Abstract
Increasing evidence demonstrated that inactivation of tumor suppressor genes (TSGs) by aberrant promoter methylation is an early event during carcinogenesis. Aiming at developing early diagnostic or prognostic tools for various tumors, we took an EBV-associated tumor, nasopharyngeal carcinoma (NPC), as a model and developed a powerful assay based on “multiplex methylation specific-PCR (MMSP)”. The MMSP assay was designed to detect tumor-specific methylation status of several NPC-related genes and was capable of acquiring multiplex information simultaneously through a single PCR reaction with the tiny tumor DNA derived from the direct body fluid close to the primary tumor. In this study, we collected paired nasopharyngeal (NP) swabs and NPC biopsies from 49 NPC patients and twenty noncancerous controls. A panel of markers including two EBV, and two cellular TSG markers were applied in this NPC-specific-MMSP assay. We optimized the working condition of MMSP so that it provides information equal to that from the corresponding separate PCRs. The results showed that MMSP patterns of NPC swab were largely consistent with those of corresponding biopsies and significantly distinguished themselves from those of 20 noncancerous volunteers. Among the 69 samples (49 NPCs and 20 normal controls), the sensitivity of detecting NPC from NP swabs is 98%. The specificity is as high as 100%. In conclusion, being characterized by its noninvasiveness, high reproducibility and informativeness, MMSP assay is a reliable and potential diagnostic tool for NPC. It paves the way for the development of population screening and early diagnosis approaches for various tumor types.
Highlights
Nasopharyngeal carcinoma (NPC) is the second most common cancer in southern China and constitutes the main menace in this area
The feasibility of Multiplex Methylation Specific PCR (MMSP) In order to test the feasibility of MMSP to achieve the same readout as those from single methylation specific PCR (MSP), we used Namalwa cell line as a testing material
There was an excellent concordance with the relative density of the five bands between MMSP and separate PCR reactions
Summary
Nasopharyngeal carcinoma (NPC) is the second most common cancer in southern China and constitutes the main menace in this area. Patients in stage I and II disease have a significantly longer overall survival compared with those in stage III and IV. Early diagnosis for NPC is essential in achieving a satisfactory therapy effect. Because of the non-specific local symptoms and the un-convenience of a fully clinical examination of the nasopharynx, the majority of NPC patients are only diagnosed when the tumor has reached an advanced stage [2]. 70% of newly-diagnosed NPC patients presented as advanced diseases with a poor prognosis [2,3]. To find biomarkers for detecting early stage NPC and monitoring residual or recurrent tumor would help greatly in elevating survival probability and the choice of subsequent therapy
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