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Lysosomal inhibition elicited by palmitoyl-protein thioesterase 1 (PPT1) inhibitors such as DC661 can produce cell death, but the mechanism for this is not completely understood. Programmed cell death pathways (autophagy, apoptosis, necroptosis, ferroptosis, and pyroptosis) were not required to achieve the cytotoxic effect of DC661. Inhibition of cathepsins, or iron or calcium chelation, did not rescue DC661-induced cytotoxicity. PPT1 inhibition induced lysosomal lipid peroxidation (LLP), which led to lysosomal membrane permeabilization and cell death that could be reversed by the antioxidant N-acetylcysteine (NAC) but not by other lipid peroxidation antioxidants. The lysosomal cysteine transporter MFSD12 was required for intralysosomal transport of NAC and rescue of LLP. PPT1 inhibition produced cell-intrinsic immunogenicity with surface expression of calreticulin that could only be reversed with NAC. DC661-treated cells primed naive T cells and enhanced T cell-mediated toxicity. Mice vaccinated with DC661-treated cells engendered adaptive immunity and tumor rejection in 'immune hot' tumors but not in 'immune cold' tumors. These findings demonstrate that LLP drives lysosomal cell death, a unique immunogenic form of cell death, pointing the way to rational combinations of immunotherapy and lysosomal inhibition that can be tested in clinical trials., Introduction With encouraging activity in clinical trials that involve the lysosomal inhibitor hydroxychloroquine (HCQ) (1), the identification of palmitoyl-protein thioesterase 1 (PPT1) as the molecular target of chloroquine derivatives (2), [...]

Targeting multiple components of the MAPK pathway can prolong the survival of patients with BRAFV600E melanoma. This approach is not curative, as some BRAF-mutated melanoma cells are intrinsically resistant to MAPK inhibitors (MAPKi). At the systemic level, our knowledge of how signaling pathways underlie drug resistance needs to be further expanded. Here, we have shown that intrinsically resistant BRAF-mutated melanoma cells with a low basal level of mitochondrial biogenesis depend on this process to survive MAPKi. Intrinsically resistant cells exploited an integrated stress response, exhibited an increase in mitochondrial DNA content, and required oxidative phosphorylation to meet their bioenergetic needs. We determined that intrinsically resistant cells rely on the genes encoding TFAM, which controls mitochondrial genome replication and transcription, and TRAP1, which regulates mitochondrial protein folding. Therefore, we targeted mitochondrial biogenesis with a mitochondrium-targeted, small-molecule HSP90 inhibitor (Gamitrinib), which eradicated intrinsically resistant cells and augmented the efficacy of MAPKi by inducing mitochondrial dysfunction and inhibiting tumor bioenergetics. A subset of tumor biopsies from patients with disease progression despite MAPKi treatment showed increased mitochondrial biogenesis and tumor bioenergetics. A subset of acquired drug-resistant melanoma cell lines was sensitive to Gamitrinib. Our study establishes mitochondrial biogenesis, coupled with aberrant tumor bioenergetics, as a potential therapy escape mechanism and paves the way for a rationale-based combinatorial strategy to improve the efficacy of MAPKi., Introduction Cutaneous melanoma is a devastating disease, with a 10-year survival rate of less than 10% in patients diagnosed with stage IV melanoma (1). Approximately 50% of melanoma patients' tumors [...]

Melanomas that result from mutations in the gene encoding BRAF often become resistant to BRAF inhibition (BRAFi), with multiple mechanisms contributing to resistance. While therapy-induced autophagy promotes resistance to a number of therapies, especially those that target PI3K/mTOR signaling, its role as an adaptive resistance mechanism to BRAFi is not well characterized. Using tumor biopsies from [BRAF.sup.V600E] melanoma patients treated either with BRAFi or with combined BRAF and MEK inhibition, we found that BRAFi-resistant tumors had increased levels of autophagy compared with baseline. Patients with higher levels of therapy-induced autophagy had drastically lower response rates to BRAFi and a shorter duration of progression-free survival. In [BRAF.sup.V600E] melanoma cell lines, BRAFi or BRAF/MEK inhibition induced cytoprotective autophagy, and autophagy inhibition enhanced BRAFi-induced cell death. Shortly after BRAF inhibitor treatment in melanoma cell lines, mutant BRAF bound the ER stress gatekeeper GRP78, which rapidly expanded the ER. Disassociation of GRP78 from the PKR-like ER-kinase (PERK) promoted a PERK-dependent ER stress response that subsequently activated cytoprotective autophagy. Combined BRAF and autophagy inhibition promoted tumor regression in BRAFi-resistant xenografts. These data identify a molecular pathway for drug resistance connecting BRAFi, the ER stress response, and autophagy and provide a rationale for combination approaches targeting this resistance pathway., Introduction The 50% response rate and 7-month progression-free survival (PFS) observed in clinical trials of BRAF inhibition (BRAFi) using the inhibitors vemurafenib (also known as PLX4032) and dabrafenib represents a [...]

The mechanisms underlying adaptive resistance of melanoma to targeted therapies remain unclear. By combining ChIP sequencing with microarray-based gene profiling, we determined that ERBB3 is upregulated by FOXD3, a transcription [...]

RAS-driven cancers exhibit variable dependency on autophagy for survival; however, it is not fully understood how. In this issue of the JCI, Cheong and colleagues demonstrate that RAS-dependent elevation of casein kinase 1a (CK1α) negatively regulates autophagy at the level of autophagy gene transcription. Moreover, combined inhibition of both CK1α and autophagy reduced proliferation of RAS-driven tumors. The results of this study provide insight into the connection between mutant RAS and autophagy, and suggest targeting CK1α as a potential therapeutic strategy to modulate autophagy in RAS-driven cancers., The challenge of directly targeting RAS for cancer therapy RAS is a canonical oncogenic driver, with RAS-activating mutations identified in 20%-30% of cancers. Constitutive RAS activation turns on many signaling [...]

Autophagy--a process of "self-eating" that involves enzymatic digestion and recycling of cellular constituents in response to stress--contributes to both cancer cell death and survival. In this issue of the JCI, Lu et al. report that controlled induction of tumor suppressor gene aplasia Ras homolog member I (ARHI) results in autophagic cell death of human ovarian cancer cells in vitro (see the related article beginning on page 3917). However, within xenograft tumors in mice, multiple factors within the tumor microenvironment switched ARHI-induced autophagy to a mechanism of tumor cell survival, leading to tumor dormancy. Since ARHI expression is suppressed in the majority of breast and ovarian cancers but is high in premalignant lesions, ARHI-induced autophagy could be manipulated for therapeutic benefit., Cells that encounter a variety of stresses undergo an evolutionarily conserved process of self-digestion termed autophagy. The importance of this intracellular damage response for pathophysiology has been established across multiple [...]

Autophagy is a lysosome-dependent degradative pathway frequently activated in tumor cells treated with chemotherapy or radiation. Whether autophagy observed in treated cancer cells represents a mechanism that allows tumor cells [...]