Abstract 3412: Liquid biopsy for t(11;14)/blc2 identification in multiple myeloma

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Abstract Background: Multiple myeloma (MM) is a spatially heterogeneous plasma cell malignancy of the bone marrow (BM). Venetoclax is a potent inhibitor of the anti-apoptotic protein BCL2 with evidence in MM that t(11;14) and high BCL2 expression confers greater sensitivity to venetoclax. BM biopsy is required for fluorescent in-situ hybridisation (FISH) detection of t(11;14) and the evaluation of MM cell BCL2 expression, with success being entirely dependent on the quality of the BM sample procured. We hypothesised that a liquid biopsy strategy to enable interrogation of cell-free circulating tumour (ct)DNA and extra-cellular (ex)RNA would represent a more practical and less invasive alternative strategy to BM biopsy for the detection of t(11;14) and the determination of MM cell BCL2 expression, respectively. Methods: Blood and BM were obtained from 24 FISH characterised MM patients - normal karyotype (n=9), t(11;14) (n=11) and t(4;14) (n=4). Plasma derived ctDNA and exRNA was extracted utilising the QIAamp circulating nucleic acid kit and matched PBMC DNA was obtained for germ line controls. BM CD138+ MM cells for DNA and RNA extraction were selected with Miltenyi Biotec CD138 microbeads. A low density custom panel of unique capture probes (SSEL XT HS2, SureSelect Custom Tier 2, Agilent Technology), spanning approximately 10% of the CCND1 and IGH loci involved in t(11;14), was used for sequencing with approximately 5 million reads per sample with adapter sequences trimmed from the reads and aligned to the hg38 using bwa mem algorithm. Structural variant caller, Lumpy was used for identifying translocations, with low confidence reads excluded with a defined Z-score threshold. Droplet digital PCR (ddPCR) for BCL2 was performed on exRNA. Results: The low-density capture seq identified t(11;14) in 6 of 11 (55%) of the CD138+ BM samples, but not in paired germ-line samples nor in the non-t(11;14) cohort BM samples. Interrogation of ctDNA identified t(11;14) in 1 of 11 samples but also in ctDNA from a patient labelled as being t(4;14) positive raising questions about the validity of the FISH attribution. We are now utilising a modification of the assay using ultra-high density capture probes with coverage approaching 100% of the IGH region and higher ctDNA input to detect translocations involving the IGH region on chromosome 14 with any partner chromosome, including translocations undetectable by FISH. BCL2 exRNA transcripts were detected in all patients with a range of 2-120 copies/ml of plasma with 79% of patients classified as ‘high’ BCL2 expression (>40 copies/ml of plasma) compared to 59% of the matched CD138+ BM samples. Conclusion: We provide proof-of-concept of the potential of ctDNA and exRNA for t(11;14) detection and ddPCR quantitation of BCL2 expression, respectively, providing a rationale for further exploration of ctDNA and exRNA liquid biopsy testing as a strategy for identifying patients with t(11;14) and/or high BCL2. Citation Format: Andrew Spencer, Sridurga Mithraprabhu, Jaynish Shah, Kawa Choi, Ashley George, Carlos Hader, Tiffany Khong. Liquid biopsy for t(11;14)/blc2 identification in multiple myeloma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3412.

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