Cancer genomics studies have nominated thousands of putative cancer driver genes
1
; a major challenge is to develop high-throughput and accurate models to define their functions. Here we devised a scalable cancer spheroid model and performed genome-wide CRISPR screens in 2D-monolayers and 3D lung cancer spheroids. CRISPR phenotypes in 3D more accurately recapitulate those of
in vivo
tumors, and genes with differential sensitivities between 2D and 3D are strongly enriched for significant mutations in lung cancers. These analyses also revealed novel drivers essential for cancer growth in 3D and
in vivo
, but not in 2D. Notably, we discovered that CPD (Carboxypeptidase D) is responsible for removal of a c-terminal RKRR motif
2
of IGF1R α-chain, critical for receptor activity. CPD expression correlates with patient outcomes in lung cancer, and loss of CPD reduced tumor growth. Our results reveal key differences between 2D and 3D cancer models, and establish a generalizable strategy to perform CRISPR screens in spheroids to uncover cancer vulnerabilities.
The intestinal epithelium is a rapidly renewing cellular compartment. This constant regeneration is a hallmark of intestinal homeostasis and requires a tightly regulated balance between intestinal stem cell (ISC) proliferation and differentiation. Since intestinal epithelial cells directly contact pathogenic environmental factors that continuously challenge their integrity, ISCs must also actively divide to facilitate regeneration and repair. Understanding niche adaptations that maintain ISC activity during homeostatic renewal and injury-induced intestinal regeneration is therefore a major and ongoing focus for stem cell biology. Here, we review recent concepts and propose an active interconversion of the ISC niche between homeostasis and injury-adaptive states that is superimposed upon an equally dynamic equilibrium between active and reserve ISC populations.
The characterization of cancer genomes has provided insight into somatically altered genes across tumors, transformed our understanding of cancer biology, and enabled tailoring of therapeutic strategies. However, the function of most cancer alleles remains mysterious, and many cancer features transcend their genomes. Consequently, tumor genomic characterization does not influence therapy for most patients. Approaches to understand the function and circuitry of cancer genes provide complementary approaches to elucidate both oncogene and non-oncogene dependencies. Emerging work indicates that the diversity of therapeutic targets engendered by non-oncogene dependencies is much larger than the list of recurrently mutated genes. Here we describe a framework for this expanded list of cancer targets, providing novel opportunities for clinical translation.
Mutations in ARID1A rank amongst the most common molecular aberrations in human cancer.However, oncogenic consequences of ARID1A mutation in human cells remain poorly defined due to lack of forward genetic models. Here, CRISPR/Cas9-mediated ARID1A knockout in primary TP53 -/human gastric organoids induced morphologic dysplasia, tumorigenicity and mucinous differentiation. Genetic Wnt/-catenin activation rescued mucinous differentiation, but not hyperproliferation, suggesting alternative pathways of ARID1A KO-mediated transformation.ARID1A mutation induced transcriptional regulatory modules characteristic of MSI and EBV subtype human gastric cancer, including FOXM1-associated mitotic genes and BIRC5/survivin.Convergently, high-throughput compound screening indicated selective vulnerability of ARID1Adeficient organoids to inhibition of BIRC5/survivin, functionally implicating this pathway as an essential mediator of ARID1A KO-dependent early-stage gastric tumorigenesis. Overall, we define distinct pathways downstream of oncogenic ARID1A mutation, with non-essential Wntinhibited mucinous differentiation in parallel with essential transcriptional FOXM1/BIRC5stimulated proliferation, illustrating the general utility of organoid-based forward genetic cancer analysis in human cells.Research.
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