origin of replication, and viral
protein-1 capsid protein. The use of these
different sets of primers to test human tumor specimens for SV40 produce a
different percentage of positive results. This is because these experiments
revealed that some primers are more specific than others which may also detect
sequences belonging to other DNA papovaviruses. Therefore, the combined use of
different sets of primers is recommended when it is important to distinguish
SV40 from other related papovaviruses such as BK and JC, which can also be
occasionally present in human cells. Furthermore, these experiments demonstrated
that polymerase chain reaction analyses for simian virus 40 can be performed
better and easier when using deoxyribonucleic acid extracted from fresh and/or
frozen tissue. Deoxyribonucleic acid from paraffin embedded specimens should not
be used routinely for simian virus 40 testing because of the high risk of
obtaining false negative results. However, these paraffin derived
deoxyribonucleic acids can be used reliably in molecular laboratories
specialized in these type of analyses. This paper describes the methods
that we have developed to test simian virus 40 in human specimens.
Detection of SV40 sequences in human
mesothelioma.
Griffiths DJ, Nicholson AG, Weiss RA.
Institute of Cancer Research, Chester Beatty Laboratories, London, UK.
The presence of SV40 DNA sequences in British cases of mesothelioma was
investigated using previously reported PCR assays. Three different SV40 primer
sets were used which gave positive detection rates of 100% (SV.for3/SV.rev), 42%
(PYV.for/rev) and 15% (SV.for2/SV.rev). When 26 mesothelioma biopsies were
tested. Negative samples included 22 semen samples from AIDS patients, 10 blood
donor DNAs and the human mesothelioma cell line H-MESO-1.
The problems encountered with these assays and
the interpretation of the results are discussed.
Cardinal Bernardin Cancer Center and Department of Pathology, Loyola University
of Chicago, Maywood, Illinois 60153, USA.
Simian virus 40 (SV40) is a DNA tumour virus
which is highly oncogenic in hamsters. Only specific histologic types of tumours
develop in hamsters injected with SV40, and these are influenced by the route of
virus inoculation. When SV40 is injected systemically to expose most different
cell types to the virus, the animals develop mesotheliomas, osteosarcomas,
sarcomas, and lymphomas within six months. When the virus is injected
subcutaneously, sarcomas at the site of injection develop. If hamsters are
injected intracranially with SV40, they develop ependymomas. These same tumour
types have been found to contain SV40.
Simian virus 40 footprints in normal human
tissues, brain and bone tumours of different histotypes.
Martini F, Lazzarin L, Iaccheri L, Corallini A, Gerosa M, Trabanelli C, Calza
N, Barbanti-Brodano G, Tognon M.
Institute of Histology and General Embryology, University of Ferrara, Italy.
SV40 footprints were investigated by PCR in normal human tissues and tumours of
different histotypes, followed by Southern blot hybridization with a specific
internal oligoprobe for SV40 DNA. Specific SV40
amplification products were detected at high prevalence in primary human brain
tumours: 83% of choroid plexus papillomas, 75% ependymomas, 47% astrocytomas and
37% glioblastomas. SV40 footprints were also revealed in primary bone tumours:
35% osteosarcomas and Ewing's tumours. Positive normal tissue samples ranged
from 45% of sperm fluids to 8% of brain tissue. Normal bone tissue specimens
were SV40 negative. These results indicate that SV40 is associated with human
brain and bone neoplasms, whereas normal bone and brain tissues were either SV40
negative or positive at low grade. SV40 footprints were found in other normal
samples such as PBC, B- and T-lymphocytes and sperm fluids, indicating that SV40
is latent in these cells. Therefore, these cells may be vectors of SV40 in other
host tissues and may spread SV40 infection by blood transfusion and sexual
transmission in the human population.
Evidence for and implications of SV40-like
sequences in human mesotheliomas and osteosarcomas.
Rizzo P, Di Resta I, Stach R, Mutti L, Picci P, Kast WM, Pass HI, Carbone M.
Cardinal Bernardin Cancer Center and Department of Pathology, Loyola University
of Chicago, Maywood, Illinois 60153, USA.
Pleural mesotheliomas and osteosarcomas develop in hamsters injected
intracardially with SV40. Using primers specific for the RB-pocket binding
domain of SV40 we analysed with the polymerase chain reaction frozen specimens
from 48 human mesotheliomas and 145 human bone tumours. We found that 60% of
human mesotheliomas and 33% of human bone tumours contained SV40-like DNA.
Immunostaining, Western blot and RNA in situ hybridization experiments revealed
SV40 Tag expression in human mesotheliomas. Osteosarcomas were not studied for
Tag expression because not enough material was available. Finally, antibodies
anti-Tag were detected in the sera collected from patients with mesothelioma.
These data indicate that SV40, or a closely
related virus, is/are present in human mesothelioma and osteosarcoma.
Correspondence re: F. Martini et al., SV40
early region and large T antigen in human brain tumors, peripheral blood cells,
and sperm fluids from healthy individuals. Cancer Res., 56: 4820-4825, 1996.
Division of Molecular Virology, Baylor College of Medicine, Houston, TX
77030-3498, USA.
Authentic simian virus 40 (SV40) has been detected in association with human
choroid plexus and ependymoma tumors, and SV40-like DNA sequences have been
found in some human osteosarcomas. We report here an analysis of human
osteosarcoma samples for the presence of SV40 DNA using PCR and primers directed
at 4 distinct sites of the SV40 genome, coupled with sequence analysis.
Authentic SV40 DNA sequences were detected in 5 of 10 osteosarcoma tumor
samples. The SV40 regulatory region in each case was identical and of archetypal
length (non-duplicated enhancer), as is usually found in natural isolates of
SV40 from monkeys and in human brain tumors. A section of the gene that encodes
a viral late gene product (VP1) was detected in 5 of 10 tumors and had an exact
match with the known sequence of SV40. Two separated segments of the large
T-antigen (T-ag) gene were found in the same 5 tumors. Analysis of the DNA
sequences encoding the T-ag carboxy terminus revealed sequence variation among
the tumors, as observed previously in viral DNA associated with human brain
tumors. There does not appear to be a preferential association of a T-ag
variable domain sequence with a given tumor type. No sequences from the
regulatory region of human polyomaviruses JCV and BKV were detected in the bone
tumors. We also noted less efficient recovery of SV40 DNA from tumor samples
fixed in paraffin as compared to frozen tumors.
Our results confirm the presence of SV40 DNA in human bone tumors and, based on
the sequence variation observed for the carboxy terminus of the T-ag gene,
suggest that there is not a specific SV40 strain associated with human
osteosarcomas.
Large T antigen coding sequences of two DNA
tumor viruses, BK and SV40, and nonrandom chromosome changes in two glioblastoma
cell lines.
Tognon M, Casalone R, Martini F, De Mattei M, Granata P, Minelli E, Arcuri C,
Collini P, Bocchini V.
Institute of Histology and General Embryology, School of Medicine, University of
Ferrara, Italy.
The T antigen (TAg) coding sequences of two DNA
tumor viruses, BKV and SV40, were detected by Polymerase Chain Reaction (PCR)
amplification followed by Southern-blot hybridization in two human glioblastoma
multiforme derived cell lines. RT-PCR analysis indicated that these two
TAg coding sequences were expressed in both tumor cell lines carrying the viral
early region DNAs. Moreover, analytical polyacrylamide gel electrophoresis
(PAGE) and DNA sequence analyses showed that the amplified PCR products are
indistinguishable from the TAg coding sequences of BKV and SV40 wildtype
strains. Cytogenetic study performed in the two cell lines showed unbalanced
changes, mainly gains of chromosomes 3p, 5, 6, 7, and 19 and losses of
chromosomes 3, 3q, 16, 9p22-->pter, 18, and 20. Excess of chromosomes 6 and 7
are common to the two cell lines. The putative role of the TAg of the two DNA
tumor viruses in transformation and karyotype changes is discussed.
Chromosomal changes and progressive
tumorigenesis of human bronchial epithelial cell lines.
Ohnuki Y, Reddel RR, Bates SE, Lehman TA, Lechner JF, Harris CC.
Huntington Medical Research Institutes, Pasadena, California 91101, USA.
A simian virus 40 (SV40)-transformed human bronchial epithelial cell line,
BEAS-2B, underwent progressive changes, including the development of
tumorigenicity, during extended in vitro passaging. Karyotypic changes occurred
in parallel with the phenotypic changes. For the first 12 passages following
viral transformation, there were random karyotypic changes. Immortalization
occurred between passages 12 and 21, corresponding with the accumulation of four
characteristic abnormal chromosomes-m-1: add(15)(p11.1); m-2: der(8;9)(q10;q10);
m-3: add(16)(p13); and m-4: mar4- and the loss of one homolog of chromosomes 8,
15, 16, 21, and 22. With further passaging (from 21 to 63), the acquisition of
weak tumorigenicity was observed, accompanied by an increased frequency of cells
containing all four common abnormal chromosomes, m-1 through m-4, and missing
one normal homolog of chromosomes 8, 15, 16, and 22. Four tumor cell lines
(B39-TL, B39-TR, B61-T4 and B61-T7) were established from tumors induced by the
injection of these weakly tumorigenic BEAS-2B 39th- and 61st- passage cells into
athymic nude mice. One of the cell lines, B39-TL, is significantly more
tumorigenic than the others. It is notable that B39-TL showed two specific
abnormal chromosomes, del(3p);der(3;15) (q10;q10) and m-6;
der(21)t(3;21)(p14.2;p12) inducing deletion of a short arm of chromosome 3.
Fluorescence in situ hybridization analysis with a probe for protein tyrosine
phosphatase-gamma demonstrated loss of heterozygosity in the 3p14 region. The
development of step-wise karyotypic changes in this in vitro carcinogenesis
model parallels changes documented in several common human cancers.
Short exposures to tunicamycin induce
apoptosis in SV40-transformed but not in normal human fibroblasts.
Carlberg M, Dricu A, Blegen H, Kass GE, Orrenius S, Larsson O.
Department of Tumor Pathology, Karolinska Institutet, Stockholm, Sweden.
When SV40-transformed fibroblasts (line 90VAVI)
were exposed to tunicamycin, an inhibitor of N-linked glycosylation, an
extensive cell death occured compared with untransformed fibroblasts. A
considerable cell loss was obtained within 24 h after tunicamycin addition, and
after 72 h there were hardly any virus-transformed cells alive. A 2-h
pulse treatment with tunicamycin was found to be almost as effective as a
continuous 48-h treatment in killing the cells. Even such a short exposure as 7
min resulted in a drastically decreased cell viability (54%). The morphology of
the dying tunicamycin-treated 90VAVI cells suggested that they were undergoing
apoptosis. This was also supported by the appearance of nuclear condensation, as
assayed by propidium iodide uptake, which was detectable within 2 h after
tunicamycin addition. Furthermore, analysis of DNA from tunicamycin-treated
90VAVI cells by field inversion gel electrophoresis revealed DNA degradation
into 50 kbp fragments within 2 h, and conventional agarose gel electrophoresis
showed 'DNA laddering', indicating internucleosomal DNA cleavage, detectable
after 36 h. Together with the finding that tunicamycin within seconds caused an
elevation of [Ca2+]i, a well documented early feature of apoptosis in many
experimental systems, these results strongly suggest that tunicamycin-induced
cell death in 90VAVI is due to apoptosis. The
short tunicamycin exposure required to trigger cell death in 90VAVI indicates
that the apoptotic process is irreversibly induced soon after its addition.
It seems unlikely that the pool of one or several specific N-linked
glycoproteins could be depleted during such a short period. Instead the overall
accumulation of unglycosylated proteins in ER might contribute to the apoptotic
response in 90VAVI. Tunicamycin also killed and induced DNA degradation in the
breast cancer cell line MDA-231.
Immortalisation of human oesophageal
epithelial cells by a recombinant SV40 adenovirus vector.
Inokuchi S, Handa H, Imai T, Makuuchi H, Kidokoro M, Tohya H, Aizawa S,
Shimamura K, Ueyama Y, Mitomi T, et al.
Department of Critical and Emergency Medicine, Tokai University School of
Medicine, Kanagawa, Japan.
We introduced the origin-defective SV40 early gene into cultured human
oesophageal epithelial cells by infection of a recombinant SV40 adenovirus
vector. The virus-infected cells formed colonies 3-4 weeks after infection in
medium containing fetal calf serum. When the cells derived from
'serum-resistant' colonies were then maintained in the serum-free medium with a
low calcium ion concentration, some of them passed the cell crisis and kept
growing for over 12 months. These cells, regarded as immortalised cells,
resembled the primarily cultured oesophageal epithelial cells in morphology and
had some of their original characteristics. Treatment of the cells with a high
calcium concentration induced phenotypic changes. These cells still responded to
transforming growth factor beta. When the immortalised cells were injected into
severe combined immunodeficient mice, they transiently formed epithelial cysts,
although the typical differentiation pattern of the oesophageal epithelium was
not observed. These cysts regressed within 2 months without development into
tumours. The results indicated that human
oesophageal epithelial cells were reproducibly immortalised by infection with a
recombinant SV40 adenovirus vector at relatively high efficiency. The
immortalised cells should be useful in studies on oesophageal carcinogenesis and
in assessing the cooperative effects with other oncogene products or
carcinogens.
The role of chemical, physical, or viral
exposures and health factors in neurocarcinogenesis: implications for
epidemiologic studies of brain tumors.
Berleur MP, Cordier S.
Institut National de la Sante et Recherche Medicale, Villejuif, France.
This review highlights some future prospects and implications for epidemiologic
research on the etiology of nervous system tumors. It reviews some points
regarding physiology of the nervous system, in connection with mechanisms of
neurocarcinogenesis, and experimental studies in animals. The results of
epidemiologic studies are summarized in the light of the biological and
experimental observations. The following aspects are particularly emphasized: (i)
higher susceptibility of the developing nervous system to neurocarcinogenic
agents (in the fetus and after birth); (ii) possible implications of knowledge
about mechanisms of neurocarcinogenesis regarding crossing of the blood-brain
barrier, activation of oncogenes and inactivation of anti-oncogenes,
relationship between chemical structure and neurocarcinogenic action; (iii)
necessity of further investigation concerning the occurrence of nitrosoureas and
their precursors in the environment, and the potential role of nitroso compounds
in the development of human brain tumors; (iv) lack of information about
promoting or inhibiting neurocarcinogenic effects, and co-carcinogenesis--among
others, interaction between X-irradiation and exposure to neurocarcinogenic
nitrosoureas; (v) need for studying the
potential neurocarcinogenic risk of polyomaviruses BKV, JCV, and SV40 to humans.
Regulation of the cell cycle by viral
oncoproteins.
Vousden KH.
Ludwig Institute for Cancer Research, St Mary's Hospital Medical School, London,
UK.
Human papillomavirus (HPVs) adenovirus and simian virus 40 (SV40) are small DNA
viruses which can show oncogenic activity. Although not otherwise related, all
three have adopted very similar strategies to deregulate cell growth; each virus
encoding oncoproteins which interact with the same cellular targets. Of
particular interest are the interactions with the cell encoded pRB and p53
proteins, products of tumour suppressor genes. Somatic mutation results in the
loss of the pRB and p53 function in many cancers and the contribution of the
viruses to tumour development appears to reflect their ability to inactivate
these cellular proteins. Both pRB and p53 negatively regulate progress through
the cell cycle and the action of the viral proteins has highlighted the central
importance of these tumour suppressor proteins in maintaining normal cell
growth.
Regulation by tumour antigens defines a role
for PP2A in signal transduction.
Mumby M.
Department of Pharmacology, University of Texas Southwestern Medical Center,
Dallas 75235-9041, USA.
Research conducted over the last decade has provided a wealth of new information
on the molecular mechanisms utilized by DNA tumour viruses. Studies of tumour
viruses have led to important insights into the functions of viral proteins and
into the regulation of normal cellular proliferation. DNA tumour viruses can
stimulate growth factor signaling pathways, alter gene transcription, and
inactivate growth suppressor proteins. Members of the polyoma and adenovirus
families express proteins that interact with protein serine/threonine
phosphatase 2A (PP2A). In the case of SV40
virus, this interaction plays an accessory role in transformation of most cells,
while it is essential for transformation of some cell types. The topics
discussed in this review include the interactions of these viral proteins with
PP2A, the effects of these interactions on phosphatase activity, and how these
interactions alter cellular signal transduction pathways.
Carcinogenesis and related cell and tissue
responses to asbestos: a review.
Mossman BT.
A review of the literature reveals that chrysotile asbestos has a mulitiplicity
of effects on cells and tissues which can provide a framework for assessment of
its role(s) in initiation, promotion, or as a co-carcinogen which acts in
concert with chemical carcinogens found in cigarette smoke in the development of
lung cancer. Several caveats important in the interpretation of these data
include the general lack of dose-response protocols both for in vivo and for in
vitro studies as well as the absence, in many studies, of minerals which are
appropriate positive or negative controls based on epidemiological data in man.
Other factors which may account for disparities in results between studies
include the use of different preparations of chrysotile fibres with distinct
chemical and physical compositions and different cell types and species. Whether
chrysotile is an initiator of lung cancer or mesothelioma in human cells is
unclear, as evidence of chromosomal aberrations in human bronchial epithelial
cells are for the most part negative (Kodoma et al., 1993; see Mossman, B., this
Workshop): only one study employing pleural mesothelial cells from four
individuals, two of whom exhibited chromosomal abnormalities before exposure to
asbestos, has documented chromosomal changes with chrysotile, crocidolite and
amosite asbestos, and this study did not use a non-pathogenic dust as a negative
control. Studies using human lymphocytes show chromosomal changes after exposure
to latex beads or chrysotile at equal weight concentrations (Korkina et al.,
1992; see Mossman, B., this Workshop). Lastly,
although both chrysotile and crocidolite asbestos demonstrated dose-dependent
increase in aberrant anaphases in an SV40 T antigen-transformed human
mesothelial cell line (Pelin et al., 1992, see Mossman, B., this Workshop),
erionite, a potent mesotheliomagenic fibre in rodents and humans, caused no
aberrations in this bioassay. Several studies have been performed to
determine the interactions of chrysotile with rodent cells or isolated bacterial
DNA. Results in a number of bioassays have been positive, but chrysotile is less
potent on a fibre number comparative basis than crocidolite and
no-observed-effect-levels (NOELs) have been observed in several systems. Cell
proliferation by asbestos may be a more relevant phenomenon to tumour
development and promotion, and the ability of chrysotile to stimulate cell
proliferation, using a number of biomarkers, has been demonstrated both in vitro
and after inhalation by rats.(ABSTRACT TRUNCATED AT 400 WORDS)
The p53 tumor suppressor gene as a common
cellular target in human carcinogenesis.
Chang F, Syrjanen S, Kurvinen K, Syrjanen K.
Department of Pathology, Kuopio Cancer Research Centre, University of Kuopio,
Finland.
The p53 gene is a 16-20 kb of cellular DNA located on the short arm of human
chromosome 17 at position 17p13.1. This gene encodes a 375-amino acid nuclear
phosphoprotein which involves in the regulation of cell proliferation. The p53
gene was originally regarded as a dominant oncogene because its overexpression
resulted in the immortalization of rodent cells, and the p53 gene could
transform rat embryonic fibroblasts in concert with an activated ras gene. It
soon became clear, however, that many of the p53 clones that had been studied
were in fact mutated versions of the gene, and the wild-type p53 actually acts
as a tumor suppressor. Loss of normal p53 function has been associated with the
cell transformation in vitro and the development of neoplasms in vivo. More than
one-half of human malignancies derived from the epithelial, mesenchymal,
hematopoietic, and lymphoid tissues, as well as the central nervous system,
analyzed thus far, were shown to contain an altered p53 gene. Most p53 gene
alterations are the missense mutations, giving rise to an altered protein. These
mutations are most frequently located in the evolutionally conserved areas.
Furthermore, it has been demonstrated that the
SV40 large T antigen, the adenovirus E1B protein, and papillomavirus E6 protein
can bind to wild-type p53 protein and presumably lead to inactivation of this
gene product as well. Therefore, the inactivation of normal (or wild-type) p53
is currently regarded as an important genetic pathway for human carcinogenesis
generated by endogenous factors and exogenous carcinogens, as well as several
tumor viruses. The current data on the p53 gene and its alterations in
human malignancies, particularly those in the gastrointestinal tract, are
reviewed.
p53 mutations in human immortalized
epithelial cell lines.
Lehman TA, Modali R, Boukamp P, Stanek J, Bennett WP, Welsh JA, Metcalf RA,
Stampfer MR, Fusenig N, Rogan EM, et al.
Laboratory of Human Carcinogenesis, National Cancer Institute, Bethesda, MD
20892.
Although rodent cells have been immortalized following transfection with a
mutant p53 gene, the role of p53 in the immortalization of human cells is
unknown. Therefore, human epithelial cell lines were examined for p53 mutations
in exons 4-9 which include the evolutionarily conserved regions. A spontaneously
immortalized skin keratinocyte cell line, HaCat, and three ras-transfected
clones, have a p53 mutational spectrum that is typical of ultraviolet light
induced mutations. A normal finite lifespan cell strain (184) and two
benzo[a]pyrene immortalized mammary epithelial cell lines derived from 184
(184A1 and 184B5) contain wild type p53 sequences in exons 4-9, although
elevated levels of nuclear p53 indicate an alteration in the stability of the
normally transient protein. Wild type p53 was found in human bronchial,
esophageal and hepatic epithelial cells immortalized by SV40 T antigen gene and
human renal epithelial cells immortalized by adenovirus 5. BEAS-2B, an SV40 T
antigen immortalized bronchial epithelial cell line and two subclones, have a
germline polymorphism at codon 47. Inactivation
of p53 by mechanisms such as mutation or complexing with proteins of DNA tumor
viruses appears to be important in the immortalization of human epithelial cells.
Tumourigenesis associated with the p53 tumour
suppressor gene.
Chang F, Syrjanen S, Tervahauta A, Syrjanen K.
Department of Pathology, University of Kuopio, Finland.
The p53 gene is contained within 16-20 kb of cellular DNA located on the short
arm of human chromosome 17 at position 17p13.1. This gene encodes a
393-amino-acid nuclear phosphoprotein involved in the regulation of cell
proliferation. Current evidence suggests that loss of normal p53 function is
associated with cell transformation in vitro and development of neoplasms in
vivo. More than 50% of human malignancies of
epithelial, mesenchymal, haematopoietic, lymphoid, and central nervous system
origin analysed thus far, were shown to contain an altered p53 gene. The
oncoproteins derived from several tumour viruses, including the SV40 large T
antigen, the adenovirus E1B protein and papillomavirus E6 protein, as well as
specific cellular gene products, e.g. murine double minute-2 (MDM2), were found
to bind to the wild-type p53 protein and presumably lead to inactivation of this
gene product. Therefore, the inactivation of p53 tumour suppressor gene is
currently regarded as an almost universal step in the development of human
cancers. The current data on p53-associated tumourigenesis are briefly
discussed in this minireview.
Selective expression of foreign genes in
glioma cells: use of the mouse myelin basic protein gene promoter to direct
toxic gene expression.
Miyao Y, Shimizu K, Moriuchi S, Yamada M, Nakahira K, Nakajima K, Nakao J,
Kuriyama S, Tsujii T, Mikoshiba K, et al.
Department of Neurosurgery, Osaka University Medical School, Japan.
We have previously demonstrated that retrovirus-mediated genes were transferred
to mouse glioma cells in a meningeal gliomatosis model (Yamada et al.: Japanese
Journal of Cancer Research 83:1244-1247, 1992). This retrovirus vector contains
the Escherichia coli. beta-galactosidase (beta-gal) gene as a marker for
integration of the lacZ gene, which is
controlled by the SV40 early promoter. We investigated whether lacZ genes
could be specifically controlled in mouse glioma cells by glial-specific
promoters, including the 2.5 kb 5' flanking region of the mouse glial fibrillary
acidic protein (GFAP) gene, the 1.3 kb 5' flanking region of the myelin basic
protein (MBP) gene, and the 1.5 kb 5' flanking region of the myelin proteolipid
protein (PLP) gene. Psi-2 packaging cells were transfected with each retrovirus
vector (GFAP promoter-, MBP promoter-, and PLP promoter-lacZ) and the infectious
virus particles were recovered from the supernatants. Blue staining for beta-gal
was detected in various fibroblast, myeloma, and glioma cell lines transduced
with the retrovirus BAG vector. On the other hand, blue staining was only
detected in glioma cells after transduction with the lacZ gene-bearing
retrovirus controlled by glial-specific promoters. The strongest promoter
activity was detected after transduction with the retrovirus in which the MBP
promoter controlled the lacZ gene. Mouse glioma cells transduced with retrovirus
containing the MBP promoter directing the herpes simplex virus type 1 thymidine
kinase (HTK) gene were extremely sensitive to ganciclovir, while the parental
cells and cells transduced with retrovirus containing the lacZ gene were not
sensitive to ganciclovir.(ABSTRACT TRUNCATED AT 250 WORDS)
Why are human cells resistant to malignant
cell transformation in vitro?
Kuroki T, Huh NH.
Department of Cancer Cell Research, University of Tokyo.
Transformation of human cells, both induced and spontaneous, is an extremely
rare event, whereas rodent cells are relatively easily transformed when treated
with a single carcinogenic agent. The present review addresses the question of
why human cells are resistant to malignant transformation in vitro. To
facilitate understanding of the problem, the process of transformation is
divided operationally into two phases, i.e. phase I, immortalization; and phase
II, malignant transformation. In human cells, one-phase transformation, i.e.,
the consecutive occurrence of phases I and II due to the action of a single
carcinogenic agent, is observed only rarely. Once human cells are immortalized,
however, malignant transformation by chemical carcinogens or oncogenes proceeds,
suggesting that for human cells, phase I immortalization is a prerequisite for
such transformation to take place. To date, about 20 papers have been published
describing protocols for the two-phase transformation of a variety of human
epithelial cells and fibroblasts. In most
experiments, SV40, human papilloma viruses and their transforming genes are
utilized for induction of phase I (immortalization) followed by the use of
chemical carcinogens or activated oncogenes for induction of phase II (malignant
transformation). Possible mechanisms that would render human cells
refractory to transformation are discussed below.
SV40 large T antigen trans-activates the long
terminal repeats of a large family of human endogenous retrovirus-like
sequences.
Feuchter AE, Mager DL.
Terry Fox Laboratory, B. C. Cancer Agency, Vancouver, Canada.
The Simian Virus 40 (SV40) large T antigen (T)
is required for the initiation of viral replication, the autoregulation of early
gene expression, and the activation of late gene expression in productively
infected cells. In addition to these roles, T has been implicated in the
transcriptional activation of a variety of viral and cellular promoters.
We have used the chloramphenicol acetyltransferase (CAT) reporter gene system to
study the effect of T on the long terminal repeats (LTRs) of a large family of
human endogenous retrovirus-like sequences, RTVL-H. Here we show that T can
activate expression from certain RTVL-H LTRs 5- to 30-fold. Competition
experiments in which an excess of plasmid containing only an RTVL-H LTR was
cotransfected with an LTR-CAT reporter gene construct confirmed that this effect
is specific for RTVL-H sequences. Restriction enzyme analysis using methylation-sensitive
enzymes has shown that this activation is not due to plasmid replication. We
have also observed this trans-activation effect in two CV-1 cells lines
containing stably integrated LTR-CAT constructs.
These results demonstrate that a known
transforming protein can alter the transcriptional capabilities of RTVL-H LTRs.
As there are approximately 3000 related LTRs in the genomes of humans and other
primates, these findings suggest that a large number of these promoters and
their associated transcripts may be transcriptionally stimulated by this and
other oncogens.
Department of Biological Sciences, Columbia University, New York 10027.
The p53 protein is an important determinant in human cancer and regulates the
growth of cells in culture. It is known to be a sequence-specific DNA-binding
protein with a powerful activation domain, but it has not been established
whether it regulates transcription directly. Here we show that intact purified
wild-type human and murine p53 proteins strongly activate transcription in
vitro. This activation depends on the ability of p53 to bind to a template
bearing a p53-binding sequence. By contrast, tumour-derived mutant p53 proteins
cannot activate transcription from the template at all, and when complexed to
wild-type p53, these mutants block transcriptional activation by the wild-type
protein. Moreover, the simian virus 40 large T
antigen inhibits wild-type p53 from activating transcription. Our results
support a model in which p53 directly activates transcription but this activity
can be inhibited by mutant p53 and SV40 large T antigen through interaction with
wild-type p53.
Switch to the angiogenic phenotype during
tumorigenesis.
Folkman J, Hanahan D.
Department of Surgery, Children's Hospital, Boston, Massachusetts.
Tumor growth and metastasis are angiogenesis-dependent. Virtually all solid
tumors are neovascularized by the time they are detected. However, there is a
prevascular phase during early tumor development where few or no tumor cells are
angiogenic and expansion of the tumor is restricted to a few mm3. When enough
tumor cells become angiogenic, the tumor can expand progressively and shed
metastatic cells. This angiogenic switch has recently been quantitated for human
breast cancer, as well as for prostate cancer.
We have studied the problem of how tumors switch to the angiogenic phenotype by
using transgenic mice in which tumors develop at a predictable time and in
discrete prevascular and vascular stages. When the transgene is the
bovine papilloma virus (BPV) genome, angiogenic fibrosarcomas develop from non-angiogenic
precursors called fibromatoses. The fibrosarcomas secrete growth factors for
capillary endothelial cells. In contrast, the fibromatoses do not secrete
endothelial cell growth factors. When the
transgene consists of the large "T" antigen of SV40 under the control of the rat
insulin promoter, 70% of pancreatic islets become hyperplastic and 4-10% of
these become angiogenic at 6-7 weeks. Tumors arise from these
neovascularized hyperplastic islets and reach > 1000 x the volume of the
preangiogenic islets. The onset of angiogenic activity coincides with the
secretion of acidic fibroblast growth factor (aFGF) and other growth factors not
fully identified at this writing. These studies help to explain the switch to
the angiogenic phenotype during tumorigenesis and provide models to discover
antiangiogenic therapies directed at the source of angiogenic activity. Such
therapy, when developed, may be co-administered with currently available
angiogenesis inhibitors which are directed at the target of angiogenic activity,
vascular endothelial cells.
Department of Biology, Lewis Thomas Laboratory, Princeton University, New Jersey
08544-1014.
The retinoblastoma sensitivity protein (Rb) and the p53 gene product both appear
to function as negative regulators of cell division or abnormal cellular growth
in some differentiated cell types. Several types of cancers have been shown to
be derived from cells that have extensively mutated both alleles of one or both
of these genes, resulting in a loss-of-function mutation. In the case of the p53
gene, this mutational process appears to occur in two steps, with the first
mutation at the p53 locus resulting in a trans-dominant phenotype. The mutant
p53 gene product enters into an oligomeric protein complex with the wild-type
p53 protein derived from the other normal allele and such a complex is inactive
or less efficient in its negative regulation of growth control. This
intermediate stage of carcinogenesis selects for the proliferation of cells with
one mutant allele, enhancing the probability of obtaining a cancer cell with
both alleles damaged. The DNA tumor viruses have evolved mechanisms to interact
with the Rb and p53 negative regulators of cellular growth in order to enhance
their own replication in growing cells. SV40
and adenovirus type 5 produce viral encoded proteins that also form oligomeric
protein complexes with p53 and Rb, presumably inactivating their functions.
These viral proteins are also the oncogene products of these viruses. Thus, the
mechanisms by which cancer may arise in a host, via mutations or virus
infections, have fundamental common pathways effecting the same cellular genes
and gene products; Rb and p53.
Department of Pathology, University of California, San Diego, La Jolla 92093.
It is clear that the RB-deficient tumor cells
lost their tumorigenicity in nude mice after regaining the RB gene expression.
However, the mechanism of tumor suppression by the RB gene is still unknown.
More studies on the biological activities of RB protein, pp110RB, are necessary
to answer this question. Recent studies have
shown that several oncogenic viral proteins, such as SV40 large T antigen (47)
and adenoviral E1A protein (48), bind to RB protein. The significance of these
bindings remains unclear; nevertheless, they suggest that depletion of
functional RB protein by viral proteins may provide another mechanism of RB
inactivation. Continued study of naturally occurring as well as
engineered RB mutants may give us some information on the biological activity of
RB protein, and its roles in oncogenesis, differentiation, development and gene
regulation. Additionally, direct detection of RB gene mutations would have great
clinical utility. Probes for the RB gene and gene product will be useful for
genetic diagnosis of cancer susceptibility in affected families. Therefore,
antibodies to the RB protein will be excellent tools for diagnostic and/or
prognostic application in clinical medicine.
Cell alterations induced by the large
T-antigens of SV40 and polyoma virus.
Dilworth SM.
Department of Chemical Pathology, Royal Postgraduate Medical School, London, UK.
The large T-antigens of SV40 and polyoma virus
are nuclear, multifunctional proteins that are essential for replication of the
respective viruses. They can also 'transform' cells in culture to varying
extents; both can immortalise primary cells, and SV40 large T can additionally
induce full transformation. Recently, p105Rb, the protein product of the
anti-oncogene RB-1, has been shown to interact with both large T-antigens. SV40
large T-antigen also binds to a p105Rb related protein, p107.
SV40 large T differs from that of polyoma virus
in its ability to associate with another anti-oncogene product, p53. The
significance of these properties to the transforming potential of both viruses
is considered.
Imperial Cancer Research Fund Laboratory of Molecular Genetics, Institute of
Child Health, London, UK.
A novel class of oncogene has been recognised
whose loss-of-function results in the expression of the malignant phenotype. Two
examples of such genes are the human retinoblastoma predisposition gene (RB1)
and the gene encoding the cellular protein p53. These genes are thought to
regulate and limit normal proliferation of cells and, as a consequence, can
suppress tumorigenicity when introduced into transformed cells. They are hence
frequently described as 'tumour suppressor genes'. Both RB1 and p53 gene
products are bound by various transforming early proteins encoded by the DNA
tumour viruses SV40, adenovirus and human papilloma virus. It is thought that
they are thus sequestered and rendered inactive. Thus, a coherent model is
emerging whereby inactivation, either by mutation of sequestration, of these
tumour suppressor genes may contribute to natural and experimental carcinogenic
processes.
Characterization of hamster tumors induced by
simian virus 40 small t deletion mutants as true histiocytic lymphomas.
Carbone M, Lewis AM Jr, Matthews BJ, Levine AS, Dixon K.
Viral Pathogenesis Section, National Institute of Allergy and Infectious
Diseases, Bethesda, Maryland 20892.
Random bred Syrian hamsters given s.c. injections of SV40 small t deletion
mutants dl883, dl884, and dl890 rapidly develop reticulum cell sarcomas in the
abdominal cavity in addition to slowly developing s.c. fibrosarcomas at the site
of virus inoculation. Injection of wild type SV40 s.c. induces only
fibrosarcomas at the site of inoculation. In an attempt to understand why
mutations in the SV40 small t gene should lead to this difference in
tumor-inducing capacity in hamsters, we studied cells from 12 abdominal
reticulum cell sarcomas which were induced by the s.c. injection of SV40
mutants. Morphological and functional analyses indicate that these tumor cells
are derived from MAC-2+ macrophages. They are highly granulated, vacuolated, and
multinucleated, and they generally adhere to glass and plastic. In addition,
they (a) phagocytose latex beads; (b) express high levels of class II major
histocompatibility complex antigens; (c) contain beta-glucuronidase, acid
phosphatase, and fluoride-inhibited nonspecific esterase; (d) contain lysozyme
and fibronectin; and (e) express cell surface MAC-2 antigens.
Thus, the small t deletions in the SV40 genome
appear to permit the virus to transform cells that are distant from the site of
virus inoculation; at this distant site, the cells transformed are of a
specific lineage, MAC-2+ peritoneal macrophages. This specific tropism may
reflect a unique characteristic of MAC-2+ cells or their precursors that renders
these cells susceptible to SV40 mutants which are otherwise restricted in the
range of cells that they can transform.
Establishment of a human in vitro mesothelial
cell model system for investigating mechanisms of asbestos-induced mesothelioma.
Ke Y, Reddel RR, Gerwin BI, Reddel HK, Somers AN, McMenamin MG, LaVeck MA,
Stahel RA, Lechner JF, Harris CC.
Division of Cancer Etiology, National Cancer Institute, Bethesda, Maryland
20892.
Normal human mesothelial (NHM) cells were transfected with a plasmid containing
SV40 early region DNA. Individual colonies of transformed cells from several
donors were subcultured for periods of 5 to 6 months and 60 to 70 population
doublings (PDs) before senescence, in contrast to a culture lifespan of
approximately 1 month and 15 PDs for NHM cells. One such culture, designated
MeT-5A, escaped senescence and has been passaged continuously for more than 2
years. These cells had a single integrated copy of SV40 early region DNA in
their genome, expressed SV40 large T antigen, and exhibited features of
mesothelial cells including sensitivity to the cytotoxic effects of asbestos
fibers. One year after injection subcutaneously or intraperitoneally in athymic
nude mice, these cells remain nontumorigenic, and therefore are a potential
model system for in vitro fiber carcinogenesis studies.
Does the product of the RB-1 locus have a
cell cycle regulatory function?
Livingston DM, DeCaprio JA, Ludlow JW.
Dana-Farber Cancer Institute, Boston, Massachusetts 02115.
The product of the retinoblastoma (RB) susceptibility locus suppresses
neoplastic growth and exists as a family of differentially phosphorylated, DNA
binding proteins. The unphosphorylated member(s) of this group can interact
specifically with the transforming product of the SV40 genome, large T antigen
(T). The genetics of this interaction further suggest that T-RB complex
formation is an important step in the mechanism of T function as a transforming
element and that T may operate in this regard by perturbing one or more aspects
of the RB growth suppression function. Results of various analyses of the
details of complex formation suggest at least one generic function for RB,
contributing to the regulation of the G1/S transition in the mammalian cell
cycle.
Association of polyomaviruses JC, SV40, and
BK with human brain tumors.
Dorries K, Loeber G, Meixensberger J.
Human brain tumors of 11 different types were analyzed by Southern blot analysis
for the presence of JCV, SV40, and BKV. In 21
tumor specimens examined with JCV- and SV40-specific probes no positive
hybridizations were obtained. Analysis for BKV DNA, however, revealed the
presence of BKV-specific sequences in 11 of 24 tumor specimens. No hybridization
was found in DNA from CNS tissues from different areas of 29 individuals without
CNS tumors. The BKV DNA sequences were associated with high molecular weight
cellular DNA, suggesting a chromosomal location. These data provide evidence for
the involvement of BKV in the development of human brain tumors.
Antibodies to simian vacuolating virus 40 in
bladder cancer patients.
Bravo MP, del Rey-Calero J, Olivares I, Vizcaino MJ.
Department of Preventive Medicine, School of Medicine, Autonomous University of
Madrid, Spain.
The occurrence of antibodies to herpes simplex virus type 2 (HSV-2) and simian
vacuolating virus 40 (SV40) (two presumptive oncogenic viruses) was investigated
by the technique of enzyme-linked immunosorbent assay in 233 patients with
bladder cancer (200 males and 33 females) and in 466 controls (400 males and 66
females). The age of both the bladder cancer patients and controls varied from
55 to 65 years. A statistically significant
association between bladder cancer and antibodies to SV40 was found in both
males and females. No association between HSV-2 and bladder cancer was
observed.
Simian virus 40 (SV40) is a small, DNA-containing tumour virus. One of its gene
products, the large tumour antigen (T-ag), is essential for both viral
replication and cell transformation. SV40 T-ag can be considered a dual oncogene
protein; it is a composite transforming protein that provides distinct functions
at different subcellular locations. In addition to its roles in virus
replication, T-ag exerts numerous effects on host cells. Those cellular effects
reflect viral stimulation of host cell entry into S phase. Numerous chemical
modifications have been ascribed to T-ag. They might be involved in defining
subpopulations of T-ag that are, in turn, responsible for mediating various T-ag
biochemical functions. The T-ag polypeptide, 90,000-100,000 in molecular weight,
appears to contain multiple, discrete functional domains; several biological
activities have been assigned to relatively small defined regions of the
molecule. The cellular progenitors of the T-ag biochemical activities are not
obvious. A cellular protein, p53, thought to be involved in regulation of cell
proliferation, becomes complexed with T-ag in transformed cells and is
stabilized. The interaction of T-ag with this cellular substrate may play an
important part in SV40 transformation. T-ag and T-ag/p53 complexes are localized
in both the nucleus and plasma membrane of transformed cells. T-ag is
transported to the nucleus because of a 7-residue nuclear transport signal
contained within its primary sequence. Its migration to the membrane is by an
unknown pathway. Only a minor fraction of the total cellular T-ag is present at
the cell surface. Both amino and carboxy termini of the T-ag polypeptide are
exposed on the extracellular face of the cell. Nuclear and membrane T-ag are
structurally very similar, although a portion of membrane T-ag is acylated and
nuclear T-ag is not. The nuclear and membrane forms of T-ag apparently provide
separate and complementary functions necessary for cell transformation. Nuclear
T-ag is important in immortalizing primary cells and membrane T-ag may mediate
more pronounced morphological changes. A model is presented, postulating how the
two forms of T-ag might cooperate to mediate phenotypic transformation.
From two human brain tumors SV40 genomes were
isolated by recombinant DNA techniques. The SV40 genome cloned from a human
meningioma DNA was shown to be indistinguishable from wild-type SV40. In
contrast, the SV40 genome cloned from a human astrocytoma proved to be a
nonviable deletion mutant with a truncated early region removing most of the
large T-coding region. In addition, this mutant also carries a tandem
duplication of an intact origin of replication.
Angiosarcomas in hamsters after inoculation
of brain tissue from a case of progressive multifocal leukoencephalopathy.
Brun A, Jonsson N.
Subcutaneous inoculation of minced brain tissue
from a case of progressive multifocal leukoencephalopathy into newborn Syrian
hamsters produced transplantable angiosarcomas in 2 of 11 animals.
Immunostaining revealed the presence of SV40 in the progressive multifocal
leukoencephalopathy (PML) tissue, indicating this virus as the tumor-inducing
agent.
Investigations on the presence of papova
virus in certain forms of human cancer. Note 2. Brain tumors.
Stoian M, Suru M, Hozoc M, Iosipenco M, Athanasiu P, Nastac E, Arseni C,
Alexianu D.
Two serially transmissible tumors (metastatic lymphosarcomas) were induced in
the hamster by inoculation of human brain tumor material--tissue homogenate from
a malignized fibroma or DNA extracted from a meningioma.
SV40 V antigen was made evident by indirect
immunofluorescence in 4 out of 18 human tumors, while SV40 T antigen could only
be demonstrated in the tumors experimentally induced in hamsters. Antibodies to
SV40 T antigen were detected in 12 out of 60 patients with different tumors, in
1 out of 40 blood donors and in 7 out of 18 hamsters carrying experimentally
induced tumors.
Structure of simian virus 40 DNA in primary
and derived mouse tumors.
Naimski P, Defendi V.
Primary tumors induced by Simian Virus 40 (SV40) in nude mice, lines derived
from them, and subsequent transplanted tumors were analyzed as to the state of
the viral DNA. It was found that SV40 DNA is present only as integrated
molecules, each tumor having a different integration pattern; all tumors were
SV40 T-antigen positive by immunofluorescence. One tumor and its derivatives in
vitro and in vivo have maintained a single integration structure consisting of a
head-to-tail arrangement of a molecule slightly larger than whole-length viral
DNA (approximately equal to 1.1). This tumor expresses a full-sized T-antigen
and a Mr 54,000 host-associated protein.
Medulloblastoma in childhood: an
epidemiological study.
Farwell JR, Dohrmann GJ, Flannery JT.
The authors have reviewed 143 cases of
medulloblastoma in children aged 19 years or younger who were treated in a
42-year period and reported in the Connecticut Tumor Registry. About 20 cases
have occurred in each 5-year period since 1950, but 31 were seen between 1955
and 1959. Correspondingly, an excessive number of children born in the period
1954 to 1958 have developed medulloblastomas. A relationship to polio vaccine
contaminated with SV40 virus may exist. Children with medulloblastomas
had an increased number of immediate family members with brain tumors, leukemia,
and childhood cancer when compared to controls. In this series, the male to
female ratio was 1.33:1. Average age at diagnosis was 6 1/2 years, with most
children being diagnosed at 3 years old and fewer cases appearing in each
successive hemidecade from birth to 20 years of age. Probability of survival at
6 months was 0.687; at 1 year, 0.444; at 2 years, 0.314; and at 5 years, 0.222.
Survival probability was statistically significantly better in the years 1968 to
1977 than in previous decades, in part due to fewer autopsy diagnoses and
lowered operative mortality, but also due to a decreased mortality rate in
children several years after diagnosis. Fifty-one percent were treated with
operation and irradiation, 17% with operation alone, 12% with irradiation alone,
and 5% with operation, irradiation, and chemotherapy. Fifteen percent were not
treated. One- and 5-year survival rates in patients with operation and
irradiation were, respectively, 0.615 and 0.307; with operation, 0.125 and
0.042; with irradiation, 0.688 and 0.277; and with operation, irradiation, and
chemotherapy, 0.857 and 0.643. All seven children who received chemotherapy were
diagnosed after 1968, and five are still alive. Perhaps due to short follow-up
time, the course and mortality rate of children treated with all three
modalities were not statistically significantly different from those of children
treated since 1968 with operation and radiation therapy.
Mutagenic effects of DNA-containing oncogenic
viruses and malignant transformation of mammalian cells.
Shapiro NI, Marshak MI, Varshaver NB.
It was discovered in the 1970s that oncogenic
viruses could induce gene mutations in mammalian cells. The phenomenon seems to
be widespread: it was observed with all groups of DNA-containing viruses and
some retroviruses. The mutagenic effects of the tested viruses at gene
level are not locus specific. The viruses induce point mutations, including base
substitutions, as well as deletions and insertions. The mutagenic effect of SV40
is controlled by the activity of the early A gene, which encodes the T antigen.
Presumably, the process of integration creates the possibility for occurrence of
mutations early after infection. Mutagenesis seems to be induced by an
integrated virus, though to a much smaller extent. Virus-induced mutagenesis may
be connected with an activation of the cell error-prone repair systems. The sum
total of the experimental data shows that virus-induced mutagenesis and
transformation are interrelated: (A) viruses, like other carcinogenes, display
mutagenic activity; (B) viruses that are far removed from each other
systematically, whose only similarity lay in being oncogenic and capable of
integration, simultaneously showed the ability to induce gene mutations; (C)
agents changing the rate of transformation also changed the rate of gene
mutations: (D) The function of mutagenicity was mapped in the oncogene of SV40
(gene A); and the DNA of (E) mouse mammary carcinoma virus (MMTV) and avian
leukosis virus (ALLV) induced tumors has been found to contain nucleotide
sequences that transform 3T3NIH cells but do not carry any viral genetic
information. Mutagenesis induced by oncogenic
viruses may play a part in the multistage process of malignant transformation,
though its contribution may be different in various specific cases and for
different groups of viruses. Further studies of the uncommon mutagens, which
viruses seem to be, may greatly increase our knowledge of the virus-cell
relationship. An understanding of the extent of genetic danger inherent in
viruses and live viral vaccines is necessary for practical medicine.
A case of epidermodysplasia verruciformis (Lewandowsky-Lutz,
1922) with skin cancer: histopathology of malignant cutaneous changes.
Tanigaki T, Endo H.
Histopathology of tumors which occurred on the face of a 34-year-old man with
epidermodysplasia verruciformis was studied. The following three different types
of histological changes were recognized in the tumors: (1) intraepidermal
initial changes; (2) Bowen-like changes, and (3) squamous cell carcinoma.
Carcinogenetic processes in this case were
compatible with those reported by Todaro et al. in 1966, in which case tumors
were caused in vitro by SV40 virus. According to their results, initial
transformed cells were observed to occur at the lower layer of the epidermis. It
seemed likely that viral infection occurred at the epidermal lower layer and
immediately the infected cells divided and some of them changed to transformed
cells which then abnormally proliferated to show a malignant feature. Similar
histological changes were recognized and viral oncogenesis was suggested in this
case too.
Sixth Daniel C. Baker, Jr. Memorial Lecture,
Induction of cancer by DNA viruses.
Watson JD.
The life cycles of the tumor virus SV40 and
polyoma are discussed with particular emphasis on the role of the viral coded
proteins which mediate the transformation of normal cells into their cancerous
equivalents. One of more of these proteins possibly act by stimulating
directly the synthesis of cellular DNA, while others may mimic the action of
polypeptide mitogens that act at the cell surface.
Human papovaviruses of the polyoma-SV40
subgroup are common infectious agents. The clinical picture of the primary
infection of these viruses is unknown, but one of the viruses, the JC virus, is
associated with and probably causes progressive multifocal leukoencephalopathy.
These viruses cause tumours and transform cells of several species of laboratory
animals. Current results seem to rule them out as a major aetiollogical factor
in human cancer. However, their relationship to human tumours requires further
study, especially in immunologically compromised patients showing activation of
a latent infection.
The cells of tumours induced by many oncogenic
DNA viruses, or cells transformed in vitro, contain virus-specific T and
transplantation antigens; these have been described for SV40 virus, polyoma
virus and adenoviruses. The investigation of viruses as causes of malignant
disease in man has sought to establish whether tumour cells possess these
virus-specific proteins; however, to date and with the limitations of present
techniques, this enquiry has not demonstrated the above viruses as causal of
human cancer. More recent studies with herpesvirus type 2 (HSV-2) have
shown this virus to transform animal and human cells in culture, and induce
cancer in experimental animals: for these reasons, many researchers have
suggested that this agent may be an agent of some forms of cancer, in particular
carcinoma of the cervix. The possible association of HSV-2 with human malignant
disease is discussed.
Sarcomas induced by injection of simian virus
40 into neonatal CFW mice.
Hargis BJ, Malkiel S.
Sarcomas were induced in CFW mice by the iv
inoculation of simian virus 40 (SV40) in neonatal animals. Infection with murine
malaria parasites, Plasmodium berghei yoelli, decreased the latency and
increased the incidence and invasiveness of the tumors. All mice given both SV40
and P. berghei yoelli had sarcomas of the liver and spleen at 9 months of age.
At 11 months of age, 70% of the SV40-inoculated mice had sarcomas of the liver
indistinguishable from those in the group given both pathogens. Only 1 lung
metastasis was seen in the SV40-treated group. The sarcomas contained SV40
T-antigen as revealed by the indirect immunofluorescence technique. Among adult
CFW mice given iv injections of SV40, only 2 tumors were found at 11 or 12
months after virus inoculation. Both tumors were in the lungs; 1 was an adenoma
and 1 was a papillary adenocarcinoma. Neither gave a positive reaction with the
immunofluorescence test.
Isolation of a SV40-like Papovavirus from a
human glioblastoma.
Scherneck S, Rudolph M, Geissler E, Vogel F, Lubbe L, Wahlte H, Nisch G,
Weickmann F, Zimmermann W.
A human glioblastoma multiforme (M27) tested in
early cell cultures by indirect immunofluorescence staining showed SV40-related
tumor (T)-antigen, 95% of the cells being positive. SV40-related viral
capsid (V)-antigen was absent in all cells tested. Experiments to rescue this
virus were performed by fusing M27 cells with CV-I monkey cells, which were
permissive for SV40, using polyethylene glycol (PEG) as fusion factor. We
succeeded in isolating virus particles SV40-GBM which electron microscopy showed
to correspond in size and morphology to papovaviruses. Serological tests (hemagglutination,
neutralization, fluorescent antibody) revealed that the virus is
indistinguishable from SV40. Despite this apparent antigenic identity SV40-GBM
differs slightly from SV40 wild type. This virus can propagate and produce CPE
in both CV-I cells and primary fetal human kidney cells. Furthermore digestion
of SV40-GBM DNA with the HindII/III restriction endonucleases revealed minor
differences compared with the SV40 DNA. Therefore the virus SV40-GBM obtained
from glioblastoma cells seems to be closely related to the SV40-PML viruses
described earlier.
Screening of human brain tumors for
SV40-related T antigen.
Tabuchi K, Kirsch WM, Low M, Gaskin D, Van Buskirk J, Maa S.
A series of 39 human brain tumors has been
screened for the presence or absence of SV40-related T antigen by the direct and
indirect immunoperoxidase methods. Two tumors of ependymal origin (malignant
ependymoma, choroid plexus papilloma) revealed markedly positive nuclear
staining for T antigen both in vivo and in vitro. The relationship of
these tumors to their experimental counterparts inducible by recent human
papovavirus isolates is discussed.
Investigation of human urogenital tract
tumors of papovavirus etiology: brief communication.
Shah KV, Daniel RW, Stone KR, Elliott AY.
Cells cultured from human urogenital cancer and other cancers as well as cells
from noncancerous tissues were examined by immunofluorescent staining with
antibodies to T-antigens and capsid antigens of papovaviruses BK virus (BKV), JC
virus, and simian virus 40(SV40), and to capsid antigens of herpes simplex virus
types 1 and 2 and human cytomegalovirus (CMV). Cells from early passage cultures
of 123 primary tissues and from 14 continuous lines derived from transitional or
renal cell carcinoma were tested. None of the cell preparations was specifically
stained with any of the antisera. A serologic comparison of patients with
bladder cancer, patients with prostate cancer, and normal control groups of BKV
hemagglutination-inhibiting and SV40-neutralizing antibodies showed no
differences among the 3 groups. None of the sera in the 3 groups had SV40 or BKV
T-antibodies. In tests of supernatants of 35 primary cultures for presence of
virus, a single isolation, that of a cytomegalovirus, was made.
The study revealed no evidence that infection
with papovaviruses of the SV40-polyoma subgroup has any part in the production
of bladder and prostate cancer.
Induction of sister chromatid exchanges by
transformation with simian virus 40.
Nichols WW, Bradt CI, Toji LH, Godley M, Segawa M.
The frequency of sister chromatid exchange (SCE) has been followed sequentially
after the addition of SV40 to human diploid fibroblast cultures. The SCE
frequency was nearly the same in uninfected controls and in infected cultures
before they became tumor antigen positive. When cells exhibited tumor antigen,
the SCE frequency increased over a wide range, and changes in chromosome number
and structure were observed simultaneously. Cells with induced chromosome
abnormalities without increased SCE's and the reverse present the possibility
that the two phenomena have different viral mechanisms.
This increase in SCE can be added to the
previously demonstrated change in chromosome number and increase in chromosome
breakage and rearrangement as indicators of genetic damage associated with viral
transformation.
Susceptibility of human skin fibroblasts to SV40 virus infection has been
suggested as a marker of cancer risk. To evaluate the role of heritable factors
in the regulation of SV40 T-antigen, fibroblasts from 9 pairs of identical twins
and 129 members of cancer-prone families, including 16 with cancer, were tested
in a 3-day immunofluorescence assay. In the twin study, the variance of
T-antigen values was significantly less in identical than in fraternal or
non-twin sibs, suggesting a heritable component in the regulation of SV40
infection. In the families, T-antigen values of parents and children were
compared to models of Mendelian inheritance. At least three modes of
inheritance--autosomal dominant, recessive, and X-linker--were observed. The
distribution of offspring values compared to those of their parents suggested
that interaction of multiple genetic factors influences the T-antigen value in
individual patients. With the exception of Fanconi's anemia, the values for
patients with cancer or predisposing syndromes were not uniformly elevated.
The utility of this assay as a marker of cancer
risk appears limited because of the complexity of factors that influence
T-antigen expression in individual cases.
Fibroblast cultures were prepared from skin biopsies from 29 patients and tested
for their susceptibility to transformation by simian virus SV40. Cells with a
normal chromosome complement showed a mean transformation frequency of 25/106
cells but for cells from a single patient with Fanconi's anaemia, the value was
152/106 cells. An increased susceptibility to transformation was observed for
cells from 6 patients with Down's syndrome 3 patients with trisomy 18, a patient
with trisomy 18 for 5% of cells and a patient with trisomy 13. No increased
susceptibility to transformation was found for cells with a chromosome
complement of XO, XXY, XX/XX + 8, XX + partial 15q or XX + 9p.
The susceptiability to transformation was
related to susceptibility to SV40 virus infection, as measured by the number of
infected cells which contained SV40 virus induced T antigen. This latter test
was technically easier to perform and could serve to detect persons of increased
susceptiability to transformation, since this may indicate an increased risk of
natural malignant disease.
Studies on the presence of viral antibodies
in patients with various forms of malignant neoplasia.
Stoian M, Hozoc M, Ionescu T, Lungu M, Predescu E, Nastac E.
The study of viral antibodies in 6714 sera
demonstrated that the level of antibodies to infectious viruses was the same in
cancer patients as in controls. However, the patients with various forms of
neoplasia showed a considerable percentage and high antibody levels to viruses
with oncogenic potential for animals (adenovirus, SV40, Rous virus) or involved
in human carcinogenesis (herpes virus).
ALL INFORMATION, DATA, AND
MATERIAL CONTAINED, PRESENTED, OR PROVIDED HERE IS FOR GENERAL INFORMATION
PURPOSES ONLY AND IS NOT TO BE CONSTRUED AS REFLECTING THE KNOWLEDGE OR OPINIONS
OF THE PUBLISHER, AND IS NOT TO BE CONSTRUED OR INTENDED AS PROVIDING MEDICAL OR
LEGAL ADVICE. THE DECISION WHETHER OR NOT TO VACCINATE IS AN IMPORTANT AND
COMPLEX ISSUE AND SHOULD BE MADE BY YOU, AND YOU ALONE, IN CONSULTATION WITH
YOUR HEALTH CARE PROVIDER.
"A foolish faith in authority is the worst enemy of truth."
-- Albert Einstein, letter to a friend, 1901
"I know of no safe depository of the ultimate powers of the society but the people themselves, and if we think them not enlightened enough to exercise control with a wholesome discretion, the remedy is not to take it from them, but to inform their discretion by education."
-- Thomas Jefferson, letter to William C. Jarvis, September 28, 1820
"What's the point of vaccination if it doesn't protect you from the unvaccinated?"
-- Sandy Gottstein
"Who gets to decide what the greater good is and how many will be sacrificed to it?"
