The Classical Illegitimate Switch Translocation Model Is Unable To Account for at Least Half of the Translocation Breakpoints from t(4;14)(p16;q32) Multiple Myeloma.
Authors: Keats Jonathan J. , Strachan Erin , Belch Andrew R. , Pilarski Linda M. 
Published: April 2020

Abstract
Translocations involving the IgH locus are one of the most common genetic abnormalities observed in multiple myeloma (MM). Unlike several hematological malignancies, MM IgH translocations involve multiple partner chromosomes. Although IgH translocations are not unique to MM, the molecular anatomy of the translocations appears to be different from that observed in most B-cell malignancies. In general, the breakpoints occur within the switch regions of the IgH locus and the translocations appear to result from illegitimate class switch recombination (CSR) events. Previous analysis of the breakpoint junctions from t(4;14) samples suggested that the majority of these translocations result from illegitimate CSR events. These events were characterized by der(4) breakpoints containing Smu-chromosome 4 junctions and der(14) breakpoints with chromosome 4-downstream switch region junctions. However, not all t(4;14) breakpoints fit this “classical” model, as some derivative chromosomes were observed with hybrid switch regions. Unfortunately, the mechanism that generates these hybrid switch regions has been unclear. In general, only a single derivative was cloned from each patient or cell line. In the one case reported elsewhere in which both derivatives were cloned, the mechanism did not appear to be linked to the CSR process and thus represented a “non-classical” translocation. The poor prognostic impact of t(4;14) myeloma has been well established by several groups, including our own. In an attempt to identify recurrent breakpoint sites and to identify the potential mechanism(s) leading to t(4;14) translocations, we cloned the breakpoint junctions of both derivative chromosomes from 4 cell lines and 5 patients with MB4-2 and MB4-3 breakpoints. Furthermore, we cloned der(4) breakpoints from 4 additional patients, three of which are FGFR3 non-expressers for which we could not detect a der(14) breakpoint using our PCR based strategy. We defined the t(4;14) breakpoint region as encompassing 64.5 kb of chromosome 4, flanked by LETM1 exon 3 and MMSET exon 5, based on combining the previously published breakpoints with our newly cloned and sequenced breakpoints. Current dogma suggests that t(4;14) translocation events are randomly distributed throughout the defined breakpoint region, but this idea is not supported by our sequencing data. We identified two hotspots, which contain breakpoints from 9 of the 27 patients or cell lines with at least one cloned derivative. Interestingly, these regions only represent 1 kb of the entire breakpoint region. Therefore 33% of the cloned breakpoints exist within only 1.5% of the total breakpoint region. Moreover, for the 13 MM samples for which both derivatives are cloned, although 6/13 (46%) fit the classical model of CSR mediated switch translocations, surprisingly, 7/13 (54%) appear to be non-classical translocations. The non-classical translocations are defined by little to no loss of sequence from the involved switch region and the presence of a hybrid switch region on one of the two derivative chromosomes. Importantly, the non-classical translocations may not involve B-cell specific mechanisms and could potentially occur before or after a successful CSR event. Therefore, the classical illegitimate CSR event model can explain only half of the t (4; 14) breakpoints cloned to date.