Covalent Targeting of Glutamate Cysteine Ligase to Inhibit Glutathione Synthesis.

Dysregulated oxidative stress plays a major role in cancer pathogenesis and some types of cancer cells are particularly vulnerable to inhibition of their cellular antioxidant capacity. Glutamate-cysteine ligase (GCL) is the first and rate-limiting step in the synthesis of the major cellular antioxidant glutathione (GSH). Developing a GCL inhibitor may be an attractive therapeutic strategy for certain cancer types that are particularly sensitive to oxidative stress. In this study, we reveal a cysteine-reactive ligand, EN25, that covalently targets an allosteric cysteine C114 on GCLM, the modifier subunit of GCL, and leads to inhibition of GCL activity. This interaction also leads to reduced cellular GSH levels and impaired cell viability in ARID1A-deficient cancer cells, which are particularly vulnerable to glutathione depletion, but not in ARID1A-positive cancer cells. Our studies uncover a novel potential ligandable site within GCLM that can be targeted to inhibit GSH synthesis in vulnerable cancer cell types.

CYP27A1-dependent anti-melanoma activity of limonoid natural products targets mitochondrial metabolism.

Three limonoid natural products with selective anti-proliferative activity against BRAF(V600E) and NRAS(Q61K)-mutation-dependent melanoma cell lines were identified. Differential transcriptome analysis revealed dependency of compound activity on expression of the mitochondrial cytochrome P450 oxidase CYP27A1, a transcriptional target of melanogenesis-associated transcription factor (MITF). We determined that CYP27A1 activity is necessary for the generation of a reactive metabolite that proceeds to inhibit cellular proliferation. A genome-wide small interfering RNA screen in combination with chemical proteomics experiments revealed gene-drug functional epistasis, suggesting that these compounds target mitochondrial biogenesis and inhibit tumor bioenergetics through a covalent mechanism. Our work suggests a strategy for melanoma-specific targeting by exploiting the expression of MITF target gene CYP27A1 and inhibiting mitochondrial oxidative phosphorylation in BRAF mutant melanomas.

Theranostic Targeting of CUB Domain Containing Protein 1 (CDCP1) in Pancreatic Cancer.

PURPOSE: The recent emergence of radioligand therapies for cancer treatment has increased enthusiasm for developing new theranostic strategies coupling both imaging and cytotoxicity in the same entity. In this study, we evaluated whether CUB domain containing protein 1 (CDCP1), a single-pass transmembrane protein highly overexpressed in diverse human cancers, might be a target for cancer theranostics. EXPERIMENTAL DESIGN: The ectodomain of CDCP1 was targeted using radiolabeled forms of 4A06, a potent and specific recombinant human antibody that we developed. Imaging and antitumor assessment studies were performed in animal models of pancreatic cancer, including two patient-derived xenograft models we developed for this study. For antitumor assessment studies, the endpoints were death due to tumor volume >3,000 mm3 or ≥20% loss in body weight. Specific tracer binding or antitumor effects were assessed with an unpaired, two-tailed Student t test and survival advantages were assessed with a log rank (Mantel-Cox) test. Differences at the 95% confidence level were interpreted to be significant. RESULTS: 89Zr-4A06 detected a broad dynamic range of full length or cleaved CDCP1 expression on seven human pancreatic cancer tumors (n = 4/tumor). Treating mice with single or fractionated doses of 177Lu-4A06 significantly reduced pancreatic cancer tumor volume compared with mice receiving vehicle or unlabeled 4A06 (n = 8; P < 0.01). A single dose of 225Ac-4A06 also inhibited tumor growth, although the effect was less profound compared with 177Lu-4A06 (n = 8; P < 0.01). A significant survival advantage was imparted by 225Ac-4A06 (HR = 2.56; P < 0.05). CONCLUSIONS: These data establish that CDCP1 can be exploited for theranostics, a finding with widespread implications given its breadth of overexpression in cancer.