Detection of human papillomavirus DNA in peri-tumor tissues and pelvic lymph nodes as potential molecular marker of micrometastasis in cervical cancer

Abstract

BackgroundThe association between high risk human papillomaviruses (HPV) and cervical cancer has been firmly established. HPV genome is present in nearly all cases of cervical cancer and detection of viral DNA could therefore be used as a surrogate marker of micrometastasis in peri-tumor tissues and lymph nodes.MethodsWe analyzed primary cervical carcinomas, peri-tumor biopsies and pelvic lymph nodes in 20 women with invasive cancer (FIGO stage I-II) who underwent radical pelvic surgery and lymphadenectomy. HPV DNA was searched by broad spectrum PCR in 142 DNA samples extracted from paraffin embedded tissues. Viral genotypes were identified by direct sequencing analysis.ResultsHPV DNA sequences were identified in all available primary cervical tumors (n = 15). The most common genotype was HPV16 (60 %), followed by HPV18 (20 %), HPV35 (7 %), HPV45 (7 %) and HPV66 (7 %). Seven out of 20 (35 %) women had metastatic spread in peri-tumor tissues and/or lymph nodes, as determined by histology. HPV DNA was detected in all histological positive samples as well as in 16 and 25 % of histological negative peri-tumor tissues and lymph nodes, respectively. Three out of 20 (15 %) women without histological evidence of metastatic spread had HPV-positive lymph nodes. HPV genotype was found always concordant between primary tumor and metastatic lesions. The remaining 10 women (50 %) were histology and HPV-negative in all peri-tumor biopsies and lymph nodes analyzed.ConclusionsEvaluation of HPV DNA in peri-tumor tissues as well as pelvic lymph nodes could be a sensitive marker to identify micrometastasis or isolated tumor cells and to monitor the risk of disease recurrence in women with cervical cancer.

Distinct profiles of TERT promoter mutations and telomerase expression in head and neck cancer and cervical carcinoma

“…Genomic DNA was extracted from approximately 10 mg of fresh frozen tissues and 5 × 10 6 cultured cells by digestion with proteinase K (150 μg per mL at 37°C for 10 hr) in 100 μL of lysis buffer (10 mM Tris–HCl pH 7.6, 5 mM EDTA, 150 mM NaCl, 1% SDS), followed by extraction with phenol–chloroform–isoamyl alcohol (25:24:1) and ethanol precipitation in 0.3 M sodium acetate (pH 4.6) . DNA was extracted from paraffin embedded tissues as described previously . Total RNA was extracted from all fresh frozen samples and cultured cells using RNeasy MiniKit (Qiagen, Hilden, Germany).…”

“…HPV detection was carried out by nested PCR amplifying 30–300 ng of genomic DNA with MY09/MY11 primer pairs for the outer reaction and MGP primer system for the inner reaction in 50 μL reaction mixture containing 5 μL of outer reaction . The method has been previously validated as proficient for the detection of twelve high risk HPV genotypes being able to detect 50 genome equivalents (GE) of HPV 16 and HPV 18 DNA in 5 μL sample with a specificity above 97% through a reference‐test proficiency panel as part of the 2010 WHO LabNet study for validating HPV genotyping tests .…”

Value of Intraoperative Cytological and Pathological Sentinel Lymph Node Diagnosis in Fertility-Sparing Trachelectomy for Early-Stage Cervical Cancer

“…The HPV detection system, in which E6/E7 mRNA is not used, results in a high false-positive rate, explained by the existence of HPV DNA from destroyed cancer cells [39]. Although evaluation of HPV DNA in lymph nodes could be a sensitive marker not only to identify micrometastasis or isolated tumor cells but also to monitor the risk of cancer recurrence [40,41], further prospective studies are required to confirm its value for this purpose.…”

A new marker based on risk stratification of human papillomavirus DNA and tumor size to predict survival of locally advanced cervical cancer

“…They found that HPV DNA was a strong independent prognostic factor for disease-free survival 9. However, the status of HPV DNA replication is associated with multiple genetic alterations which may result in cancer progression,10 suggesting that different HPV viral loads may impact differently on distant metastasis-free survival or overall survival.…”