Resistance to standard of care taxane and androgen deprivation therapy (ADT) causes the vast majority of prostate cancer (PC) deaths worldwide. We have developed RapidCaP, an autochthonous genetically engineered mouse model of PC. It is driven by the loss of PTEN and p53, the most common driver events in PC patients with life-threatening diseases. As in human ADT, surgical castration of RapidCaP animals invariably results in disease relapse and death from the metastatic disease burden. Fatty Acid Binding Proteins (FABPs) are a large family of signaling lipid carriers. They have been suggested as drivers of multiple cancer types. Here we combine analysis of primary cancer cells from RapidCaP (RCaP cells) with large-scale patient datasets to show that among the 10 FABP paralogs, FABP5 is the PC-relevant target. Next, we show that RCaP cells are uniquely insensitive to both ADT and taxane treatment compared to a panel of human PC cell lines. Yet, they share an exquisite sensitivity to the small-molecule FABP5 inhibitor SBFI-103. We show that SBFI-103 is well tolerated and can strongly eliminate RCaP tumor cells in vivo. This provides a pre-clinical platform to fight incurable PC and suggests an important role for FABP5 in PTEN-deficient PC.

Krüppel-like factors (KLFs) belong to the family of transcription factors with three highly conserved zinc finger domains in the C-terminus. They regulate homeostasis, development, and disease progression in many tissues. It has been shown that KLFs play an essential role in the endocrine and exocrine compartments of the pancreas. They are necessary to maintain glucose homeostasis and have been implicated in the development of diabetes. Furthermore, they can be a vital tool in enabling pancreas regeneration and disease modeling. Finally, the KLF family contains proteins that act as tumor suppressors and oncogenes. A subset of members has a biphasic function, being upregulated in the early stages of oncogenesis and stimulating its progression and downregulated in the late stages to allow for tumor dissemination. Here, we describe KLFs’ function in pancreatic physiology and pathophysiology.

Oxidative phosphorylation is an essential metabolic process for cancer proliferation and therapy resistance. The ClpXP complex maintains mitochondrial proteostasis by degrading misfolded proteins. Madera Therapeutics has developed a class of highly potent and selective small-molecule activators (TR compounds) of the ClpXP component caseinolytic peptidase proteolytic subunit (ClpP). This approach to cancer therapy eliminates substrate recognition and activates nonspecific protease function within mitochondria, which has shown encouraging preclinical efficacy in multiple malignancies. The class-leading compound TR-107 has demonstrated significantly improved potency in ClpP affinity and activation and enhanced pharmacokinetic properties over the multitargeting clinical agent ONC201. In this study, we investigate the in vitro efficacy of TR-107 against human colorectal cancer cells. TR-107 inhibited colorectal cancer cell proliferation in a dose- and time-dependent manner and induced cell cycle arrest at low nanomolar concentrations. Mechanistically, TR-107 downregulated the expression of proteins involved in the mitochondrial unfolded protein response and mitochondrial DNA transcription and translation. TR-107 attenuated oxygen consumption rate and glycolytic compensation, confirming inactivation of oxidative phosphorylation and a reduction in total cellular respiration. Multiomics analysis of treated cells indicated a downregulation of respiratory chain complex subunits and an upregulation of mitophagy and ferroptosis pathways. Further evaluation of ferroptosis revealed a depletion of antioxidant and iron toxicity defenses that could potentiate sensitivity to combinatory chemotherapeutics. Together, this study provides evidence and insight into the subcellular mechanisms employed by colorectal cancer cells in response to potent ClpP agonism. Our findings demonstrate a productive approach to disrupting mitochondrial metabolism, supporting the translational potential of TR-107.

Supplementary Table S1. Proteomics data

Supplementary Figure S8. TR-107 reduces total ATP production confirming an inactivation of OXPHOS and suppressed glycolytic compensatory response. Colorectal cancer cells (A) DLD-1, (B) LS1034, (C) HT29, and (D) HCT116 were treated with vehicle control (0.1% DMSO) or indicated concentrations of TR-107 for 12 hours. Quantification of OCR and PER was measured with the Seahorse XF Real-Time ATP Rate Assay. Mitochondrial, glycolytic, and total ATP production rates were calculated using OCR and PER values as described in Materials and Methods. Data are presented as mean ± SD values (N=6). *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

Supplementary Table S3. Transcriptomic and proteomic data analysis.