PF512 NEXT‐GENERATION SEQUENCING‐BASED CLONALITY ASSESSMENT OF IMMUNOGLOBULIN GENE REARRANGEMENTS DISTINGUISHES RELAPSE FROM SECOND PRIMARY CLASSICAL HODGKIN LYMPHOMA

Abstract

Background:Classic Hodgkin's lymphoma (cHL) has a high cure rate with current therapy, however relapse still occurs in 5%>10% of early stage and 15%>30% of advanced cHL. Interestingly, case reports and small series have shown that some of these relapses appear to be a new primary cHL. Assessment of clonal relationship between primary diagnosis and relapsed cHL samples after treatment will distinguish true relapse from second primary lymphoma. Clonality detection in cHL using conventional clonality assays has thus far been severely hampered by the low frequency of Hodgkin and Reed‐Sternberg (HRS) cells and limited DNA quality obtained from formalin‐fixed paraffin‐embedded (FFPE) material. Within the EuroClonality‐NGS Working Group, we have developed a novel approach to detect immunoglobulin (IG) heavy chain (IGH) and k light chain (IGK) gene rearrangements.Aims:The objective or our study is to determine the clonal relationship between HL diagnosis and relapse samples to assess the incidence of second primary malignancies in HL recurrences.Methods:We have collected 70 paired primary and relapse cHL cases. In 38/70 cases relapse occurred within three years after diagnosis, in 32/70 cases relapse occurred later than three years after diagnosis. EuroClonality‐NGS gene‐specific IGH‐VJ‐FR3, IGH‐DJ, IGK‐VJ and Intron‐Kde primer sets for small amplicon sizes were used to perform next‐generation sequencing (NGS)‐based clonality analysis with Ion Torrent PGM. Bioinformatic analyses with the interactive web‐based immunoprofiler ARResT/Interrogate allowed run/sample quality control followed by accurate identification of clonotypes.Results:We show that NGS‐based clonality assessment is highly suitable for detecting IG rearrangements in FFPE tissue specimens, including HL tissue specimens. Our preliminary data analysis of 7 paired diagnosis and relapse samples demonstrates the presence of identical clonotypes in 2 cases, while distinct clonotypes were observed in 3 other cases suggesting a new primary lymphoma. In the remaining 2 cases no specific clonotype could be identified in either diagnosis and/or relapse. The analysis of additional cHL recurrences by NGS‐based clonality assessment will reveal the true incidence of clonally unrelated lymphoma in relapsed cHL.Summary/Conclusion:This study is an important step towards establishment of NGS‐based clonality assessment in clinical practice for cHL, and eventually the improvement of therapeutic management of second primary cHL.