Human papillomavirus DNA in plasma of patients with cervical cancer

BACKGROUND: Human papillomavirus (HPV) is a crucial etiological factor for cervical cancer (CC) development. From a diagnostic view-point, the consistent presence of HPV in CC allows the viral DNA to be used as a genetic marker. The aims of this study were to evaluate the presence, physical status and clinical significant of HPV DNA in circulation of CC patients. RESULTS: Whereas 6 out of 50 (12%) HPV positive CC patients revealed plasma HPV DNA, it was detected in none of 20 normal controls or 13 HPV negative CC cases. The plasma DNA exhibited an HPV type identical to the HPV in the primary tumors and the DNA from both sources was integrated into host genome. Interestingly, several findings suggested an association between plasma HPV DNA and metastasis. First, three of the HPV DNA positive cases were CC patients with clinical stage IVB or recurrence with distance metastases (P = 0.001, RR = 15.67). Second, the amount of plasma HPV DNA from metastatic patients to be three times more than three other patients without metastases. Finally, the later cases had tendency to develop recurrence distant metastases within one year after complete treatment when compared with other HPV associated CC patients with the same stage but without the present of plasma HPV DNA. CONCLUSIONS: The plasma HPV DNA originated from the CC, was associated with metastasis and could be used as a marker representing the circulating free CC DNA

Cyclin A1 promoter hypermethylation in human papillomavirus-associated cervical cancer

BACKGROUND: The aim of this study was to evaluate epigenetic status of cyclin A1 in human papillomavirus-associated cervical cancer. Y. Tokumaru et al., Cancer Res 64, 5982-7 (Sep 1, 2004)demonstrated in head and neck squamous-cell cancer an inverse correlation between cyclin A1 promoter hypermethylation and TP53 mutation. Human papillomavirus-associated cervical cancer, however, is deprived of TP53 function by a different mechanism. Therefore, it was of interest to investigate the epigenetic alterations during multistep cervical cancer development. METHODS: In this study, we performed duplex methylation-specific PCR and reverse transcriptase PCR on several cervical cancer cell lines and microdissected cervical cancers. Furthermore, the incidence of cyclin A1 methylation was studied in 43 samples of white blood cells, 25 normal cervices, and 24, 5 and 30 human papillomavirus-associated premalignant, microinvasive and invasive cervical lesions, respectively. RESULTS: We demonstrated cyclin A1 methylation to be commonly found in cervical cancer, both in vitro and in vivo, with its physiological role being to decrease gene expression. More important, this study demonstrated that not only is cyclin A1 promoter hypermethylation strikingly common in cervical cancer, but is also specific to the invasive phenotype in comparison with other histopathological stages during multistep carcinogenesis. None of the normal cells and low-grade squamous intraepithelial lesions exhibited methylation. In contrast, 36.6%, 60% and 93.3% of high-grade squamous intraepithelial lesions, microinvasive and invasive cancers, respectively, showed methylation. CONCLUSION: This methylation study indicated that cyclin A1 is a potential tumor marker for early diagnosis of invasive cervical cancer

Cytotoxic function of gamma delta (<img src='/image/spc_char/gamma2.gif' border=0>/<img src='/image/spc_char/delta1.gif' border=0>) T cells against pamidronate- treated cervical cancer cells </span></span>

597-605The cytotoxic function of polyclonal expanded / T cells against pamidronate-treated cervical cancer cells in vitro and in vivo were determined. The / T cells were isolated and purified from PBMCs by using miniMACS and were later treated with 10 μM pamidronate. The expansion of / T cells was 15 times more than the non-stimulated cells. Among the expanded / T cells, 47% were V9/V2 T cells with a purity of 87%. Analyzing the cytotoxic function of / T cells against 3 cervical cancer cells in vitro by LDH cytotoxicity test revealed that the killing efficacy increased if the cervical cancer cells (HeLa, SiHa and CaSki) were pretreated with pamidronate. The presence of CD107 on / T cells indicated the degranulation of perforin and granzyme pathway is one of the mechanisms used by the / T cells to kill cancer cells. The killing ability of / T cells against cancer cells in vivo was preliminary assessed by using mouse baring HeLa cells. The results demonstrated that / T cells induce apoptosis in tumor cells. Our study supports the usefulness of / T cells in future development of immunotherapy for cervical cancer. </span

Mouse acquired HPV tumor using dorsal skin-fold window chamber

327-332Human papillomavirus (HPV) plays important role in developing several types of cancer especially cervical cancer. In order to understand the viral pathogenesis, the animal model of HPV infection is very necessary. This communication reports establishment of an animal model carrying implanted HeLa cells, a human cervical cancer cell line via dorsal skin-fold window chambers. Nude mice were divided into 4 groups; each group contained different amount of HeLa cells, 2.5×105, 5×105, and 1×106 cells, and cell free medium (control), respectively. The results showed that even using the low number of HeLa cells (2.5×105), the tumor microvasculature was developed at 2 weeks after implantation with the enlarged tumor margin which then progressed to tumor mass in the following week. The existing tumor was confirmed to be HeLa-cell type by PCR, in situ hybridization, and HPV genotyping. By using linear regression analysis, it indicated that means of tumor size from each group significantly increased in relation to number of HeLa cells used (R2 = 0.98, y = 0.1171x+4.35). This mouse model will be useful for the further HPV studies particularly anti-cancer drugs efficacy