Efficient and multiplexed somatic genome editing with Cas12a mice.

Somatic genome editing in mouse models has increased our understanding of the in vivo effects of genetic alterations. However, existing models have a limited ability to create multiple targeted edits, hindering our understanding of complex genetic interactions. Here we generate transgenic mice with Cre-regulated and constitutive expression of enhanced Acidaminococcus sp.

Combinatorial In Vivo Genome Editing Identifies Widespread Epistasis and an Accessible Fitness Landscape During Lung Tumorigenesis.

Lung adenocarcinoma, the most common subtype of lung cancer, is genomically complex, with tumors containing tens to hundreds of non-synonymous mutations. However, little is understood about how genes interact with each other to enable the evolution of cancer in vivo, largely due to a lack of methods for investigating genetic interactions in a high-throughput and quantitative manner. Here, we employed a novel platform to generate tumors with inactivation of pairs of ten diverse tumor suppressor genes within an autochthonous mouse model of oncogenic KRAS-driven lung cancer.

A STAG2-PAXIP1/PAGR1 axis suppresses lung tumorigenesis.

The cohesin complex is a critical regulator of gene expression. STAG2 is the most frequently mutated cohesin subunit across several cancer types and is a key tumor suppressor in lung cancer. Here, we coupled somatic CRISPR-Cas9 genome editing and tumor barcoding with an autochthonous oncogenic KRAS-driven lung cancer model and showed that STAG2 is uniquely tumor-suppressive among all core and auxiliary cohesin components.

A STAG2-PAXIP1/PAGR1 axis suppresses lung tumorigenesis.

Unlabelled: The cohesin complex is a critical regulator of gene expression. is the most frequently mutated cohesin subunit across several cancer types and is a key tumor suppressor in lung cancer. Here, we coupled somatic CRISPR-Cas9 genome editing and tumor barcoding with an autochthonous oncogenic KRAS-driven lung cancer model and show that STAG2 is uniquely tumor suppressive among all core and auxiliary cohesin components.

Functional mapping of epigenetic regulators uncovers coordinated tumor suppression by the HBO1 and MLL1 complexes.

Epigenetic dysregulation is widespread in cancer. However, the specific epigenetic regulators and the processes they control to drive cancer phenotypes are poorly understood. Here, we employed a novel, scalable and high-throughput method to perform iterative functional screens of over 250 epigenetic regulatory genes within autochthonous oncogenic KRAS-driven lung tumors.

Aging represses lung tumorigenesis and alters tumor suppression.

Most cancers are diagnosed in persons over the age of sixty, but little is known about how age impacts tumorigenesis. While aging is accompanied by mutation accumulation - widely understood to contribute to cancer risk - it is also associated with numerous other cellular and molecular changes likely to impact tumorigenesis. Moreover, cancer incidence decreases in the oldest part of the population, suggesting that very old age may reduce carcinogenesis.

Combinatorial genome editing identifies widespread epistasis during lung tumorigenesis.

Lung adenocarcinoma, the most common subtype of lung cancer, is genomically complex, with tumors containing tens to hundreds of non-synonymous mutations. However, little is understood about how genes interact with each other to enable tumorigenesis , largely due to a lack of methods for investigating genetic interactions in a high-throughput and multiplexed manner. Here, we employed a novel platform to generate tumors with all pairwise inactivation of ten tumor suppressor genes within an autochthonous mouse model of oncogenic KRAS-driven lung cancer.

Efficient and multiplexed somatic genome editing with Cas12a mice.

Somatic genome editing in mouse models has increased our understanding of the effects of genetic alterations in areas ranging from neuroscience to cancer biology and beyond. However, existing models are limited in their ability to create multiple targeted edits. Thus, our understanding of the complex genetic interactions that underlie development, homeostasis, and disease remains incomplete.

Dissecting metastasis using preclinical models and methods.

Metastasis has long been understood to lead to the overwhelming majority of cancer-related deaths. However, our understanding of the metastatic process, and thus our ability to prevent or eliminate metastases, remains frustratingly limited. This is largely due to the complexity of metastasis, which is a multistep process that likely differs across cancer types and is greatly influenced by many aspects of the in vivo microenvironment.

Multiplexed screens identify RAS paralogues HRAS and NRAS as suppressors of KRAS-driven lung cancer growth.

Oncogenic KRAS mutations occur in approximately 30% of lung adenocarcinoma. Despite several decades of effort, oncogenic KRAS-driven lung cancer remains difficult to treat, and our understanding of the regulators of RAS signalling is incomplete. Here to uncover the impact of diverse KRAS-interacting proteins on lung cancer growth, we combined multiplexed somatic CRISPR/Cas9-based genome editing in genetically engineered mouse models with tumour barcoding and high-throughput barcode sequencing.

Combinatorial Inactivation of Tumor Suppressors Efficiently Initiates Lung Adenocarcinoma with Therapeutic Vulnerabilities.

Unlabelled: Lung cancer is the leading cause of cancer death worldwide, with lung adenocarcinoma being the most common subtype. Many oncogenes and tumor suppressor genes are altered in this cancer type, and the discovery of oncogene mutations has led to the development of targeted therapies that have improved clinical outcomes. However, a large fraction of lung adenocarcinomas lacks mutations in known oncogenes, and the genesis and treatment of these oncogene-negative tumors remain enigmatic.

The scaffold protein IQGAP1 is crucial for extravasation and metastasis.

IQGAP1 is a scaffold protein involved in a range of cellular activities, including migration, invasion, adhesion and proliferation. It is also oncogenic in a variety of cancers, promoting primary tumor growth and invasiveness. However, the role of IQGAP1 in tumor progression and metastasis remains unclear.

Proteomic Profiling of the ECM of Xenograft Breast Cancer Metastases in Different Organs Reveals Distinct Metastatic Niches.

Metastasis causes most cancer-related deaths, and one poorly understood aspect of metastatic cancer is the adaptability of cells from a primary tumor to create new niches and survive in multiple, different secondary sites. We used quantitative mass spectrometry to analyze the extracellular matrix (ECM), a critical component of metastatic niches, in metastases to the brain, lungs, liver, and bone marrow, all derived from parental MDA-MB-231 triple-negative breast cancer cells. Tumor and stromal cells cooperated in forming niches; stromal cells produced predominantly core, structural ECM proteins and tumor cells produced a diverse array of ECM-associated proteins, including secreted factors and modulators of the matrix.

Macrophage-Secreted TNFα and TGFβ1 Influence Migration Speed and Persistence of Cancer Cells in 3D Tissue Culture via Independent Pathways.

The ability of a cancer cell to migrate through the dense extracellular matrix within and surrounding the solid tumor is a critical determinant of metastasis. Macrophages enhance invasion and metastasis in the tumor microenvironment, but the basis for their effects is not fully understood. Using a microfluidic 3D cell migration assay, we found that the presence of macrophages enhanced the speed and persistence of cancer cell migration through a 3D extracellular matrix in a matrix metalloproteinases (MMP)-dependent fashion.

Conditionally reprogrammed cells represent a stem-like state of adult epithelial cells.

The combination of irradiated fibroblast feeder cells and Rho kinase inhibitor, Y-27632, conditionally induces an indefinite proliferative state in primary mammalian epithelial cells. These conditionally reprogrammed cells (CRCs) are karyotype-stable and nontumorigenic. Because self-renewal is a recognized property of stem cells, we investigated whether Y-27632 and feeder cells induced a stem-like phenotype.

The human papillomavirus type 16 E5 oncoprotein translocates calpactin I to the perinuclear region.

The human papillomavirus type 16 (HPV-16) E5 oncoprotein is embedded in membranes of the endoplasmic reticulum and nuclear envelope with its C terminus exposed to the cytoplasm. Among other activities, E5 cooperates with the HPV E6 oncoprotein to induce koilocytosis in human cervical cells and keratinocytes in vitro. The effect of E5 on infected cells may rely on its interactions with various cellular proteins.

The human papillomavirus type 16 E5 oncoprotein inhibits epidermal growth factor trafficking independently of endosome acidification.

The human papillomavirus type 16 E5 oncoprotein (16E5) enhances acute, ligand-dependent activation of the epidermal growth factor receptor (EGFR) and concomitantly alkalinizes endosomes, presumably by binding to the 16-kDa "c" subunit of the V-ATPase proton pump (16K) and inhibiting V-ATPase function. However, the relationship between 16K binding, endosome alkalinization, and altered EGFR signaling remains unclear. Using an antibody that we generated against 16K, we found that 16E5 associated with only a small fraction of endogenous 16K in keratinocytes, suggesting that it was unlikely that E5 could significantly affect V-ATPase function by direct inhibition.