Study on the Role of Platelet-Activating Factor in Promoting Relapse of Multiple Myeloma after CAR-T Therapy

Aims: Multiple myeloma (MM) is a malignant plasma cell disease. While CAR-T therapy is highly effective in the early stages of MM treatment, most patients still face relapse. Platelet-activating factor (PAF) has been implicated in tumor development, but its role in MM recurrence after CAR-T treatment remains unexplored. This study aims to investigate the potential involvement of PAF in MM relapse following CAR-T therapy.

Methods: We analyzed data from 33 MM patients who received CAR-T cell therapy at our department. Plasma metabolomics characteristics were analyzed using high-performance liquid chromatography. PAF levels in patient plasma were measured by enzyme-linked immunosorbent assay (ELISA), and PAF receptor (PAFR) protein expression in bone marrow tissue was assessed by immunohistochemistry. QPCR was used to compare PAFR mRNA expression in normal peripheral blood mononuclear cells (PBMCs) and MM cell lines. The high PAFR-expressing OPM2 cell line was selected to construct a luciferase-overexpressing stable strain (OPM2-luc) for proliferation assays with varying PAF concentrations. After co-culturing PAF and BCMA CAR-T cells, the effects of PAF on CAR-T cell proliferation and killing function were evaluated using trypan blue cell counting and a luciferase-based killing assay, respectively.

Results: The overall remission rate for CAR-T cell therapy was 81.8%, with 9 patients achieving complete remission, 11 very good partial remission, and 7 partial remission. However, 17 patients (63%) relapsed. Plasma metabolomics analysis revealed significantly lower levels of phospholipid molecules, including palm carnitine ester and lysophosphatidylcholine (LPC), in the relapse group compared to the remission group (P<0.05). Pathway analysis enriched these differentially metabolized molecules in LPC and PAF metabolic pathways. ELISA results showed significantly higher PAF levels in relapsed patients' plasma, with increased PAFR protein expression in their bone marrow tissue. QPCR results showed that compared with PBMC, the expression levels of PAFR mRNA in 8226, H929, U266, and OPM2 cell lines were significantly increased, among which the expression level in OPM2 cells was the highest (P < 0.05). Various concentrations of PAF were introduced to OPM2-luc cells. After a 96-hour incubation period, significant differences in proliferation effects were observed across the concentration range, with the lowest concentration being 0.1 nmol. Flow cytometry confirmed 92.16% BCMA target antigen expression in OPM2-luc cells. Co-culture experiments showed that PAF had no significant impact on CAR-T cell proliferation or killing function.

Conclusions: Our findings suggest that the PAF remodeling pathway is enhanced following CAR-T treatment of MM, with increased PAF synthesis promoting MM cell proliferation, potentially through the PAF/PAFR pathway. Targeting the PAF pathway may represent an effective strategy to enhance MM CAR-T therapy.