OA-25: MM B cells and pre-plasma cell progenitors are the cellular origin of MM disease relapse following effective therapy targeting plasma cells

Introduction: Multiple Myeloma (MM) is a mature B cell neoplasm distinguished by the accumulation of plasma cells (PCs) within the bone marrow (BM). Despite treatment advances, MM remains incurable in the majority of patients. The failure to achieve pervasive cure in MM despite the attainment of deep PC responses with PIs, IMiDs, anti-CD38 mAb, melphalan and SCT, bispecific antibodies and CAR-T, suggests the existence of intra-tumoral heterogeneity and rare drug-resistant MM cells with full tumorigenic capacity.

Methods: We examined MM patient BM samples using a combination of FACS, single cell protein immunoflourescence-FISH (IF-FISH), whole exome sequencing (WES), custom-capture targeted deep sequencing (CC-Seq) and single cell RNA sequencing (scRNAseq) to characterize the genomic landscape of intraclonal MM cell subpopulations and to track these subpopulations over time in patients.

Results: FACS-IF-FISH studies of MM patient BM samples (n=140) identified MM progenitor subpopulations that resemble the maturation stages of post germinal centre B cells through to PCs. These clonal subpopulations include rare MM cells that resemble CD20+CD38-CD138-Irf4- Xbp1s- B cells, CD20-CD38-CD138-Irf4-Xbp1s- pre-plasmablasts and CD38+CD138-Irf4+XBP1s+ pre-PCs, as well as tumor-bulk CD38+CD138+IRF4+Xbp1s+ PCs. MM progenitors possess all of the chromosomal translocations and copy number variations (CNV) present in MM PCs including ploidy changes, translocations such as t(4;14), t(14;16) and t(11;14), and secondary aberrations such as gain(1q), del(1p) and del(17p). To examine if MM progenitor cells also possess the single nucleotide variations (SNV) present within MM PCs, and are thus fully malignant, and to track MM progenitor subpopulations within patients over time, we performed WES (n=60) plus CC-Seq (n=210) on purified cell subpopulations from 17 patients, including 5 whom we sampled serially over >3 years. Five cellular subpopulations were isolated from each BM, and each was sequenced to a depth of 4–20,000x. From these data, SNVs in tumor-bulk MM PCs were regularly detected in MM progenitors, although the low frequency of progenitors often prevented capture of complete SNV profiles. Notably, analyses of serial BM samples taken pre and post treatment suggests that relapsing MM PCs typically do not derive from pre-treatment PCs, and often lack multiple SNVs present in pre-treatment PCs. Instead, crucially, relapsing MM PCs commonly derive from MM progenitor cells, as relapse-specific SNVs present in relapsing PCs were detected within MM B cells and progenitor cells isolated from BMs collected up to 3 years prior to relapse. Moreover MM progenitors were also detected as the sole MRD in PC-negative patients.

Conclusions: MM B cells and progenitors possess all of the genomic aberrations required for full malignant potential. Tracking of these cells in vivo in patients by their SNV profile implicates them as the cellular origin of disease relapse. Cure of MM requires eradication of MM progenitor cells.