HHV-8 Sequences Research Articles

HHV-8 positive clinically advanced castrate-resistant prostate cancer (mCRPC): A potentially distinct molecular subset.

163 Background: Herpesvirus HHV-8 has been detected in normal prostate tissue and has been linked to the acquisition of androgen-independent growth of prostate cancer cells in vitro, early development of anti-androgen therapy resistance, and more aggressive disease in a subset of patients with mCRPC. We used comprehensive genomic profiling (CGP) to test the hypothesis that HHV-8 associated mCRPC is a distinct molecular subset of mCRPC featuring altered AR signaling compared to HHV-8 negative mCRPC. Methods: Using a hybrid capture-based FDA-approved CGP assay, a series of 4,918 mCRPC were sequenced to evaluate all classes of genomic alterations (GAs). Tumor mutational burden (TMB) was determined on up to 1.1 Mb of sequenced DNA and microsatellite instability (MSI) was determined on 114 loci. PD-L1 expression was determined by IHC (Dako 22C3). HHV-8 status was determined by CGP-based identification of unique viral reads. Results: Overall, 128 (2.6%) of the mCRPC cases harbored HHV-8 sequences (Table). The patient age distribution was similar in the HHV-8+ and HHV-8− groups. GAs in AR, predominantly amplifications, and RAD21 were slightly reduced in HHV-8+ mCRPC cases (P = .01 for both). The frequencies of GAs in genes associated with mCRPC including PTEN, BRCA2, ATM, CDK12, and the TMPRSS2: ERG fusion were similar in the two groups. HHV-8+ patients were less likely to be of European ancestry (P = 0.001). Putative biomarkers associated with immunotherapy response (MSI status, TMB, and PD-L1 expression) were also similar in the two groups. Conclusions: HHV-8+ mCRPC is a rare sub-type of mCRPC with lower frequency of AR GAs compared to HHV-8− mCRPC cases, suggesting a potential role of HHV-8 in inducing androgen-independent prostate cancer growth. Additional alterations associated with potentially enhanced PARP inhibitor response seen in HHV-8+ mCRPC (e.g., RAD21) suggest that early performance of CGP may inform precision treatment strategies to be tested in clinical trials. [Table: see text]

Cancer Diagnostic and Predictive Biomarkers 2015.

Early cancer diagnosis and prediction of responsivity to treatments are crucial elements for favorable prognosis and increased effectiveness of therapies in cancer patients. This special issue compiles relevant articles focused on the development of innovative cancer biomarkers and their validation. Availability of advanced molecular biology tools, including deep-sequencing and RNA-seq, allowed the identification of new unique biomarkers for more sensitive and specific diagnosis and the possibility to select new candidate therapeutic targets for personalized medicine. The negative prognostic role of stem cell markers, with higher progression rate, has been reported by G. Chene et al. in preinvasive tubal lesions of ovarian carcinoma, F. Collina et al. in triple negative breast cancers, and S. H. Kim et al. in prostate cancer patients, where high level of GAPDH mRNA is a significant predictor of recurrence. The negative prognostic value, particularly for head and neck cancers, of cell-to-cell internalization was reported by M. Schwegler et al., and inverse correlation with expression levels of iNOS was reported by L. Yang et al. Different biomarkers have been analyzed for early diagnosis and follow-up monitoring spanning within serum detection of glycosylated biomarkers (described by A. Kirwan et al.), serum levels of 15-F2t-isoprostane in non-melanoma skin cancer (B. Freitas et al.), and methylation levels of GALR1 promoter in endometrial lesions by pyrosequencing analysis (C. Kottaridi et al.). Early diagnosis of liver metastasis by CT perfusion has been reported by Y. Wang et al. Predictive therapeutic biomarkers have been described to chemoradiation in locally advanced rectal cancer by R. Conde-Muino et al. and to radiotherapy in localized prostate cancer by A. Wilkins et al. The role of miRNAs in cancers and their prognostics relevance has been analyzed by A. Min et al. in oral carcinogenesis. J. Long et al. have focused on miRNA 193b in human breast cancer, and S. Wu et al. on miRNA 125b in gastric cancer. Finally identification of HHV-8 sequences in conjunctival neoplasia of Uganda patients offers the possibility to unveil a further cancer association to such herpesvirus and highlights the need to look for indirect role of virus with low-modest oncogenic potential in immunodeficient patients (including low risk HPVs in HIV-positive patients). By compiling these papers, we hope to enrich our readers and researchers with respect to the current wide range of cancer biomarkers, actively pursued by several worldwide research groups. Franco M. Buonaguro C. David Pauza Maria Lina Tornesello Pierre Hainaut Renato Franco

Detection of human herpesvirus 8 sequences in cutaneous cherry angiomas

Cherry angiomas (CAs) are common vascular benign skin tumors, characterized by abnormal angiogenesis, whose etiology is still unclear and poorly studied. We investigated the presence of HHV8 in CAs due to virus ability of inducing neoangiogenesis in endothelial cells. A total of 29 patients were enrolled in a randomized, controlled, blinded analysis of skin specimens including various vascular lesions. All clinical samples were anonymized and analyzed by three different biomolecular assays to minimize the risk of false positive/negative results. Results showed that 53% of eruptive CAs harbor HHV8 sequences, with the highest viral loads in samples derived from immunosuppressed patients. By contrast, no paucilesional CAs were positive for HHV8. Considering HHV8 prevalence in the Mediterranean population (10-15%), results obtained in eruptive CAs are significant and suggest for the first time a possible involvement of HHV8 in eruptive cherry angiomas development, particularly in the context of immunosuppression, similar to that recognized for major HHV8-induced pathologies.

Molecular epidemiology of human herpesvirus 8 variants in Kaposi's sarcoma from Iranian patients

Kaposi's sarcoma (KS) is a rare cancer in Iran and there is no epidemiological and molecular information about HHV-8 variants circulating among the Iranian population. In this study HHV-8 sequences have been analyzed in 43 cutaneous KS biopsies from Iranian patients mainly affected by classic KS. DNA samples were subjected to PCR amplification of HHV-8 ORF26, T0.7 and K1 followed by direct nucleotide sequencing and phylogenetic analysis. The analysis of ORF26 showed that 30 (69.8%) and 13 (30.2%) samples belonged to subtypes A/C and K, respectively. In general, the clustering of HHV-8 T0.7 variants paralleled that of ORF26. Genotyping of K1 sequences showed that the majority of samples (39 out of 41) fall into the large C clade with only 2 belonging to the A clade. In conclusion, HHV-8 variants identified among classic Iranian KS are largely related to Eurasian genotypes previously identified in KS from Mediterranean, Middle East, and East Asian regions.

Primary Mediastinal Large B-Cell Lymphoma: A Single-Center Study of Clinicopathologic Characteristics

Primary mediastinal large B-cell lymphoma (PMLBCL) is a subset of LBCL with unique clinicopathologic features. Some studies have raised the question of differences in biological features and clinical course among patients from different parts of the world. We conducted a retrospective clinicopathologic analysis of 24 patients with PMLBCL from a single center in Croatia. We also conducted the first investigation of the frequency of lymphotropic viruses human herpesvirus 6 (HHV-6) and HHV-8 in lymphoid lesions of this disease. The clinical characteristics of the patients were as expected, with high International Prognostic Index scores, elevated serum lactate dehydrogenase (LDH) levels, and bulky disease being adverse prognostic factors. Only 6 patients (25%) showed CD30 expression, and Bcl-6 protein expression was, in our series, prognostically favorable (P = .0401). One patient's tumor had detectable HHV-6 genome sequence, but no HHV-8 sequences were detected in any tumors. Two thirds of the patients received CHOP chemotherapy (cyclophosphamide, hydroxydaunomycin, vincristine, and prednisone) with a relatively low complete remission rate (43.8%; median follow-up, 33.8 months). This study confirmed the moderate preponderance among PMLBCL patients of young females with B symptoms and elevated LDH levels. The CHOP regimen proved effective as first-line therapy only in patients with limited disease. Therefore, other third-generation chemotherapy protocols may be considered for treatment, especially in patients with bulky and advanced disease.

Kaposi's sarcoma-associated herpesvirus/human herpesvirus 8-associated polyclonal body cavity effusions that mimic primary effusion lymphomas

Department of Clinical and Experimental Medicine, Amedeo Avogadro University of Eastern Piedmont, NovaraDear Sir,Kaposi’s sarcoma-associated herpesvirus/human herpesvirus8 (KSHV/HHV-8) is involved in the pathogenesis of Kaposi’ssarcoma (KS) and 2 lymphoproliferative diseases, primaryeffusion lymphoma (PEL) and the plasmablastic variant ofmulticentric Castleman’s disease (MCD). PEL is defined as aliquid-phase lymphoma harboring monoclonal rearrangementsof immunoglobulin (Ig) variable genes or, more rarely, of T-cell receptor (TCR) genes. As a rule, these elements are usedfor PEL diagnosis together with the demonstration of HHV-8infection of the tumor clone. However, as pointed out by Bou-langer and colleagues in the 1 July 2005 issue of the Interna-tional Journal of Cancer,

Human Herpesviruses in Primary Ocular Lymphoma

Several human lymphotropic herpesviruses have been found in certain lymphoproliferative disorders and implicated as possible etiologic factors or as modulating elements of the diseases. To assess a possible association of the human herpesviruses with lymphomas arising from mucosa-associated lymphoid tissue (MALT), we evaluated the presence of four human herpesviruses, Epstein-Barr virus (EBV), human herpesvirus 6 (HHV-6), HHV-7, and HHV-8, in the biopsied specimens from 14 patients with primary ocular MALT lymphomas. EBV DNA sequences were detected in four specimens by polymerase chain reaction (PCR), and four cases were positive for HHV-6 DNA. In situ hybridization showed that three and two of 14 cases were positive for EBV mRNA and HHV-6 DNA, respectively. Neither HHV-7 nor HHV-8 sequences could be detected by PCR. These findings would stimulate further investigation as to the involvement of these lymphotropic viruses in the pathogenesis of a subset of low-grade primary ocular lymphomas.

Absence of Kaposi's sarcoma-associated virus (human herpesvirus-8) sequences in angiosarcoma.

Kaposi's sarcoma-associated virus (human herpesvirus-8 [HHV8]) sequences have been identified in both AIDS-associated and AIDS-non-associated Kaposi's sarcoma, but data relating to the detection of HHV8 sequences in angiosarcoma are variable. One study showed HHV8 sequences in 29% (7/24) of angiosarcomas, but others have not confirmed these results. We evaluated 33 angiosarcomas for HHV8 sequences to determine the frequency of this virus in angiosarcomas and its possible pathogenetic significance. Five cases of Kaposi's sarcoma from HIV-positive patients were used as positive controls. Five additional cases of Kaposi's sarcoma collected approximately 40-50 years ago were also analysed, and three were HHV8 positive. None of the angiosarcomas revealed HHV8 sequences after the standard 35 cycles of PCR. In 6 cases, nested PCR revealed the presence of HHV8 sequences. These results were not reproducible when outer primers (based on sequences outside of the earlier PCR products) were used for amplification. This suggests that the HHV8 sequences detected in 6 cases represent a low level of contamination. In contrast, HHV8 sequences were found in all Kaposi's sarcomas with well-preserved DNA after standard 35-cycle PCR amplification. These findings confirm a close association between Kaposi's sarcoma and HHV8 infection and suggest that HHV8 is not involved in the pathogenesis of angiosarcoma.

HHV-8 infection is specific for cell lines derived from primary effusion (body cavity-based) lymphomas.

Human herpes virus 8 (HHV-8; or KSHV, Kaposi's sarcoma-associated herpes virus) is a gamma herpes virus with sequence homology to Epstein-Barr virus (EBV). It was first isolated from Kaposi's sarcoma tumor cells and subsequently from tumor cells and peripheral blood mononuclear cells from patients with primary effusion lymphomas (PEL; or body cavity-based lymphomas). PEL has been recognized as an individual nosologic entity based on its distinctive biological-pathological features and its consistent infection with HHV-8 (commonly, but not always co-infected with EBV), occurring predominantly in human immunodeficiency virus (HIV)-infected patients but occasionally also in HIV-negative cases. Whether HHV-8 sequences can be found also in non-hematopoietic tumor cells other than Kaposi's sarcoma and in malignant hematopoietic malignancies other than PEL, has been the focus of the present studies. We examined the presence of HHV-8 sequences by polymerase chain reaction (PCR) using (1) a panel of 133 human cell lines established from a large variety of solid tumors; (2) a spectrum of 114 hematopoietic cell lines derived from the different cell lineages including 50 B cell leukemia/lymphoma-derived cell lines and seven cell lines established from patients with PEL. Besides the seven PEL cell lines, 46 cell lines that were derived from malignant pleural effusion or ascitic fluid material (25 non-hematopoietic and 21 hematopoietic cell lines) were examined. Except for the seven PEL cell lines that were strongly HHV-8+ in the PCR, all solid tumor cell lines and all hematopoietic cell lines scored consistently negative for the presence of HHV-8 sequences. These results confirm the absolute specificity of HHV-8 infection (within the hematopoietic malignancies) for PEL. PEL cell lines represent useful tools for the analysis of the biology of this neoplasm and of the pathogenetic role of the virus in the disease development.

Detection and semiquantitative analysis of human herpesvirus 8 DNA in specimens from patients with Kaposi's sarcoma.

Kaposi's sarcoma (KS) is the most common neoplasm in patients with AIDS. Epidemiologic evidence and the recent identification of herpesvirus-like DNA sequences in patients with KS have suggested a role for viral agents in the etiopathogenesis of this disease. It is unclear if these sequences are present in all types of KS and if the copy number of these sequences has a correlation with disease severity (staging). In order to clarify these issues, we retrospectively analyzed, by PCR and Southern blotting, formalin-fixed, paraffin-embedded biopsy specimens from 12 patients previously diagnosed with KS by histopathologic examination of these specimens between the years of 1977 and 1996. We also analyzed tissue samples from these patients taken from dermal sites without KS lesions and control tissues from healthy subjects. Of the 12 patients, 6 had classic KS, 5 had AIDS-associated KS, and 1 had the endemic type of KS. We tested the specimens for other herpesviruses, including cytomegalovirus, human herpesvirus 6 (HHV-6), HHV-7, HHV-8, and Epstein-Barr virus. Of 20 biopsy specimens from patients previously diagnosed with KS, 19 were positive for HHV-8 sequences (95%), while PCR for the DNAs of other herpesvirus agents was negative. Uninvolved tissue from these patients and control tissue from healthy subjects gave negative results for all viruses. Semiquantitative analysis of Southern blots showed higher levels of HHV-8 DNA in those patients with multicentric and visceral involvement than in those patients with localized involvement. In addition, in patients with localized skin disease, the nodular stage had higher levels of HHV-8 DNA than the patch or plaque stages. Our data confirmed that HHV-8 is involved in the etiopathogenesis of all types of KS and that there is a correlation between HHV-8 DNA load and the severity and staging of this disease.

RISK FACTORS AND HUMAN HERPES VIRUS-8 (HHV8) DURING KAPOSI'S SARCOMA (KS) IN KIDNEY TRANSPLANT PATIENTS

62 Pathogenesis of post-transplant KS is still unknown despite recent reports of association with HHV8. In order to analyze KS risk factors and value of HHV8 detection among GCIF kidney transplant patients (TP) alive between 1996-1998, we retrospectively compared TP who had developed post transplant KS before january 1996 with 2 matched controls (C) for age, sex, transplant year and center. Before KS diagnosis or at same follow-up for C, we analyzed: ethnic origin; number of blood transfusions, grafts and rejection episodes; immunosuppressive regimen; tuberculosis, i.v. treated fungal, CMV or herpes infections; serological status for HIV 1-2, CMV, Hepatitis B and C, Herpes; blood cell count, renal function. We searched for HHV8 viremia by PCR at time of study (ts), HHV8 antibodies (Ab) before and after the graft. Sera were tested blindly using 2 indirect immunofluorescence assays (IFA) with 2 derived HHV8(+) EBV(-) cell lines and ELISA (recombinant protein encoded by HHV8 orf 65) confirmed by western blot. We then prospectively followed HHV8 detection within KS lesions, normal skin and peripheral blood mononuclear cells (PBMC) in TP with KS onset at ts. Among 30 TP with KS 21±34 months (M) after grafting, 24 were in remission, 1 had stable KS, 5 were lost from follow-up at ts. 9 other TP had KS onset at ts. After, conditional logistic regression analysis, TP geographic origin (OR=15.1, p<0.001), antiHbc Ab (OR = 5.9, p<0.005), hemoglobin <12g/dl (OR=6.3, p<0.001), lymphocytes <750/mm3 (OR=9.0, p<0.05) were KS risk factors but not use of cyclosporine (OR=0.2, p<0.05). Stepwise logistic regression showed independant increased risk with African origin (OR=50.0, p<0.001) and initial use of polyclonal antilymphocytes sera (OR=11.3, p<0.05). HHV8 Ab were detected before graft in 100% of 5 KS vs 10% of 20 C(p<0.001); after graft, in 92% of 13 KS at 74±66 M vs 34% of 23 C at 87±58 M(p<0.01). At ts, HHV8 viremia was detected in 11% of 9 (1 stable, 8 remission) KS vs 0% of 13 C (ns). Prospective study detected HHV8 sequences in recipient's KS lesions (6/6), normal skin (3/5), PBMC (7/9) and HHV8 Ab in 88% of 6 KS at KS onset with negativation of viremia in 3/3 TP with clinical resolution. This first case-control analysis identifies KS risk factors. It suggests that HHV8 Ab detection before graft is predictive of KS and viremia is related to tumor burden and evolution.

RISK FACTORS AND HUMAN HERPES VIRUS-8 (HHV8) DURING KAPOSI'S SARCOMA (KS) IN KIDNEY TRANSPLANT PATIENTS.

62 Pathogenesis of post-transplant KS is still unknown despite recent reports of association with HHV8. In order to analyze KS risk factors and value of HHV8 detection among GCIF kidney transplant patients (TP) alive betwen 1996-1998, we retrospectively compared TP who had developed post transplant KS before january 1996 with 2 matched controls (C) for age, sex, transplant year and center. Before KS diagnosis or at same follow-up for C, we analyzed: ethnic origin; number of blood transfusions, grafts and rejection episodes; immunosuppressive regimen; tuberculosis, i.v. treated fungal, CMV or herpes infections; serological status for HIV 1-2, CMV, Hepatitis B and C, Herpes; blood cell count, renal function. We searched for HHV8 viremia by PCR at time of study (ts), HHV8 antibodies (Ab) before and after the graft. Sera were tested blindly using 2 indirect immunofluorescence assays (IFA) with 2 derived HHV8(+) EBV(-) cell lines and ELISA (recombinant protein encoded by HHV8 orf 65) confirmed by western blot. We then prospectively followed HHV8 detection within KS lesions, normal skin and peripheral blood mononuclear cells (PBMC) in TP with KS onset at ts. Among 30 TP with KS 21±34 months (M) after grafting, 24 were in remission, 1 had stable KS, 5 were lost from follow-up at ts. 9 other TP had KS onset at ts. After, conditional logistic regression analysis, TP geographic origin (OR=15.1, p<0.001), antiHbc Ab (OR=5.9, p<0.005), hemoglobin <12g/dl (OR=6.3, p<0.001), lymphocytes<750/mm3 (OR=9.0, p<0.05) were KS risk factors but not use of cyclosporine (OR=0.2, p<0.05). Stepwise logistic regression showed independant increased risk with African origin (OR=50.0, p<0.001) and initial use of polyclonal antilymphocytes sera (OR=11.3, p<0.05). HHV8 Ab were detected before graft in 100% of 5 KS vs 10% of 20 C(p<0.001); after graft, in 92% of 13 KS at 74±66 M vs 34% of 23 C at 87±58 M(p<0.01). At ts, HHV8 viremia was detected in 11% of 9 (1 stable, 8 remission) KS vs 0% of 13 C (ns). Prospective study detected HHV8 sequences in recipient's KS lesions (6/6), normal skin (3/5), PBMC (7/9) and HHV8 Ab in 88% of 6 KS at KS onset with negativation of viremia in 3/3 TP with clinical resolution. This first case-control analysis identifies KS risk factors. It suggests that HHV8 Ab detection before graft is predictive of KS and viremia is related to tumor burden and evolution.

Detection of human herpesvirus-8 DNA in Kaposi’s sarcomas from iatrogenically immunosuppressed patients

Background: Kaposi’s sarcoma (KS) accounts for more than 5% of malignancies in immunosuppressed organ transplant patients (OKS). A new herpesvirus (HHV-8) was identified with high prevalence in biopsy specimens of AIDS-KS, endemic KS, and classic KS and in OKS. KS has also been associated with other underlying diseases in patients treated with corticosteroids, but this subset of KS has been reported to contain HHV-8 in only a few case reports. Objective: In this larger study, we determined the prevalence of HHV-8 in seven patients of Jewish origin in whom KS developed during immunosuppressive therapy for different primary diseases (ISKS). Methods: The study included HHV-8 DNA detection by polymerase chain reaction (PCR) coupled with Southern blot and sequence analysis as well as by in situ hybridization. Results: HHV-8 sequences were detected by PCR with confirmation by Southern blot and sequence analysis in 100% of the ISKS samples. Direct sequencing revealed several previously unknown base changes within the 208 bp region from open reading frame 26 (ORF26 208 ) of HHV-8 in ISKS. Conclusion: Ours is the largest known study describing the presence of HHV-8 in iatrogenic KS from immunosuppressed nontransplant patients and provides data of previously unknown sequence variations within the ORF26 of HHV-8 DNA. (J Am Acad Dermatol 1998;38:429-37.)

HHV-8 and AIDS-related CNS lymphoma.

Human herpesvirus 8 (HHV-8), the Kaposi's sarcoma (KS)-associated herpesvirus, is closely related to the transforming Epstein-Barr virus (EBV) and may be detected in AIDS-related, EBV-containing, body-cavity-based lymphomas. Accordingly, we analyzed 27 EBV-positive, AIDS-related CNS lymphomas for HHV-8 sequences. Only one tumor yielded HHV-8 sequences; this tumor arose in a patient with concomitant systemic lymphoma and a history of KS. We conclude that HHV-8 is unlikely to have a major role in the pathogenesis of AIDS-related, EBV-associated CNS lymphomas.

Establishment, characterization and Scid mice injection of B-cell lines harboring HHV-8 and EBV from a spleen of an HIV infected patient with multicentric Castleman's disease.

Backround: Previous studies demonstrated an association between HHV-8 and multicentric Castleman's disease (MCD) especially in HIV infected patients. Until now, persistant cell lines harboring HHV-8 with and without EBV could only be established from body-cavity-based lymphomas (BCBL). herein, we report the first establishment of HHV-8 positive cell lines from a patient with MCD. Methods: Fresh samples were obtained from an HIV infected patient after a therapeutic splenectomy for MCD. Recovered cells from disrupted spleen fragments were Ficoll® purified, maintained in RPMI 1640 and 10 % FCS and weekly passaged. HHV-8 and EBV DNA sequences were detected by PCR. Ultracentrifuged supernatent samples were tested for the presence of HHV-8 DNA sequences after DNAse treatment. Cells typing were performed by conventional flow cytometry and cytology. CB17 scid/scid mice were intraperitoneally inoculated with 15 millions of each of our cell lines (15th passage). Results: Since the 12th passage flow cytometry analysis showed the emergence of 3 lymphocytes lines with a B-cell phenotype. The presence of HHV-8 and EBV DNA sequences was demonstrated until now (26th passage). Until the 12th passage, we could also demonstrated the presence of DNAse resistant HHV-8 sequences in culture supernatent suggesting the production of encapsidated genome. Four weeks after inoculation, the PCR analysis of mice samples showed the presence of HHV-8 sequences in heart, peritoneal wash, lung. Moreover, an intraperitoneal tumor corresponding to a lymphoma harboring HHV-8 was observed in one mice. Conclusion: As far as we know, this is the first description of HHV-8 positive B-cell lines obtained from a patient with MCD. Preliminary results on Scid mice suggest that a systemic dissemination could occurred. Further characterization of these cell lines and of the Scid intraperitoneal tumor are in progress.

Human herpesvirus type 8 DNA sequences in biological samples of HIV-positive and negative individuals in Sicily.

To evaluate the circulation of a new human herpesvirus (HHV), HHV-8 or Kaposi's sarcoma (KS)-associated herpesvirus in a geographical area where a high incidence rate of classical KS was already present before the appearance of the AIDS epidemic. The study was carried out by analysing: (i) bioptic samples from classic, AIDS-associated KS, and controls; (ii) peripheral blood mononuclear cells (PBMC) from classic KS, HIV-positive subjects with and without KS and healthy HIV-negative individuals; (iii) semen samples from heterosexual HIV-positive and HIV-negative individuals affected or not by KS; and (iv) cervical swabs from HIV-negative healthy heterosexual females. All specimens were tested for the presence of HHV-8 DNA sequences by a two-step polymerase chain reaction. Positive results were obtained in 90% of bioptic samples of classic KS and in 100% of AIDS-associated KS. Viral sequences were also present in 50% of PBMC of subjects with classic KS and AIDS-associated KS, in 10% of AIDS patients without the angiosarcoma and in 11% of healthy HIV-negative individuals. Finally, HHV-8 DNA was detected in 13% of semen of HIV-negative heterosexual individuals and in 10% of AIDS patients without KS. Both PBMC and ejaculates from the same individual gave positive results. No HHV-8 sequences were found in cervical swabs. HHV-8 is widespread in the general population in Sicily since it was detected in PBMC and semen of heterosexual HIV-negative individuals and is not found only in high-risk groups. The viral load appears to be more elevated in a high-risk population and it may be ascribed to a viral reactivation. The higher incidence rates of KS in Sicily compared with northern Italy and other European countries might be related to the presence of HHV-8 in the general population.

Human herpesvirus type 8 DNA sequences in cell-free plasma and mononuclear cells of Kaposi's sarcoma patients.

Human herpesvirus (HHV) type 8 has been detected in both classical and AIDS-related Kaposi's sarcoma, body-cavity lymphomas, and other types of tumors. HHV-8 has also been detected in DNA from peripheral blood mononuclear cells (PBMC) of some Kaposi's sarcoma patients and more readily in B cell fractions derived from panned cell subpopulations. Two patients were followed using several methods; in situ hybridization, solution-based polymerase chain reaction (PCR), and in situ PCR. HHV-8 was intermittently detected in plasma, and detection correlated with detection in PBMC. In situ PCR demonstrated HHV-8 sequences in both peripheral blood B lymphocytes and, to a lesser extent, T lymphocytes. HHV-8 may undergo periods of viremia while at other times it is undetectable and infects circulating B cells and some T cells.

Distribution of human herpesvirus-8 sequences throughout the spectrum of AIDS-related neoplasia.

AIDS frequently associates with certain malignancies, including Kaposi's sarcoma, non-Hodgkin's lymphoma (NHL), and anogenital neoplasia. In this study we aimed to define the frequency of infection by human herpesvirus (HHV)-8 throughout the spectrum of AIDS-related neoplasia in Europe. A tumour panel representative of the distinct types of AIDS-related neoplasms was tested for the presence of HHV-8 DNA sequences. Autologous uninvolved tissues were also tested in selected cases. The presence of HHV-8 DNA sequences was assayed by a combination of polymerase chain reaction followed by oligohybridization and Southern blot hybridization of genomic DNA with an HHV-8-specific probe. HHV-8 sequences were detected in 100% of AIDS-related Kaposi's sarcoma (all 35 cases). Among AIDS-related NHL, HHV-8 sequences selectively clustered with body-cavity-based lymphomas (BCBL; all three cases), although they were consistently negative in small non-cleaved cell lymphomas (none in 18 cases), diffuse large cell lymphomas (none in seven), or anaplastic large cell lymphomas (none in three). No HHV-8 sequences were found in cases of anogenital neoplasia (out of 14) or Hodgkin's disease (out of three). HHV-8 DNA sequences were also positive in the uninvolved skin of all six AIDS-related Kaposi's sarcoma patients, but not in the circulating lymphocytes of a BCBL patient. Positivity for HHV-8 sequences occurred in patients belonging to all major AIDS risk categories. These data confirm that HHV-8 sequences associate at high frequency with selected types of AIDS-related neoplasia, namely Kaposi's sarcoma and BCBL, although they are consistently absent in other types of AIDS-NHL and AIDS-related anogenital neoplasia.