Inhibition of the transformed phenotype of carcinoma cells that contain human papillomavirus.
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Carcinoma↗
Biomedical subjects
E J Shillitoe
Publications and source records attributed to E J Shillitoe.
Effect on cancer cells of plasmids that express antisense RNA of human papillomavirus type 18.
Some human squamous cell carcinomas contain DNA of human papillomaviruses (HPV) and express RNA from the E6 and E7 genes. We have examined the effect of plasmids that express antisense RNA of these genes on the growth of the human cancer cell lines HeLa, C4-1, and 1483, which contain HPV type 18 DNA. As controls, the human cancer cell line 183 and the Vero line of monkey kidney cells were used, which do not contain HPV. Plasmids were introduced into the cells by electroporation; cells that contained HPV type 18 accepted the antisense-expressing plasmids at a lower frequency than the cells that lacked HPV. Cell lines were developed from HeLa cells that contained sense- or antisense-expressing plasmids, and lines that contained antisense-expressing plasmids showed slower growth, reduced ability to form colonies in soft agar, and increased serum requirements. The use of antisense HPV RNA might be a suitable approach to gene therapy of HPV-expressing human cancers.
Animals↗
Oral virus infections: the potential for gene transfer in treatment and prevention.
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Animals↗
Prevalence of Epstein-Barr virus and human papillomavirus in oral mucosa of HIV-infected patients.
Epstein-Barr virus (EBV) has been implicated in the genesis of oral hairy leukoplakia (OHL). Initially, OHL was also associated with human papillomavirus (HPV) as evidenced by staining with antiserum to papillomavirus common structural antigens and reports of two HPV-positive OHL as detected by in situ DNA hybridization. The aims of this study were to determine the prevalence of EBV and HPV DNA in OHL and normal oral mucosa and to explain the basis for the staining of OHL tissues with antibodies to papillomavirus common structural antigens. EBV DNA was detected by in situ hybridization in 47 of 47 cases of OHL from human immunodeficiency virus (HIV)-seropositive individuals and in 1 of 10 biopsies of clinically normal buccal mucosa from the same group of individuals. Twenty-five of 35 OHL specimens stained with antibody to papillomavirus common structural antigens. There was no staining of two EBV-containing lymphoblastoid lines, indicating that the staining with anti-papillomavirus antibody was not due to antigenic cross-reactivity with EBV antigens. HPV DNA was detected by polymerase chain reaction amplification in 10 of 18 OHL specimens and in 6 of 10 normal buccal mucosa specimens. Our results indicate that EBV and HPV are present frequently in OHL and that HPV can be found regularly in histologically normal mucosa.
Acquired Immunodeficiency Syndrome↗
HIV-related infections of the oral cavity.
Oral infections are among the first manifestations of infection by the human immunodeficiency virus (HIV). They include fungal, viral, and bacterial infections and range from being essentially trivial, through troublesome to life threatening. Although some infections are due to overproliferation of the normal oral flora, others are due to organisms that normally are not found in the mouth. The clinical features of many of these infections have now been characterized, and clinical trials have indicated the optimal management. However, for many infections, the underlying processes are still not clear, and improvements in treatment are necessary.
Acquired Immunodeficiency Syndrome↗
Analysis of complex mutations induced in cells by herpes simplex virus type-1.
The shuttle vector plasmid pZ189 was used as a target for mutagenesis in COS-1 cells. Complex mutations were analyzed in 5 plasmids that were recovered from noninfected cells and 15 plasmids that were recovered from cells that were infected with herpes simplex virus type-1 (HSV-1). Complex mutations in noninfected cells consisted of duplications and rearrangements of plasmid DNA, while those from cells that were infected with HSV-1 included 8 that were enlarged due to insertion of other DNA sequences. Plasmids that contained inserted DNA showed an increase in size of from 118 bp up to around 4500 bp. Maps were constructed based on restriction enzyme digestion, and some or all of each plasmid was examined by DNA sequencing. The inserted DNA was not derived from HSV-1 in any case, since it did not hybridize to DNA from HSV-1 and showed no sequence similarities to the virus. Instead, inserted DNA was found to hybridize to HindIII-digested cellular DNA as a single or double band in 5 plasmids and contained multiple repeat sequences such as alpha satellite, Alu or Kpn repeats in 4 plasmids. In four enlarged plasmids the identity of the inserted sequences could not be determined. The junctions between the shuttle vector and the inserted DNA did not show features of transposable elements and no homology was detected between inserted sequences and the sequence at the insertion site. No preferred site for recombination was detected. Although no similarities were found among the inserted sequences, it is possible that the cellular sequences represent cellular targets for virus-mediated rearrangement. It appears that HSV-1 stimulates nonhomologous recombination between DNA sequences in virus-infected cells.
Animals↗
Viruses and oral cancer.
Oral cancer is a disease with a complex etiology. There is evidence for important roles of smoking, drinking, and genetic susceptibility, as well as strong indications that DNA viruses could be involved. The herpes simplex virus type 1 has been associated with oral cancer by serological studies, and animal models and in vitro systems have demonstrated that it is capable of inducing oral cancer. Papillomaviruses are found in many oral cancers and are also capable of transforming cells to a malignant phenotype. However, both virus groups depend on co-factors for their carcinogenic effects. Future research on viruses and oral cancer is expected to clarify the role of these viruses, and this will lead to improvements in diagnosis and treatment of the disease.
Animals↗
Relationship of viral infection to malignancies.
The epidemiologic features and the biologic mechanisms of the oral tumor viruses have been clarified during the period under review. The use of more sensitive detection methods has confirmed that the prevalence of papillomaviruses in the mouth in patients with various lesions, as well as in healthy individuals, is high. Mechanisms by which papillomaviruses are regulated by cells and by which they may control cell behavior have been described. Herpes simplex virus has been shown to induce a variety of mutations in cells. Human herpesvirus type 6 has been found in oral tissues of a number of healthy individuals, and the virus was shown to be capable of transforming cells to a malignant phenotype. Hairy leukoplakia was found in other individuals who were not infected with the human immunodeficiency virus, and a new animal model for hairy leukoplakia was introduced. Because of the high frequency of oral viruses in nondiseased individuals, more attention must be paid to their mechanisms of action before it will be possible to define the role of viruses in oral malignancies.
Animals↗
A computer program that simulates cloning of DNA.
Easy Cloner is a computer program that manipulates DNA sequences as in cloning experiments and produces maps of the resulting plasmids. The program runs in the graphics mode of an IBM PC or compatible computer and is operated by using a mouse to point to the required actions. The program is available in the public domain.
Cloning, Molecular↗
Cytogenetic characterization of 20 lymphoblastoid lines derived from human individuals differing in bleomycin sensitivity.
Forty lymphoblastoid (lymphoid) lines were established from 42 volunteer blood donors, including healthy individuals and patients with head and neck carcinomas. Each peripheral blood sample was split into two portions, one for the establishment of a lymphoid line and the other for short-term culture, which was used to estimate bleomycin sensitivity by cytogenetic procedures. Twenty lymphoid lines were selected at random to compare bleomycin sensitivity with data obtained from short-term lymphocyte cultures. In each set, bleomycin sensitivity of lymphoid cells was similar to that of the lymphocytes. The lymphoid lines, which can be propagated for an unlimited supply of relatively homogeneous cellular material, will be useful for a variety of future investigations.
Bleomycin↗
Oral melanotic macules in patients infected with human immunodeficiency virus.
A group of 217 patients seropositive for human immunodeficiency virus (HIV) were studied for 2 years, during which time pigmented lesions of the oral mucosa developed in 14 (6.4%) of them. The lesions were well circumscribed in some cases and diffuse in others. In some patients the macules enlarged or recurred after surgical excision. In two patients the macules appeared during the administration of zidovudine. Clinical and laboratory evidence of adrenal insufficiency was not detected in any of the patients examined. The histologic appearances were those of melanotic macules. No ultrastructural alterations of the melanocytes were observed. Two of these macules also contained Epstein-Barr virus, and in one case normal oral mucosa was examined and also contained Epstein-Barr virus in the epithelial cells. As a control group we examined 180 health care workers who did not belong to any risk category, and 30 intravenous drug abusers who tested seronegative to HIV. Oral melanotic pigmentation was found in eight of the control subjects (3.6%). The difference was not statistically significant (p = 0.3097). Our study shows that oral macules do not occur more frequently in HIV-infected patients. However, the clinical behavior of these lesions appears to be different during the course of HIV infection. In some HIV-infected patients the cause of the macules might relate to the administration of zidovudine and antifungal or antibacterial drugs. In others the cause remains unknown and could be due to multiple factors.
Adult↗
DNA sequence of mutations induced in cells by herpes simplex virus type-1.
The shuttle vector plasmid pZ189 was used to find the kinds of mutations that are induced in cells by herpes simplex virus type-1 (HSV-1). A significant increase in mutation frequency was detected as early as 2 hr after infection, and reached a peak of two- to sevenfold over background at 4 hr after infection. Several differences were detected between spontaneous mutants and those induced by HSV-1 when they were analyzed by gel electrophoresis and DNA sequencing. Point mutations accounted for 63% of spontaneous mutants but for only 44% of HSV-1-induced mutants (P less than 0.05). In each case the predominant type of point mutation was the G:C to A:T transition, which comprised 51% of point mutations induced by HSV-1, and 32% of spontaneous point mutations. Deletions of DNA were seen in HSV-1-induced mutants at a frequency of 44%, compared with only 29% in spontaneous mutants. HSV-1-induced deletions were less than half the length of spontaneous deletions, and 3 contained short filler sequences. An increase in size was seen in 13% of HSV-1-induced mutants and was due either to duplication of plasmid DNA, or, in 8 instances, to insertion of sequences derived from cellular DNA. Among spontaneous mutants, only 8% were increased in size and none of them had inserted cellular DNA. The proportion of complex mutants increased as infection by the virus progressed and they accounted for 79% of mutants at 24 hr after infection. The observed mutations have implications for understanding the "hit and run" mechanism of malignant transformation of cells by HSV-1.
Animals↗
Prevalence of human papillomavirus in inverted nasal papillomas.
Human papillomaviruses (HPVs) have been associated with both benign and malignant neoplasms of the head and neck. To determine the prevalence and types of HPVs in this neoplasm, we examined fixed tissue from 21 patients by in situ hybridization. Human papillomavirus types 6b and 11 probes were used, and hybridization was positive to both types in lesions from 16 (76%) of 21 patients. Hybridization to the HPV-11 probe resulted in the most intense nuclear staining throughout the epithelium, probably indicating a high copy number. The high incidence of viral DNA sequences found in this tumor suggests a potential etiologic role. Therapeutic implications are briefly discussed.
Adult↗
Shedding of herpes simplex virus type 1 into saliva.
The shedding of herpes simplex virus Type 1 (HSV-1) into saliva was studied in 110 healthy individuals, 55 patients who were undergoing oral surgery, and 21 immunocompromised patients in an asymptomatic Japanese population. Specimens were collected regularly for up to 2 months from each individual. Virus was recovered on at least one occasion from 4.5% of the healthy individuals, 20% of the patients undergoing oral surgery and 38% of the immunocompromised patients. In those individuals that shed virus, the duration of shedding varied from a mean of 1.2 days in the healthy individuals to 5.8 days in patients undergoing oral surgery, and was 3 days in the immunocompromised patients. The majority of oral surgery patients shed HSV-1 into saliva within 7 days after surgery and immunocompromised patients shed virus almost continuously. The role of surgery in triggering the reactivation of latent HSV-1, and the differences in rates of viral shedding between American and Japanese are discussed.
Adult↗
Examination of herpes virus DNA sequences for patterns that resemble transposable elements.
Herpes viruses are associated with many types of oral tumors and can be tumorigenic in vitro. The mechanisms by which they transform cells remain unknown, but it has been suggested that they contain insertion sequences. Two algorithms were developed to search DNA sequences for the presence of patterns that characterize such elements using either selection or scoring techniques. When randomly shuffled herpes virus sequences were examined each algorithm detected many such patterns but the scoring algorithm found fewer than the selection algorithm. In the herpes virus sequences that are known to contain such patterns the computer techniques found many more than previously reported, indicating that visual inspection alone is not adequate. The frequency of these patterns suggest that although the transforming regions of herpes virus DNA could contain transposable elements experimental confirmation is necessary.
Algorithms↗
Five-year survival of patients with oral cancer and its association with antibody to herpes simplex virus.
Levels of antibody to herpes simplex virus type 1 (HSV-1) were measured in 70 patients with untreated squamous cell carcinoma of the mouth. After treatment the actuarial survival was determined at quarterly intervals for 5 years and was found to be associated with the pretreatment level of antibody to the virus. Patients with levels of IgM antibody to HSV-1 which were above the median level had a 5-year survival of only 56% whereas those with levels below the median had a higher survival of 72%. Patients with no detectable IgM antibody to HSV-1 had a 5-year survival of 81%. The reverse was seen with IgG antibody to HSV-1. Patients with higher than the median level of IgG antibody had a 5-year survival of 73%, whereas those with IgG antibody below the median had a 5-year survival of 56%. No relationship was seen between survival and levels of IgA antibody to HSV-1, or between survival and antibody of any class to cytomegalovirus. The data are consistent with the reported association between oral cancer and HSV-1.
Aged↗
Induction of mutations in bacteria by a fragment of DNA from herpes simplex virus type 1.
A bacterial assay was developed for the study of mutagenesis by DNA of herpes simplex viruses. The histidine mutations from two of the Ames mutagenesis tester strains were recombined into the Salmonella histidine operon of the F'8 plasmid and each was transferred to a derivative strain of E. coli C from which the resident histidine operon had been deleted. One tester strain could be reverted by a chemical mutagen which induces frameshift mutations and the other could be reverted by a mutagen which induces base-pair substitution mutations. The BamHI G fragment of herpes simplex virus type 1 was cloned in each orientation into the BamHI site of the expression vectors pUC7, pUC8 and pUC9 and were introduced into the new strains of E. coli. The pUC9 plasmid carrying the BamHI G fragment of herpes simplex virus type 1 with the G-E' site closest to the lac promoter showed a higher rate of reversion in the frameshift strain, which varied up to 39-fold greater than the background rate. Since many mutagens are carcinogenic these data suggest the existence of a mutagenic peptide of herpes simplex virus type 1 which might be involved in cell transformation.
Cloning, Molecular↗
Detection of herpes simplex virus proteins in cultured cells by monoclonal antibodies and the avidin-biotin-immunoperoxidase complex method.
The distribution of 5 proteins of herpes simplex virus Type 1 was observed in cells that had been infected for various periods. The cells were stained with monoclonal antibodies to ICP4, ICP5, ICP6, ICP8, and gB, using the avidin-biotin-peroxidase complex (ABC) method. Each protein had a characteristic pattern of time of appearance and translocation by which it could be distinguished from the others. The sensitivity of the ABC technique, its ease of use, and the permanence of the preparations make this method well suited for the study of viral proteins.
Animals↗