Targeting Cholesterol Biosynthesis with Statins Synergizes with AKT Inhibitors in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is responsible for a disproportionate number of breast cancer patient deaths due to extensive molecular heterogeneity, high recurrence rates and lack of targeted therapies. Dysregulation of the phosphoinositide 3-kinase (PI3K)/AKT pathway occurs in approximately 50% of TNBC patients. Here, we performed a genome-wide CRISPR/Cas9 screen with PI3Kα and AKT inhibitors to find targetable synthetic lethalities in TNBC. Cholesterol homeostasis was identified as a collateral vulnerability with AKT inhibition. Disruption of cholesterol homeostasis with pitavastatin synergized with AKT inhibition to induce TNBC cytotoxicity in vitro, in mouse TNBC xenografts and in patient-derived, estrogen receptor (ER)-negative breast cancer organoids. Neither ER-positive breast cancer cell lines nor ER-positive organoids were sensitive to combined AKT inhibitor and pitavastatin. Mechanistically, TNBC cells showed impaired sterol regulatory element-binding protein 2 (SREBP-2) activation in response to single agent or combination treatment with AKT inhibitor and pitavastatin, which was rescued by inhibition of the cholesterol trafficking protein Niemann-Pick C1 (NPC1). NPC1 loss caused lysosomal cholesterol accumulation, decreased endoplasmic reticulum cholesterol levels, and promoted SREBP-2 activation. Taken together, these data identify a TNBC-specific vulnerability to the combination of AKT inhibitors and pitavastatin mediated by dysregulated cholesterol trafficking. These findings support combining AKT inhibitors with pitavastatin as a therapeutic modality in TNBC. .

Sequential apoptotic and multiplexed proteomic evaluation of single cancer cells.

A potential cause of cancer relapse is pretreatment chemoresistant subpopulations. Identifying targetable features of subpopulations that are poorly primed for therapy-induced cell death may improve cancer therapy. Here, we develop and validate real-time BH3 profiling, a live and functional single-cell measurement of pretreatment apoptotic sensitivity that occurs upstream of apoptotic protease activation. On the same single cells, we perform cyclic immunofluorescence, which enables multiplexed immunofluorescence of more than 30 proteins on the same cell. Using cultured cells and rapid ex vivo cultures of colon cancer patient-derived xenograft (PDX) models, we identify Bak as a univariate correlate of apoptotic priming, find that poorly primed subpopulations can correspond to specific stages of the cell cycle, and, in some PDX models, identify increased expression of Bcl-XL, Mcl-1, or Her2 in subpopulations that are poorly primed for apoptosis. Last, we generate and validate mathematical models of single-cell priming that describe how targetable proteins contribute to apoptotic priming.

Developmentally upregulated transcriptional elongation factor a like 3 suppresses axon regeneration after optic nerve injury.

Projection neurons of the mammalian central nervous system (CNS) do not spontaneously regenerate axons which have been damaged by an injury or disease, often leaving patients with permanent disabilities that affect motor, cognitive, or sensory functions. Although several molecular targets which promote some extent of axon regeneration in animal models have been identified, the resulting recovery is very limited, and the molecular mechanisms underlying the axonal regenerative failure in the CNS are still poorly understood. One of the most studied targets for axon regeneration in the CNS is the mTOR pathway. A number of developmentally regulated genes also have been found to play a role in CNS axon regeneration. Here, we found that Transcriptional Elongation Factor A Like 3 (Tceal3), belonging to the Bex/Tceal transcriptional regulator family, which also modulates the mTOR pathway, is developmentally upregulated in retinal ganglion cell (RGCs) projection CNS neurons, and suppresses their capacity to regenerate axons after injury.