Long-term Analysis Of Multiple Sequential Samples Reveals Patterns Of Progression In Smoldering Myeloma

Smoldering myeloma (SMM) is an asymptomatic plasma cell disorder, distinguished from monoclonal gammopathy of undetermined significance (MGUS) by a much higher risk of progression to symptomatic multiple myeloma (MM). We postulated that a better understanding of the molecular makeup of SMM may help identify new risk factors for progression that may be used to define new inception strategies. Sequential samples from 9 SMM patients (9 controls and 54 tumors) with a median follow-up of 7 years (range: 3.5 to 12.8 years) were analysed. DNA was obtained from either CD138+ cells from the bone marrow of SMM patients through time (tumor) or from non-tumor cells from the same patient (control). 100 ng of DNA was fragmented, end-repaired, and adapters ligated, before hybridization using NimbleGen's MedExome with an additional capture for the IGH, IGK, IGL, and MYC loci. After PCR amplification hybridized libraries were sequenced on a NextSeq500 (Illumina) using 75 bp paired end reads. The median coverage was 93x (IQR 86-105) and 100x (IQR 95-103) for tumors and controls, respectively. The number of mutations per sample increased with time from diagnosis. There was a trend suggesting the mutation rate of progressors (n=6) was higher than the one of non-progressors (n=3) (F=3.9, p=0.052). Samples with hyperdiploidy had a higher rate of mutations over time than the others (F=9, p=0.009). Bi-allelic events were a common finding immediately before progression. We detected translocations in 7/9 patients. In most cases, they were seen at every time point. In one case, a t(8;14) was detected during follow up, 5.9 years from diagnosis. Using ddPCR, we quantified the rearranged MYC allele and compared the frequency to the IGH rearranged locus. This t(8;14) was not present at diagnosis, appeared in a small fraction (1%) 4.1 years after initial diagnosis and steadily increased over time reaching 37% in the last sample, 8.2 years from the initial diagnosis. Clonality analysis was available for eight patients and identified a median of seven clones per patient, most of them from branching evolution (7/8) but one yielding a linear pattern (1/8). Dramatic changes in clonal structures could be seen in all samples up to one year prior to the biological or clinical progression. Driver events were seen in 7/9 patients. Four patients had more than one driver, which were in different clones in two patients and in the same clone in two patients. In summary, comprehensive analysis of multiple SMM samples over time offers greater insight into the mechanisms of progression of SMM to MM including both novel events (such as MYC translocations and biallelic events) and changes in the clonal architecture. All these changes occurred several months before progression suggesting they could be used as early progression markers.