Abstract B010: Systematical evaluation on alternative splicing in immune-related therapy targets reveals innate and acquired therapy resistance mechanisms in multiple myeloma
Authors: Liu Enze , Johnson Travis S , Sudha Parvathi , Hamidi Habib , Zafar Faiza , Chopra Vivek S , Abonour Rafat , Walker Brian A 
Published: November 2024

Introduction: Although multiple myeloma (MM) is considered incurable, immune-related therapies are emerging as promising approaches to prolong patient survival. Alternative splicing (AS) has been identified as a mechanism causing therapy resistance to CD19 CAR-T in B-cell malignancies, but no systematic evaluation has been made on the impact towards MM immunotherapy targets. Methods: Utilizing two datasets (MMRF N=852 and IU N=122), we selected high-confidence targets based on high RNA expression (Log2(TPM+1)&gt;3), low toxicity (nTPM&gt;50 in &lt;2 normal tissues and in &lt;2 normal blood cells) and high target potential (&gt;3 surfaceome databases or established immune-therapy targets). Known resistance mechanisms of other therapies, such as immunomodulatory drugs (IMiDs) were also included. AS analysis was conducted for all identified targets. Expression and AS events between patient samples and 19 MM cell lines from DepMap database were also measured. Results: 102 high-confidence targets were identified in MM samples including six existing immunotherapy targets (BCMA, SLAMF7, GPRC5D, CD79A/B and CD38). From these 102 genes, 361 significant AS events were subsequently identified (|dPSI|&gt;0.2, FDR&lt;0.05, &gt;10 supporting reads in &gt;25% samples). One event of interest included an aberrant exon inclusion event in the existing target FCRL5 in ∼10% of newly diagnosed and relapse patient samples, despite being treatment naïve. A cryptic exon between exon 3 and 4 of the canonical transcript was found with higher usage in MM (PSI&gt;0.48) compared to 8 normal bone marrow plasma cell samples (=0.13). Inclusion of this exon led to the switch from three protein-coding transcript variants (FCRL5-201, -202, -203) to a transcript encoding a truncated protein after the first Ig domain (-206). The AS event in treatment-naïve patients may lead to the selection of this variant in treated patients as a mechanism of therapy resistance. Another key event was detected in CRBN, which encodes the target of IMiDs including lenalidomide. Serial samples in one patient treated with lenalidomide identified a novel splice junction in CRBN in the later sample (dPSI=0.2). The novel splice junction led to the skipping of exon 8 encoding the LON domain, which is essential for maintaining the functional conformation of the IMiD-binding site. Lack of exon 8 will disrupt LON domain integrity, indicating a potential AS-induced resistance mechanism. Interestingly, the novel splice junction was found in 70% relapsed samples (&gt;10 supporting reads), 60% of which were with high exon skipping level (PSI&lt;0.5). Conclusion: We have identified potential innate and acquired AS mechanisms that affect patient response towards immune-related therapies, highlighting the importance of conducting RNA-based diagnostics for MM patients.

Citation Format: Enze Liu, Travis S Johnson, Parvathi Sudha, Habib Hamidi, Faiza Zafar, Vivek S Chopra, Rafat Abonour, Brian A Walker. Systematical evaluation on alternative splicing in immune-related therapy targets reveals innate and acquired therapy resistance mechanisms in multiple myeloma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: RNAs as Drivers, Targets, and Therapeutics in Cancer; 2024 Nov 14-17; Bellevue, Washington. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(11_Suppl):Abstract nr B010.