Single-cell landscape of clonal plasma cells in t(11;14) primary light-chain amyloidosis reveals molecular insights into venetoclax response Free

Venetoclax (Ven), a selective BCL2 inhibitor, has demonstrated promising efficacy in the treatment of primary light-chain amyloidosis (pAL), particularly in patients harboring t(11;14). Despite this, approximately 17–37% of patients fail to achieve a hematologic complete response (CR) or very good partial response (VGPR). Mechanistic insights from multiple myeloma (MM) suggest that higher BCL2/BCL2L1 or BCL2/MCL1 ratios, reflecting a survival dependency biased toward BCL2, and retention of B-cell transcriptional features may predict Ven sensitivity. However, the molecular landscape of t(11;14) pAL and the extent to which Ven-resistant clonal plasma cells (cPCs) share biological features with MM remain largely unexplored.

We conducted single-cell transcriptomic and V(D)J profiling using 10x Genomics 5′ RNA sequencing on CD138⁺ magnetic bead–enriched bone marrow plasma cells from 37 newly diagnosed pAL patients prior to Ven exposure, of whom 21 exhibited t(11;14) ≥10%. Twelve patients subsequently received Ven-based therapy (7 first-line Ven+dexamethasone, 5 second-line Ven+dexamethasone ±daratumumab). Single-cell data were processed via our PRISM (Plasma cell disease Repertoire Identification and Single-cell Mapping) workflow. A total of 151,908 cPCs were identified and pseudobulked for differential analysis using DESeq2, with non-negative matrix factorization (NMF) and gene set enrichment analysis (GSEA) applied to define gene expression programs and functional pathways. Median clinical follow-up was 19 months.

Comparison by t(11;14) status revealed CCND1 overexpression (adjusted p<0.001) as expected. Both BCL2 and MCL1 trended higher in t(11;14) group (adjusted p=0.011 and 0.053, respectively), despite marked intra-group heterogeneity with extreme outliers, while BCL2L1 levels were comparable. Notably, BCL2/BCL2L1 ratio was significantly elevated in t(11;14) cPCs (p<0.001), whereas the BCL2/MCL1 ratio showed no significant difference. Multiple B-cell–associated genes (MS4A1, PAX5, PIK3AP1, BTK) were enriched, and NMF identified two gene expression programs upregulated in t(11;14) cPCs: one enriched in cellular response to tumor necrosis factor (GO:0034612) and the other in response to endoplasmic reticulum stress (GO:0034976), implying potential susceptibility to both extrinsic and intrinsic apoptotic pathways.

Among 12 Ven-treated patients, 8 achieved CR or VGPR, while 4 experienced PR or SD, including 2 with t(11;14) <10% and low CCND1 levels. Analysis of 43,795 cPCs from these patients showed no single BCL2-family gene that consistently distinguished responders; however, BCL2/BCL2L1 ratio was significantly higher in Ven-sensitive cPCs (p=0.016), whereas BCL2/MCL1 ratio lacked discriminatory power. While individual B-cell marker genes did not achieve significance, GSEA using published B-cell gene sets (GSE12366, GSE13411) indicated enrichment in optimal responders. In contrast, Ven-sensitive gene sets of MM (Gupta et al, Blood 2021) did not show significant differences between response groups. Furthermore, DESeq2 highlighted upregulation of DDIT3, TRIB3, and ERN1—core components of the intrinsic apoptotic signaling pathway in response to endoplasmic reticulum stress (GO:0070059)—in Ven-sensitive cPCs, corroborated by enrichment of “HALLMARK_UNFOLDED_PROTEIN_RESPONSE” (NES=–2.14, FDR<0.01). To the contrary, cPCs of suboptimal responders preferentially expressed gene programs associated with positive regulation of protein transport (GO:0051222) and amyloid precursor protein metabolic processes (GO:0042982), suggesting a transcriptional state associated with protein vesicular trafficking and quality control in the endoplasmic reticulum and Golgi, in line with previous reports (Gort-Freitas et al., Blood 2025).

Our study delineates a distinct molecular signature of t(11;14) pAL, defined by BCL2 dependency and a B-cell–like transcriptional phenotype, consistent with its overall sensitivity to Ven. Conversely, Ven-resistant clones exhibit lower BCL2/BCL2L1 ratios, aligning with relative BCL2 independence. In addition to these pathways, Ven-responsive clones demonstrate a robust unfolded protein response and intrinsic apoptotic priming. This unique apoptotic tendency driven by the stress of amyloid accumulation, which is not typically observed in Ven-sensitive MM cells, may underlie the superior hematologic responses to Ven in pAL compared with MM in clinical settings.