Telomere attrition predominantly occurs in precursor lesions during in vivo carcinogenic process of the uterine cervix.

Although human papillomavirus (HPV) has been defined as the pathogen for cervical carcinomas, molecular events underlying the oncogenic process are unclear. As telomere dysfunction-mediated chromosomal instability and telomerase activation have been suggested as key events in carcinogenesis, we dissected the dynamic changes in telomere length, checkpoint response, and temporal profile of telomerase expression during the evolution from precursor lesions (cervical intraepithelial neoplasia, CINs) to invasive cancers of the uterine cervix in sequential samples from 16 patients. Telomeres were significantly shortened in all CIN samples and no further substantial attritions occurred in most cases with the acquisition of malignant phenotype. Very short telomeres were coupled with constitutive activation of the DNA damage response pathway (Chk2 phosphorylation) and increased cellular proliferation in those cervical specimens. Telomerase reverse transcriptase (hTERT) expression was preferably induced at advanced CINs or invasive cancers. The present finding demonstrates that excessive telomere shortening predominantly occurs in the early carcinogenesis of the uterine cervix largely prior to telomerase activation. Widespread over-erosion of telomeres or telomere dysfunction in very early stages of cervical tumorigenesis might fuel transformation processes by driving chromosomal instability.

Predictive significance of the alterations of p16INK4A, p14ARF, p53, and proliferating cell nuclear antigen expression in the progression of cervical cancer.

PURPOSE: The purpose of this research was to evaluate the clinical significance of p16INK4A, p14ARF, p53, and proliferating cell nuclear antigen (PCNA) expression in tumor progression of cervical cancer. DESIGN: Seventeen patients (40 samples) with consecutive cervical lesions from normal squamous epithelium, inflammation of the cervix to cervical intraepithelial neoplasm (CIN) and invasive cervical squamous cell cancer (SCC), or from CIN to SCC were collected for this study. Expression of p16INK4A, p14ARF, p53, and PCNA were detected by immunohistochemistry on paraffin-embedded sections. Human papillomavirus DNA was detected simultaneously with PCR and typed according to its DNA sequence. RESULTS: p16INK4A overexpression was significantly higher in CIN (75%) and in SCC (75%) than in normal or inflammation of the cervix (12.5%; P < 0.01, P < 0.05, respectively). The positive rate of p14ARF expression was higher in SCC (83%) than in normal/inflammation of the cervix (25%; P < 0.05). PCNA expression was negative in normal or inflammation of the cervix, but an increased in expression was seen in 63.2% in CIN and 100% in SCC (P < 0.01, P < 0.05). When the time interval for disease progression from initial biopsy to CIN 3 or invasive cancer was compared with states of p16INK4A expression, cases stained positive for p16INK4A progressed within 64.2 months as compared with 122.3 months among those stained negatively (P < 0.01). Cases with increased p14ARF expression also had a short time interval for disease progression of 78.8 months as compared with 108.3 months in cases that were p14ARF negative. Cases with stable or decreased p53 expression had the shortest time interval for progression of 32.3 months in contrast to cases with no p53 expression (113.9 months). However, cases with increasing p53 expression progressed within 60.8 months. CONCLUSIONS: Our results suggested that altered states of p16INK4A, p14ARF, p53, and PCNA may be valuable markers to predict the progression of cervical neoplasia.

p16INK4a and laminin-5gamma2 chain expression during the progression of cervical neoplasia.

p16INK4a, laminin-5gamma2 chain, and PCNA were investigated to compare the expression levels in relation to histological diagnosis and time for progression. The material consisted of 37 normal cervical tissues, 35 with different grades of CIN, and 11 invasive cervical cancers. Our results showed a reduction of basement membrane staining for laminin-5gamma2 chain from 78.4% in normal squamous epithelium to 27.8% in CIN3 (p < 0.001). The intracytoplasmic staining for laminin-5gamma2 chain increases with severity of lesion. The same trend was observed with p16INK4a and PCNA expression (p < 0.001). Co-expression of p16INK4a and PCNA was seen in 85.7% of samples. Cases that were laminin-5gamma2 chain BM - /p16INK4a+/PCNA+ have the shortest interval time (average: 46.8+/-36.3 months) for progression, while cases with laminin-5gamma2 chain BM + /p16INK4a-/PCNAPCNA--have the longest time interval (average: 110.2+/-52.7 months) (p < 0.05). Thus co-expression of p16INK4a, laminin-5gamma2 chain and PCNA may be valuable for the prediction progression of cervical neoplasia.

p16INK4A and p14ARF expression pattern by immunohistochemistry in human papillomavirus-related cervical neoplasia.

Human papillomavirus is known to play an important etiological role in the genesis of cervical cancer, but only a very small proportion of infected women develop invasive cervical cancer. The purpose of cervical cancer prevention is early diagnosis of its precursors. The molecular detection of human papillomavirus DNA as a diagnostic test to cervical carcinogenesis gave a low positive predictive value as compared to the use of biomarkers. p16INK4A and possibly p14ARF have been proposed as putative surrogate biomarkers that would allow identification of dysplastic cervical epithelia. Serial consecutive biopsies representing normal cervical epithelium to cervical intraepithelial neoplasia and/or invasive cervical cancer were stained with immunohistochemistry for p16INK4A, p14ARF and proliferating cell nuclear antigen. The positive rates of these markers were significantly higher in cervical intraepithelial neoplasia and in squamous cell carcinoma than in normal cervix (P<0.01). No significant difference was noted between lesions progressing from cervical intraepithelial neoplasia to squamous cell carcinoma for both p16INK4A and p14ARF expression (P>0.05). For both biomarkers, nuclear staining was predominantly seen. However, the cytoplasmic stain of p16INK4A increased with disease progression and the pattern of expression varied between different tumors and its location within the lesion. Both nuclear and cytoplasmic staining with p16INK4A and p14ARF of affected epithelial cells were considered positive. In the adjacent normal tissue to cervical neoplasia, the positive rates of p16INK4A, p14ARF and proliferating cell nuclear antigen expression were higher than those found distant to these lesions but the findings did not reach statistical significance. No correlation was seen between the human papillomavirus types detected and the expression of p16INK4a and p14ARF. In conclusion, overexpression of p16INK4A and p14ARF act as potential biomarkers for cervical cancer progression from premalignant lesions.

Gelatinases A and B (MMP-2 and MMP-9) in endometrial cancer-MMP-9 correlates to the grade and the stage.

OBJECTIVE: Matrix metalloproteinases (MMPs) have been linked to aggressive behavior in several malignancies. Gelatinases (MMP-2 and MMP-9) in particular are prognostic factors in many adeno- and epithelial cancers. However, no conclusive data exist concerning the role of gelatinases in endometrial cancer. METHODS: Eighty-eight patients with endometrial cancer, treated between 1988 and 1993 in Umeå University Hospital, were included. MMP-2 and MMP-9 proteins were analyzed immunohistochemically from paraffin-embedded tissues by using specific monoclonal antibodies. The staining results were compared to the clinical data. RESULTS: Fifty-two percent of the cases were positive for MMP-9 and 72% for MMP-2. The overexpression of the proteins of either MMP-2 or MMP-9 was associated with poor survival. The predictive value of MMP-2 expression was most distinct in stage I cancers. An association was found between the positivity of MMP-2 and MMP-9. Only 3% of the cases were highly positive for both gelatinases and 18% of the cases were negative for both MMPs. Both gelatinases correlated to the histological grade. MMP-9 also correlated to the clinical stage of the disease, whereas MMP-2 did not. There was no apparent association with either depth of invasion, menopausal status, or the age of the patient. CONCLUSION: MMP-2 and MMP-9 could serve as markers for the clinical behavior of endometrial cancer. They may further be linked to a tendency to cancer relapse. Thus, these gelatinases may turn out to be potentially useful in decision making about the need for an adjuvant treatment.

Genetic alterations in cervical carcinomas: frequent low-level amplifications of oncogenes are associated with human papillomavirus infection.

The development of cervical carcinoma is closely associated with HPV infection. However, other genetic alterations also play an important role. In this study, we analyzed copy number alterations of several oncogene loci in a panel of 84 cervical tumors. Sixty-five (77%) tumors were HPV DNA-positive, and most were infected with type 16 or type 18 or both. The oncogenes studied include PIK3CA at 3q26.3, TERT at 5p15.33, C-MYC at 8q24, CCND1 at 11q13.3, ERBB2 at 17q21.2 and locus region 20q13.2. Amplification of 1 or more genes was detected in 55 (65%) cases using interphase FISH. PIK3CA was amplified in 43% of tumors, followed by TERT (33%), 20q13.2 (30%), ERBB2 (29%), C-MYC (25%) and CCND1 (12%). Most tumors showed low-level amplification with 3-7 copies of these genes, and complex changes involving 3 or more genes occur more frequently in tumors at advanced stages. Increased protein expression of c-erbB2 and c-myc was observed in tumors with the corresponding gene amplification. Oncogene alterations were found more often in HPV-infected cases, particularly for C-MYC and TERT. These findings indicate that HPV-associated cervical carcinomas bear frequent alterations of these genes, which may have critical biologic impact on the development and progression of carcinoma of the uterine cervix.

Amplification of the telomerase reverse transcriptase (hTERT) gene in cervical carcinomas.

The expression of telomerase reverse transcriptase (hTERT), the catalytic component of the telomerase complex, is required for activation of telomerase during immortalization and transformation of human cells. However, the biochemical and genetic mechanisms governing hTERT expression remain to be elucidated. In the present study, we examined hTERT amplification as a potential genetic event contributing to telomerase activation in cervical carcinomas. An amplification of the hTERT gene was found in 1/4 cervical cancer cell lines and 21/88 primary tumor samples derived from the patients with cervical carcinomas. An increase in the hTERT copy number was significantly correlated with higher levels of hTERT protein expression. Moreover, the hTERT alterations with the enhanced hTERT expression were exclusively observed in those tumors with high-risk human papillomavirus infection. Taken together, the hTERT gene amplification, directly or indirectly targeted by human papillomavirus, may be one of the driving forces responsible for upregulation of hTERT expression and activation of telomerase in cervical cancers.

A population-based prospective study of Chlamydia trachomatis infection and cervical carcinoma.

Persistent human papillomavirus (HPV) infection is an established cause of cervical cancer, but the role of other sexually transmitted agents, most notably Chlamydia trachomatis, has not been well defined. The women participating in the population-based cervical cancer screening program in Västerbotten county of Northern Sweden were followed up for up to 26 years to identify 118 women who developed cervical cancer after having had a normal Pap smear (on average 5.6 years later; range 0.5 months-26 years). As controls, we selected another 118 women who were matched by birth cohort, time-point of sampling of the baseline normal smear and who had a normal smear at the time when the corresponding case was diagnosed with cancer. The Pap smears and cervical cancer biopsies were analyzed by PCR for C. trachomatis DNA and for HPV DNA. At baseline, C. trachomatis DNA was present in 8% of cases but not among any one of the controls. The relative risk for cervical cancer associated with past C. trachomatis infection, adjusted for concomitant HPV DNA positivity, was 17.1 (95% CI 2.6-infinity). The presence of C. trachomatis and of HPV were not interrelated. Whereas C. trachomatis was primarily found in specimens taken many years before cancer diagnosis, HPV DNA was associated with a short lag time before cancer diagnosis. Whereas most women who were HPV DNA-positive in the prediagnostic smear were also positive for the same virus in the cervical cancer biopsy, none of the women were positive for C. trachomatis in both the prediagnostic smear and in the subsequent cervical cancer. In conclusion, a prior cervical C. trachomatis infection was associated with an increased risk for development of invasive cervical cancer.