Introduction Surface proteins in cancer not only reflect tumor–microenvironment interactions but also serve as targets for diagnostic flow cytometry and precision therapies, including monoclonal antibodies and CAR-T cell therapies. A better understanding of plasma cell immunophenotypes could inform flow cytometric prognostic assays and development of targeted therapies.

Methods RNA-seq and Seq-FISH data from CD138-enriched bone marrow plasma cells, along with corresponding clinical annotations, were obtained from the Multiple Myeloma Research Foundation-CoMMpass Study. A curated list of surface genes was assembled by integrating three sources: the Human Protein Atlas (plasma membrane and cell junction annotations), the Gene-Set Enrichment Analysis membrane-associated gene set (Ontology:0016020), and the Cell Surface Protein Atlas (validated high-confidence category). A total of 4,579 genes coding for cell surface proteins were identified. Genes below the 25th variance percentile were removed. PCA (≥90% variance) followed by k-means (k=2, silhouette) and confirmed by hierarchical clustering. A Plasma Cell Maturity (PBM) score was computed using singscore on 84 mature plasma cell–associated genes to compare cluster phenotypes. Pathway analysis was done using GSEA software for Hallmark pathways. Clinical biomarkers and survival kinetics were compared between the clusters.

Results 761 newly diagnosed patients were included. (Age: 27-93 years, median overall survival/OS: 29 months/mo). Clustering revealed two groups which we termed Atypical/Aggressive Cluster 1 (C1, n=105, 13.8%) and Classical Cluster 2 (C2, n=656, 86.2%), based on the patterns and clinical characteristics. Among non-missing cases, high-risk disease per the International Myeloma Society–International Myeloma Working Group (IMS–IMWG) definition was observed in 27.7% of patients in C2 (n = 552) and 24.7% in C1 (n = 90) (p = 0.4).C1 exhibited enrichment for t(14;16)-MAF (OR = 2.6, 95% CI: 1.1–6.1, p = 0.04) and t(11;14)-CCND1 (OR = 1.7, CI: 1.01–2.8, p = 0.04). In contrast, hyperdiploidy was more common in C2 (OR for C1 = 0.6, CI: 0.4–0.9, p = 0.04). Elevated LDH was observed in C1 (OR = 2.6, CI: 1.6–4, p < 0.001). Median serum Beta-2 microglobulin was lower in C1.(p=0.009)

Upregulated genes in C1 in decreasing order of log Fold Change(logFC): AHNAK, MT-ND1, EEF2, EZR, HLA-B, TXNIP, HLA-E, TRIB1, MCL1, RHOB, CD74, CFLAR, HLA-A, HERPUD1, RPSA, ERN1, HSP90AB1, RAPGEF2, ADGRE5, and LAPTM5. Downregulated genes in C1 in descending order of |logFC|: B2M, SDC1 (CD138), HSP90B1, SEL1L, CALR, CANX, PECAM1, SLAMF7, STT3A, TNFRSF17 (BCMA), FCRL5, STT3B, PALM2AKAP2, CD53, PARP1, CCR2, RPN1, CD38, EDNRB, and LMAN2. Upregulated pathways in C1: proliferation (MYC, E2F, PI3K/AKT/mTOR, KRAS, Wnt); evasion of apoptosis (p53, TGF-β, Notch, NF-κB, IL6/JAK/STAT3); Epithelial to Mesenchymal Transition (angiogenesis, apical junction); inflammatory (interferon, TNF-α, complement, IL2/STAT5); metabolic reprogramming (oxidative phosphorylation, lipid/cholesterol); and stress (DNA repair, unfolded protein response). The PBM score for C1 was significantly lower, suggesting a less differentiated phenotype.

C1 patients had inferior survival (n = 93, median OS 43.9 mo, p = 0.04) compared to those in the C2 cluster (n = 559, median OS 102.5 mo). Patients in the C1 + IMS–IMWG high-risk status had the shortest survival (20.7 mo), compared to C1+low (50 mo), C2/high (68.3 mo), and C2+low (not reached). In a multivariable analysis, C1 status (HR = 1.93; 95% CI: 1.1–3.4, p = 0.02) was an independent predictor, along with older age (1.05), UAMS70 (1.02), EMC-92 (1.4). In C1, early (<18 mo) progression occurred in 76.3% (n=71) (p < 0.001). In patients receiving first line of treatment(LOT1), median PFS was 39.7 mo in C1vs. 47.3 mo in C2 (p=0.8). Among patients who received stem cell transplant in LOT1, the median survival for the C2 was 102.5 mo, compared to 44.5 mo for C1 (n = 52) (p = 0.01). Among non-SCT patients on Immunomodulator + Proteasome Inhibitor therapy, C1 (n=28) had shorter survival of 50.0 vs. 71.7 C2 (n=137; p =0.8).

Conclusion Our transcriptomic analysis identified a higher-risk group of patients (C1) exhibiting downregulation of myeloma specific markers (CD38, CD138, BCMA, SLAMF7, FCRL5) and inferior outcomes. It is important to diagnose and find appropriate therapy for these patients as they don't obtain the same benefit from the current standard of therapy as the remaining patients.

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2025
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