HPV16‐E6 mRNA is superior to cytokeratin 19 mRNA as a molecular marker for the detection of disseminated tumour cells in sentinel lymph nodes of patients with cervical cancer by quantitative reverse‐transcription PCR

Abstract

About 10-15% of patients with cervical cancer suffer from recurrence despite histologically negative lymph nodes (pN0). Occult micrometastases or small tumour cell clusters may contribute to disease outcome. The aim of this study was to compare at the RNA level 2 known tumour-associated genes, HPV16-E6 and cytokeratin 19 (CK19), as molecular markers for the detection of disseminated tumour cells. Real-time reverse transcription PCR technology was used to quantify gene expression in histologically positive and negative sentinel lymph nodes (SLN) from 70 patients with cervical cancer. Lymph nodes from noncancer patients were used as controls. Calculated copy numbers were normalised to the geometric average of the most stable housekeeping genes. We observed a good correlation (R 5 0.915) between the expression of both markers in SLN with histologically confirmed metastases. However, marker gene expression differed considerably in histologically negative nodes: CK19 transcripts were detected in 90 of 112 SLN (80.4%), whereas only 38 nodes (33.9%) were positive for HPV16 E6 mRNA. In particular, 62 of 74 SLN, which were negative by histology, and HPV16 E6 mRNA expressed CK19 mRNA. Moreover, 8 of 10 lymph nodes from noncancer patients expressed CK19 mRNA. Systematic errors due to RNA degradation or incomplete cDNA could be ruled out. It is concluded that HPV16 E6 mRNA is more specific and more sensitive for the detection of tumour cells in SLN than CK19 mRNA. The specificity of CK19 is limited because of low level expression in uninvolved pelvic lymph nodes. ' 2007 Wiley-Liss, Inc.Key words: cervical cancer; disseminated tumour cells; molecular markers; HPV16-E16; cytokeratin 19; quantitative RT-PCR Cervical cancer is the second most common cancer among women world wide, accounting for 493,000 new cases in 2002. 1 The incidence rates vary considerably between different geographic regions, which reflect in part the lack of screening programs in developing countries. Despite the advancement in early diagnosis, improved surgical techniques and adjuvant therapies, approximately 40% of cervical carcinoma patients in countries with a high standard of health care die from the disease. 1 The majority of these deaths is due to metastases. 2 Different parameters such as lymph node metastases, tumour grade, lymph-vascular space involvement (LVSI) and surgical margins are prognostic factors for cervical cancer, with lymph node status as the most important one. 3,4 Patients with histologically uninvolved lymph nodes (pN0) have a better prognosis than women with histologically confirmed lymph node metastasis (pN1) showing 5 year survival rates of 88-93% and 40-63%, respectively. 5-7 Nevertheless 15% of FIGO stage IB patients with pathologically staged negative lymph nodes (pN0) suffer from recurrence. 8 The significance of micrometastases as a predictive factor for recurrent disease is the focus of several recent and ongoing studies. Extensive serial sectioning of lymph nodes that were judged to be free of disease by routine diagnos...

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

“…It is important to keep in mind, however, that cytokeratin is found in the lymph nodes and blood of healthy individuals [35]. In addition, the technique for detecting cytokeratin is reportedly nonspecific and inferior to HPV-based assays for the detection of metastatic foci in patients with cervical cancer [36].…”

Correlation between load of HPV 16 DNA in cervical cancer and HPV 16 DNA in lymph nodes

“…The E6 and E7 genes exert their transforming function by interrupting cell differentiation and inducing DNA synthesis. They accomplish this by interacting with the cellular tumor suppressor gene products p53 and pRB [4] . Zheng et al [5] found that the episomal form of HPV16 was detected in most CIN I, but that the episomal form and the integrated form coexisted in 42.9% of CIN I.…”

Circulating tumor cells: hope to diagnose and treat metastatic cancer

“…Similarly, TWIST and vimentin are considered as diagnostic markers for hepatocellular carcinoma. Häfner et al [14] have isolated CTCs from metastatic cervical cancer patients and evaluated levels of HPV16-E6 mRNA by real-time PCR, as a more sensitive molecular marker than CK19 mRNA. At the same time, Kuhlmann et al [15] have shown that ERCC1+ CTCs can predict platinum resistance therapy in ovarian cancer.…”