Abstract 1379: Discovering the essential functions of MCL-1 in hematopoiesis using gene-swap mice

Pro-survival BCL-2 proteins (e.g., MCL-1, BCL-XL, BCL-2, A1) are over-expressed in human cancers, making them critical targets for anti-cancer therapies 1. While the BCL-2 inhibitor Venetoclax is highly successful in treating certain blood cancers, inhibitors of BCL-XL and MCL-1 cause toxicity to non-malignant tissues, including hematopoietic cells 2. These findings from gene targeted mice showed that different pro-survival BCL-2 proteins are critical for the survival of distinct hematopoietic cell subsets. Notably, MCL-1 is essential for implantation, with Mcl-1-/- embryos dying at E3.5, the survival of hematopoietic stem and progenitor cells as well as many immature and mature lymphoid and myeloid cell populations3. In contrast, the essential roles for BCL-2 and BCL-XL are restricted to the survival of select hematopoietic cell populations. Of note MCL-1 (and to a minor extent BCL-XL) but not BCL-2 have been ascribed some apoptosis unrelated functions in cellular metabolism, such as mitochondrial import of long chain fatty acids. We aim to determine the unique anti-apoptotic and apoptosis unrelated roles of MCL-1 in comparison to the other pro-survival BCL-2 family members in embryogenesis and hematopoiesis.We used CRISPR to produce “gene-swap” mice in which the MCL-1 coding region was replaced with the coding region for BCL-2 (Mcl-1Bcl-2), BCL-XL (Mcl-1Bcl-xL) or A1. Remarkably, homozygous Mcl-1Bcl-xL/Bcl-xL and Mcl-1Bcl-2/Bcl-2 embryos could develop until at least E11.5 on a C57BL/6 background. This demonstrates that only the anti-apoptotic function of MCL-1 - but not its apoptosis-unrelated roles - are essential for early embryogenesis.To examine the impact of replacement of MCL-1 by BCL-XL on hematopoiesis, lethally irradiated mice were reconstituted with Mcl-1Bcl-xL/Bcl-xL hematopoietic stem progenitor cells from E12.5 embryos. Interestingly, the expression of BCL-XL in place of MCL-1 in the hematopoietic system allowed the generation of all blood cell types with a marked increase in platelets and B lymphocytes. Notably, adult heterozygous Mcl-1Mcl-1/Bcl-2 mice showed even more substantial accumulations of B cells and antibody secreting plasma cells in their spleen and bone marrow, and with progressing age they develop SLE-like autoimmune disease similar to what is seen in vav-bcl-2 transgenic mice that overexpress BCL-2 4. These findings show that distinct pro-survival BCL-2 proteins with their differences in protein stability and interactions with the pro-apoptotic members of the BCL-2 family are best suited to safeguard normal hematopoietic cell survival, with too much pro-survival activity causing disease. References1.Adams, J. M. & Cory, S. Cell Death Differ 25, 27-36 (2018).2.Kelly, G. L. & Strasser, A. Annu Rev Cancer Biol 4, 299-313 (2020).3.Opferman, J. T. et al. Nature 426, 671-676 (2003).4.Ogilvy, S. et al. Proceedings of the National Academy of Sciences 96, 14943-14948 (1999). Citation Format: Annli Tee, Andrew J. Kueh, Leonie Gibson, Shezlie Malelang, Marco J. Herold, Gemma Kelly, Andreas Strasser, Kerstin Brinkmann. Discovering the essential functions of MCL-1 in hematopoiesis using gene-swap mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1379.