SIGNIFICANCE The identification of PLK1 as a potent synthetic lethal target for FGFR-targeted therapy provides an innovative rationale for the treatment of lung and other FGFR1-amplified cancers.PI3Kα inhibitors have shown clinical activity in PIK3CA-mutated estrogen receptor-positive (ER+) patients with breast cancer. Using whole genome CRISPR/Cas9 sgRNA knockout screens, we identified and validated several negative regulators of mTORC1 whose loss confers resistance to PI3Kα inhibition. Among the top candidates were TSC1, TSC2, TBC1D7, AKT1S1, STK11, MARK2, PDE7A, DEPDC5, NPRL2, NPRL3, C12orf66, SZT2, and ITFG2. Loss of these genes invariably results in sustained mTOR signaling under pharmacologic inhibition of the PI3K-AKT pathway. Moreover, resistance could be prevented or overcome by mTOR inhibition, confirming the causative role of sustained mTOR activity in limiting the sensitivity to PI3Kα inhibition. Cumulatively, genomic alterations affecting these genes are identified in about 15% of PIK3CA-mutated breast tumors and appear to be mutually exclusive. This study improves our understanding of the role of mTOR signaling restoration in leading to resistance to PI3Kα inhibition and proposes therapeutic strategies to prevent or revert this resistance. SIGNIFICANCE These findings show that genetic lesions of multiple negative regulators of mTORC1 could limit the efficacy of PI3Kα inhibitors in breast cancer, which may guide patient selection strategies for future clinical trials.In nutrient-poor conditions, autophagy buffers metabolic stress and counteracts the effects of chemotherapy and radiation on cancer cells, which depend on autophagy for survival. However, clinical trials targeting autophagy have failed to produce successful anticancer treatments using currently available inhibitors. Recent studies have shown that PIKfyve kinase inhibitors disrupt lysosome function in autophagy and can selectively kill certain cancer cells. Analysis of biochemical changes caused by PIKfyve inhibition revealed that resistant cells contain significantly higher levels of cellular p38MAPK protein and phosphorylation. Expression of the lysosomal protein, lysosomal-associated membrane protein 2, carrying phosphomimetic mutations of the p38MAPK phosphorylation sites prevented all effects caused by PIKfyve inhibition-induced lysosome dysfunction. selleckchem Thus, the activation of p38MAPK in response to PIKfyve inhibition revealed a novel compensatory role in maintaining lysosome function in autophagy. The functional cooperation between the cellular PIKfyve and p38MAPK pathways in regulating lysosome homeostasis was especially important in cancer cells. Combined inhibition of PIKfyve and p38MAPK activities synergistically blocked autophagy-mediated protein degradation, prevented cathepsin maturation, and markedly reduced the viability of multiple cancer cell types without affecting the viability of normal cells. Furthermore, combined PIKfyve and p38MAPK inhibitors synergistically reduced tumor growth in mice bearing xenografts of human colorectal adenocarcinoma, suggesting a novel way to target cancer cells by prolonged inhibition of autophagy using lower drug concentrations. SIGNIFICANCE This study demonstrates that PIKfyve and p38MAPK cooperate to regulate lysosome homeostasis and their combined inhibition synergistically blocks autophagy to reduce cancer cell viability in vitro and in vivo.Deferoxamine (DFO) represents a widely used iron chelator for the treatment of iron overload. Here we describe the use of mitochondrially targeted deferoxamine (mitoDFO) as a novel approach to preferentially target cancer cells. The agent showed marked cytostatic, cytotoxic, and migrastatic properties in vitro, and it significantly suppressed tumor growth and metastasis in vivo. The underlying molecular mechanisms included (i) impairment of iron-sulfur [Fe-S] cluster/heme biogenesis, leading to destabilization and loss of activity of [Fe-S] cluster/heme containing enzymes, (ii) inhibition of mitochondrial respiration leading to mitochondrial reactive oxygen species production, resulting in dysfunctional mitochondria with markedly reduced supercomplexes, and (iii) fragmentation of the mitochondrial network and induction of mitophagy. Mitochondrial targeting of deferoxamine represents a way to deprive cancer cells of biologically active iron, which is incompatible with their proliferation and invasion, without disrupting systemic iron metabolism. Our findings highlight the importance of mitochondrial iron metabolism for cancer cells and demonstrate repurposing deferoxamine into an effective anticancer drug via mitochondrial targeting. SIGNIFICANCE These findings show that targeting the iron chelator deferoxamine to mitochondria impairs mitochondrial respiration and biogenesis of [Fe-S] clusters/heme in cancer cells, which suppresses proliferation and migration and induces cell death. GRAPHICAL ABSTRACT http//cancerres.aacrjournals.org/content/canres/81/9/2289/F1.large.jpg. Our 2011 report, reflecting data from 2007-2008, demonstrated that, for many pediatric subspecialties, pursuing fellowship training was a negative financial decision when compared with practicing as a general pediatrician. We provide an updated analysis on the financial impact of pediatric fellowship training and model interventions that can influence the results. We estimated the financial returns a graduating pediatric resident might anticipate from fellowship training followed by a career as a pediatric subspecialist and compared them with the returns expected from starting a career as a general pediatrician immediately after residency. We evaluated the potential effects of eliminating medical school debt, shortening the length of fellowship training, and implementing a federal loan repayment program for pediatric subspecialists. We compared the financial returns of subspecialty training in 2018-2019 to those from our previous report. Pursuing fellowship training generated widely variable financial returns when compared with general pediatrics that ranged from +$852 129 for cardiology to -$1 594 366 for adolescent medicine. Twelve of 15 subspecialties analyzed yielded negative financial returns. The differences have become more pronounced over time the spread between the highest and lowest earning subspecialties widened from >$1.4 million in 2007-2008 to >$2.3 million in 2018-2019. The negative financial impact of fellowship training could be partially ameliorated by shortening the length of training or by implementing pediatric subspecialist specific loan repayment programs. This report can be used to help guide trainees, educators, and policy makers. The interventions discussed could help maintain an adequate and balanced pediatric workforce.This report can be used to help guide trainees, educators, and policy makers. The interventions discussed could help maintain an adequate and balanced pediatric workforce.