Structure-based inhibitor development targeting RNA epitranscriptomic modifications

Epitranscriptomic modifications are chemical modifications on RNA that affect gene regulation in the cell. RNA modifications have recently been discovered to promote cancer growth. For example, the RNA modification N6 -methyladenine (m 6 A) is associated with proliferation of glioblastoma, an aggressive form of brain cancer, and acute myeloid leukemia (AML), a type of blood cancer that progresses rapidly.

Tumor progression was shown to be suppressed when the levels of m6 A erasers FTO and ALKBH5 were decreased. These findings opened up novel opportunities for anti-cancer treatment by targeting ALKBH5 and FTO using chemical compounds. However, we currently do not have inhibitors that are potent and selective to be used in clinical studies.

We hypothesize that inhibiting m 6 A erasers with small molecule inhibitors will achieve anti-cancer effects. To test our hypothesis, we propose an interdisciplinary approach combining chemical and structural biology to develop FTO and ALKBH5 inhibitors. In our approach, we use macromolecular X-ray crystallography to elucidate the binding modes of inhibitors at the atomic level. We then use the structural information to guide the design of next generation inhibitors, which will then be further investigated in vitro and in vivo.

We previously reported the first structure-based inhibition studies on FTO, and a novel structure of human ALKBH5. Based on the reported findings, we designed second generation FTO inhibitors with the guidance of crystal structures. Our preliminary data showed that we successfully (a) designed a second generation FTO inhibitor with improved potency, (b) confirmed its binding mode by X-ray crystallography, and (c) showed low toxicity in a non- target cell line. In this proposed study, we will further optimize these second-generation inhibitors using the tested structure-based approach to develop clinically relevant FTO and ALKBH5 inhibitors. The third-generation inhibitors will be validated in vitro and in vivo.

The proposed research paves way for a novel approach in cancer therapy by targeting RNA epitranscriptomics. This research will contribute to basic health research and drug discovery in various human diseases as RNA epitranscriptomic modifications have wide- ranging physiological effects, including in development and viral replication.