Understanding the Role of KAT1 and KAT2B in Melanoma Resistance To Targeted MAPKi-Therapy

• Applicant: Dr. Sina Jostes
• Subject Area: General Genetics and Functional Genome Biology
• Term: from 2019 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 429315233

The incidence of cutaneous malignant melanoma is rising and its therapeutic management is one of the most challenging for oncologists. Half of cutaneous melanoma patients present with BRAFV600E/K mutations, which result in constitutive activation of the mitogen-activated protein kinase (MAPK) pathway. BRAF-mutated melanoma has a more aggressive phenotype compared to BRAF wildtype melanoma, exhibiting increased tumour cell invasion and metastasis. The development of MAPK inhibitors (MAPKi) has dramatically improved the survival of BRAFV600E/K advanced melanoma patients. However, resistance to targeted MAPK inhibitors remains a major clinical challenge. Using CRISPR-Cas9-mediated gene editing we can now perform large-scale loss-of-function screens, in order to identify genes that upon loss provide resistance to standard therapy approaches (here: MAPKi). Since previous reports suggested that epigenetic modifications and reorganization of the chromatin template promote melanoma drug resistance, the Bernstein lab performed a CRISPR-Cas9 screen targeting ~140 chromatin factors in BRAFV600E human melanoma cells (Strub et al. 2018). Through integrated transcriptomic, epigenomic and proteomic analyses, they identified SIRT6 as a regulator of resistance to MAPKi therapy and determined IGFBP2, an insulin growth factor binding protein, as a direct target gene of SIRT6. Increased IGFBP2 levels promoted melanoma cell survival in the presence of MAPKi. Interestingly, also depletion of the histone acetyltransferases KAT1 and KAT2B conferred resistance to MAPKi treatment in melanoma cells, however so far the molecular mechanisms remain unknown. The following study will provide critical knowledge on the mechanistic role of KAT1 and KAT2B and its downstream effectors in the process of MAPKi-induced melanoma drug resistance, ultimately aiming for the identification of novel therapeutic targets for BRAFV600E/K melanoma and other MAPK-driven cancers.Aim 1: I plan to perform RNA-sequencing upon loss of KAT1 or KAT2B (+/- MAPKi) and compare it to parental KAT1 and KAT2B wildtype cells. Thereby, I will identify candidate genes (such as IGFBP2) that are altered upon loss of KAT1 or KAT2B and may promote resistance to MAPKi treatment.Aim 2: Next, I will perform ChIP-sequencing analysis for KAT1 and KAT2B, as well as for KAT1- and KAT2B- related histone modifications and non-related modifications (e.g. the active chromatin mark H3K27ac) in KAT1 or KAT2B-depleted cells (+/- MAPKi). The ChIP-seq analysis will be compared to the generated RNA-seq data to unravel the epigenetic landscape of melanoma drug resistance.Aim 3: In order to confirm the generated findings, KAT1 or KAT2B knockout cell lines will be injected into the flank of immunodeficient mice and tumour growth will be analysed (+/- MAPKi). After histopathological analyses of tumour samples and comparisons between the different treatments, the impact of KAT1 or KAT2B knockout versus control will be evaluated.

• DFG Programme: Research Fellowships
• International Connection: USA
• Host: Professorin Dr. Emily Bernstein