P846: GENERATION OF A FIRST-IN-CLASS INHIBITOR FOR THE MASTER ONCOREGULATOR HNRNP K IN HAEMATOLOGICAL MALIGNANCIES

Topic: 13. Myeloma and other monoclonal gammopathies - Biology & Translational Research Background: Haematological malignancies comprise a heterogeneous group of cancers that include diverse and biologically different subgroups of neoplasms. Although there has been an improvement in the development of new therapies and medical care of patients, the heterogeneity of haematological malignancies leads to therapy resistance, relapse, and failure of treatments, constituting a therapeutic challenge. Therefore, there is an urgent unmet clinical need to find new biomarkers and potential therapeutic targets that would contribute to overcome the current therapy resistance and dismal outcomes of these neoplasms. Heterogeneous nuclear ribonucleoprotein K (hnRNP K) is a multifunctional RBP implicated in a myriad of biological processes, such as transcription, translation, splicing, RNA stability, and chromatin remodelling. It contains three K-homology domains (KH1-KH3) responsible for nucleic acid binding, one K protein interactive domain (KI) responsible for protein binding, and a nuclear-cytoplasmic shuttling domain (KNS) that confers the ability to translocate bidirectionally through the nuclear pore complex. Interestingly, hnRNP K regulates c-Myc, c-Src, eIF4E, and p53/p21 pathways, and we have also observed that hnRNP K deregulation contributes to haematological malignancies pathogenesis and poor outcomes. Aims: Our goal is to develop a first-in-class hnRNP K inhibitor and evaluate its efficacy to overcome resistance of haematological malignancies. Methods: We carried out a high-throughput screen of over 8000 small molecules (both FDA and non-FDA approved) using AlphaLISA technology and validated both chemically and biologically the inhibition of hnRNP K with those identified compounds. Lastly, we have performed phenotypical and molecular analysis of the effects of the hnRNP K hit inhibitors in several haematological neoplasms cell lines overexpressing hnRNP K through CRISPR synergistic activation mediator (SAM). Results: Around 8000 small compounds were screened using the AlphaLISA technology, and we found 8 interesting hits. We validated three of these hits based on their specificity for hnRNP K and their ability to inhibit this target. Using L363 cells (Multiple Myeloma) and HL60 cells (Acute Myeloid Leukaemia) genetically engineered with CRISPR/SAM technology to overexpress hnRNP K, we demonstrated that these compounds had a biological effect producing changes in viability and reducing levels of hnRNP K downstream molecules. In addition, we contrasted this with the effect in both non-tumour cells (Hs1Int, NL20, and CCD-18Co) and hnRNP K-Knock out L363 and HL60 cells, obtained by CRISPR-Cas9 system. Finally, we explored the possibility of overcoming drug resistance in both Multiple Myeloma and Acute Myeloid Leukaemia cells by co-treatment assays with current therapies. Summary/Conclusion: We have identified three potential first-in-class hnRNP K inhibitors that specifically bind and inhibit hnRNP K in modified Multiple Myeloma and Acute Myeloid Leukaemia cell lines, being one of these validated hits an FDA approved compound. This could be a novel and a promising therapeutic strategy that could help overcome drug resistance in haematological neoplasms currently occurring in the clinic. This work was financially supported by CRIS contra el Cancer Association (NGO), AES ISCIII (PI18/00295 and PI21/00191), ISCIII Miguel Servet (CP19/00140), ISCIII PFIS (FI22/00234) and IF-Marie Sklodowska Curie Actions grant (MAtChinG – 101027864).Keywords: Inhibitor, Screening, Drug resistance