Andre J. Wijaya

Mutations in the Kirsten rat sarcoma viral oncogene homolog (KRAS) protein are highly prevalent in cancer. However, small-molecule concepts that address oncogenic KRAS alleles remain elusive beyond replacing glycine at position 12 with cysteine (G12C), which is clinically drugged through covalent inhibitors. Guided by biophysical and structural studies of ternary complexes, we designed a heterobifunctional small molecule that potently degrades 13 out of 17 of the most prevalent oncogenic KRAS alleles. Compared with inhibition, KRAS degradation results in more profound and sustained pathway modulation across a broad range of KRAS mutant cell lines, killing cancer cells while sparing models without genetic KRAS aberrations. Pharmacological degradation of oncogenic KRAS was tolerated and led to tumor regression in vivo. Together, these findings unveil a new path toward addressing KRAS-driven cancers with small-molecule degraders.

Propionate is an abundant catabolite in nature and represents a rich potential source of carbon for the organisms that can utilize it. However, propionate and propionate-derived catabolites are also toxic to cells, so propionate catabolism can alternatively be viewed as a detoxification mechanism. In this review, we summarize recent progress made in understanding how prokaryotes catabolize propionic acid, how these pathways are regulated and how they might be exploited to develop novel antibacterial interventions.

Abstract The ubiquitin E3 ligase cereblon (CRBN) is the target of therapeutic drugs thalidomide and lenalidomide and is recruited by most targeted protein degraders (PROTACs and molecular glues) in clinical development. Biophysical and structural investigation of CRBN has been limited by current constructs that either require co-expression with the adaptor DDB1 or inadequately represent full-length protein, with high-resolution structures of degrader ternary complexes remaining rare. We present the design of CRBN midi , a construct that readily expresses from E. coli with high yields as soluble, stable protein without DDB1. We benchmark CRBN midi for wild-type functionality through a suite of biophysical techniques and solve high-resolution co-crystal structures of its binary and ternary complexes with degraders. We qualify CRBN midi as an enabling tool to accelerate structure-based discovery of the next generation of CRBN based therapeutics.

Pseudomonas aeruginosa is an opportunistic human pathogen that, due to its unrivalled resistance to antibiotics, ubiquity in the built environment, and aggressiveness in infection scenarios, has acquired the somewhat dubious accolade of being designated a "critical priority pathogen" by the WHO. In this work, we uncover the pathways and mechanisms used by P. aeruginosa to grow on a substrate that is abundant at many infection sites: propionate. We found that if the organism is prevented from metabolizing propionate, the substrate turns from being a convenient nutrient source into a potent poison, preventing bacterial growth. We further show that one of the enzymes involved in these reactions, 2-methylcitrate synthase (PrpC), is promiscuous and can moonlight for another essential enzyme in the cell (citrate synthase). Indeed, mutations that abolish citrate synthase activity (which would normally prevent the cell from growing) can be readily overcome if the cell acquires additional mutations that increase the expression of PrpC. This is a nice example of the evolutionary utility of so-called "underground metabolism."

Targeting host proteins that are crucial for viral replication offers a promising antiviral strategy. We have designed and characterised antiviral PROteolysis TArgeting Chimeras (PROTACs) targeting the human protein cyclophilin A (CypA), a host cofactor for unrelated viruses including human immunodeficiency virus (HIV) and hepatitis C virus (HCV). The PROTAC warheads are based on fully synthetic macrocycles derived from sanglifehrin A, which are structurally different from the classical Cyp inhibitor, cyclosporine A. Our Cyp-PROTACs decrease CypA levels in cell lines and primary human cells and have high specificity for CypA confirmed by proteomics experiments. Critically, CypA degradation facilitates improved antiviral activity against HIV-1 in primary human CD4+ T cells compared to the non-PROTAC parental inhibitor, at limiting inhibitor concentrations. Similarly, we observe antiviral activity against HCV replicon in a hepatoma cell line. We propose that CypA-targeting PROTACs inhibit viral replication potently and anticipate reduced evolution of viral resistance and broad efficacy against unrelated viruses. Furthermore, they provide powerful tools for probing cyclophilin biology.