Use of HPV Insertional Mutation as a Molecular Marker of Circulating Tumour DNA
Emmanuelle Jeannot, Maura Campitelli, Martine Peter, Stephanie Saada, Virginie Fourchotte, Paul Cottu, Olivier Lantz, Jean-Yves Pierga, Jerome Couturier, Xavier Sastre-Garau. Institut Curie, Paris, France, Metropolitan
Background: The detection of circulating tumour DNA requires the identification of a specific molecular motif on primary tumour and is hampered by the low concentration of circulating tumour DNA (ct DNA) in circulating non tumour DNA.
We looked whether the genomic rearrangement secondary to the clonal integration of HPV DNA sequences into the host genome may be used as a highly specific marker to detect ct DNA in patients with HPV-associated cervical carcinoma.
Design: Nine cases of HPV16-associated invasive cervical carcinoma, corresponding to stage I (2 cases) to stage IV (one case) were analyzed. HPV integration locus was characterized using the Detection of Integrated Papillomavirus Sequenced (DIPS-PCR) method. Cell free circulating DNA was isolated from serum specimens taken before treatment in each case. Sequential serum specimens taken during the course of the disease were also available for 2 of these 9 cases. A PCR-assay was designed to specifically amplify the cell-viral DNA junctions and the minimal amount of tumour DNA detectable with this assay was determined in reconstruction experiments using serial dilutions of tumour DNA into Tris-EDTA buffer.
Results: HPV integration sites were located at different loci in the 9 cases. According to reconstruction experiments, the PCR-assay was able to detect a concentration as low as 0.5 pg/ml of tumour DNA in Tris-EDTA buffer. Using the 9 specific assays, ct DNA was detected in 7 of the 9 serum specimens with concentrations ranging from 0.03 to 42 pg/200 µl. Comparison with clinical data showed that two of the three negative cases corresponded to stage I carcinoma with tumor size ≤ 15 mm. In both cases with sequential specimens, a dynamic of ct DNA according to the tumor mass was observed during treatment and at time of relapse.
Conclusions: HPV insertional mutations can be used to detect ct DNA with a high level of specificity in patients with HPV-associated carcinoma.
Category: Gynecologic & Obstetrics
Wednesday, March 21, 2012 1:00 PM
Poster Session VI # 192, Wednesday Afternoon