RUNX1 isoform disequilibrium promotes the development of trisomy 21-associated myeloid leukemia.

Gain of chromosome 21 (Hsa21) is among the most frequent aneuploidies in leukemia. However, it remains unclear how partial or complete amplifications of Hsa21 promote leukemogenesis and why children with Down syndrome (DS) (ie, trisomy 21) are particularly at risk of leukemia development. Here, we propose that RUNX1 isoform disequilibrium with RUNX1A bias is key to DS-associated myeloid leukemia (ML-DS). Starting with Hsa21-focused CRISPR-CRISPR-associated protein 9 screens, we uncovered a strong and specific RUNX1 dependency in ML-DS cells. Expression of the RUNX1A isoform is elevated in patients with ML-DS, and mechanistic studies using murine ML-DS models and patient-derived xenografts revealed that excess RUNX1A synergizes with the pathognomonic Gata1s mutation during leukemogenesis by displacing RUNX1C from its endogenous binding sites and inducing oncogenic programs in complex with the MYC cofactor MAX. These effects were reversed by restoring the RUNX1A:RUNX1C equilibrium in patient-derived xenografts in vitro and in vivo. Moreover, pharmacological interference with MYC:MAX dimerization using MYCi361 exerted strong antileukemic effects. Thus, our study highlights the importance of alternative splicing in leukemogenesis, even on a background of aneuploidy, and paves the way for the development of specific and targeted therapies for ML-DS, as well as for other leukemias with Hsa21 aneuploidy or RUNX1 isoform disequilibrium.

Selective BH3 mimetics synergize with BET inhibition to induce mitochondrial apoptosis in rhabdomyosarcoma cells.

BH3 mimetics are promising novel anticancer therapeutics. By selectively inhibiting BCL-2, BCL-xL, or MCL-1 (i.e. ABT-199, A-1331852, S63845) they shift the balance of pro- and anti-apoptotic proteins in favor of apoptosis. As Bromodomain and Extra Terminal (BET) protein inhibitors promote pro-apoptotic rebalancing, we evaluated the potential of the BET inhibitor JQ1 in combination with ABT-199, A-1331852 or S63845 in rhabdomyosarcoma (RMS) cells. The strongest synergistic interaction was identified for JQ1/A-1331852 and JQ1/S63845 co-treatment, which reduced cell viability and long-term clonogenic survival. Mechanistic studies revealed that JQ1 upregulated BIM and NOXA accompanied by downregulation of BCL-xL, promoting pro-apoptotic rebalancing of BCL-2 proteins. JQ1/A-1331852 and JQ1/S63845 co-treatment enhanced this pro-apoptotic rebalancing and triggered BAK- and BAX-dependent apoptosis since a) genetic silencing of BIM, BAK or BAX, b) inhibition of caspase activity with zVAD.fmk and c) overexpression of BCL-2 all rescued JQ1/A-1331852- and JQ1/S63845-induced cell death. Interestingly, NOXA played a different role in both treatments, as genetic silencing of NOXA significantly rescued from JQ1/A-1331852-mediated apoptosis but not from JQ1/S63845-mediated apoptosis. In summary, JQ1/A-1331852 and JQ1/S63845 co-treatment represent new promising therapeutic strategies to synergistically trigger mitochondrial apoptosis in RMS.

The novel dual BET/HDAC inhibitor TW09 mediates cell death by mitochondrial apoptosis in rhabdomyosarcoma cells.

Targeting the epigenome of cancer cells with the combination of Bromodomain and Extra Terminal (BET) protein inhibitors and histone deacetylase (HDAC) inhibitors has shown synergistic antitumor effects in several cancer types. In this study, we investigate the antitumor potential of the novel dual BET/HDAC inhibitor TW09 in rhabdomyosarcoma (RMS) cells. TW09 reduces cell viability, suppresses long-term clonogenic survival and induces cell death in RMS cells in a dose-dependent manner. Compared to BET/HDAC co-inhibition using JQ1 and MS-275, TW09 induces similar cell death at equimolar concentrations and regulates BET and HDAC target proteins (e.g. c-MYC, H3 acetylation). Mechanistic studies revealed that TW09 upregulates BIM, NOXA, PUMA and BMF, while downregulating BCL-XL, leading to proapoptotic rebalancing of BCL-2 proteins. This results in BAK and BAX activation and caspase-dependent apoptosis, since individual genetic silencing of BIM, NOXA, PUMA, BMF, BAK or BAX, overexpression of BCL-2 or the caspase inhibition with zVAD.fmk all rescue JQ1/BYL719-induced cell death. In conclusion, TW09 shows potent antitumor activity in RMS cells in vitro by inducing mitochondrial apoptosis and may represent a promising new therapeutic option for the treatment of RMS.