Quantitative RT-PCR Assay for Detecting Lymph Node Metastasis in Endometrial Cancer: A Preliminary Study

Abstract

Objective: The detection accuracy of sentinel lymph node (SLN) metastasis by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) for endometrial cancer (EC) remains unclear and was assessed in this preliminary study. Methods: We studied primary cancer tissues and pelvic lymph nodes (PLN) from 105 patients with EC. qRT-PCR assay was performed to determine the copy numbers of CK19 mRNA in EC tissues, and negative and positive LN samples. Further, qRT-PCR results were compared with pathological findings. Results: CK19 mRNA expression was detected in 98% (104/106) of the tumors, with a median copy number of 3.0 × 10⁵/μL. Twelve LN were diagnosed as positive by pathological examination. The median copy number of CK19 mRNA for positive and negative LN was 8.1 × 10⁴/μL and 90.4/µL, respectively. CK19 mRNA expression was higher in pathologically positive LN than in pathologically negative LN (p < 0.01); the pathological and qRT-PCR findings showed no discrepancy. When the cutoff value was set at 4,500 copies/µL, qRT-PCR assay using CK19 mRNA exhibited high sensitivity and specificity. Conclusions: Our results demonstrated that qRT-PCR assay, using CK19 mRNA, exhibits a high accuracy for detecting LN metastasis in EC and represents a useful alternative to conventional pathological diagnosis of EC.

Cytokeratin Expression Pattern in Human Endometrial Carcinomas and Lymph Nodes Micrometastasis: a Mini-review

“…There are manuscripts describing advanced molecular techniques (for example qRT-PCR) in detecting EC lymph node micrometastases [94][95]. These techniques are even more sensitive than IHC, but they also carry high false-positive results [94][95].…”

“…There are manuscripts describing advanced molecular techniques (for example qRT-PCR) in detecting EC lymph node micrometastases [94][95]. These techniques are even more sensitive than IHC, but they also carry high false-positive results [94][95]. Interestingly, Pappa et al [94] assumed that "..qRT-PCR exhibits a better diagnostic accuracy compared with IHC (for the detection of lymph node micrometastasis in cervical, endometrial and vulvar cancer), while CK19 displays a consistent pattern of detection compared to carbonic anhydrase 9…".…”

Sentinel lymph node processing in gynecological cancer histopathology and molecular biology

“…Numerous candidate genes (over 24 genes for endometrial carcinoma) with specific expression in carcinomas and no expression in lymphatic or soft tissues (17) revealed cytokeratin 19 (CK19) m-RNA as the best target to use for metastatic carcinoma cells. Togami et al (18) used qRT-PCR to detect and quantify CK19 mRNA in endometrial carcinomas and found 104/106 (98%) positive cases. In this study qRT-PCR of 12 pelvic lymph node (PLN) metastases reached best sensitivity and specificity without discrepancies between pathology report and molecular test when "positivity" of PLNs was determined above 4,500 copies/µL.…”