Engineering Large Chromosomal Deletions in Mice to Advance Precision Oncology

Project Summary/Abstract
Genomic deletions, including both focal and large arm-level chromosomal deletions, are frequent genetic events
that promote cancer initiation and progression. Focal deletions have yielded significant insights into mechanisms
underlying tumorigenesis by helping identify tumor suppressor genes (TSGs) residing in the deleted loci. In
contrast, little is known about the biological and phenotypic impacts of large chromosomal deletions in cancer
despite their occurrence 30 times more frequently than focal deletions. We and other researchers have shown
that such deletions impact the activities of multiple neighboring TSGs and constitute a driving force in
tumorigenesis. Additionally, concomitant loss of multiple genes triggered by broad-scale deletions can create
phenotypes that differ fundamentally from those arising from the loss of a single TSG, and may offer
unanticipated therapeutic opportunities. These losses should therefore be considered as distinct genetic events
and their analysis should be focused on the integrated effects of chromosomal deletion rather than the impact of
a single critical TSG. Genetically engineered mouse (GEM) models provide an ideal tool for investigating the
consequences of genetic aberrations in tumor biology. However, due to the limited cloning capacity of targeting
vectors (~300 Kb) and the rarity of on-target homologous recombination events in traditional gene-targeting
technology, modelling large chromosomal deletions in mice has proven highly challenging, and the in vivo role
of such lesions in tumorigenesis has therefore been significantly understudied. To fill this critical gap, we have
developed an innovative approach to chromosomal engineering over large genetic distances through
CRISPR/Cas9 technology in mouse embryonic stem (ES) cells. The resulting ES clones carrying the desired
genome edits make possible the creation of conditional knockout mice that accurately mimic cancer-associated
large deletions. Given the prevalence and significance of large chromosomal deletions in prostate cancer, here
we propose to utilize our innovative approach to generate and fully characterize two novel GEM models that
each harbor a common but distinct prostate cancer-associated large deletion, with a goal of validating and
credentialing these models as genetically and biologically robust representations of human prostate cancer. In
Aim 1, we will develop conditional mouse lines using CRISPR/Cas9 technology to target commonly deleted large
chromosomal loci in human prostate cancer.
In Aim 2, we will develop mouse models of prostate cancer harboring
large chromosomal deletions to analyze their biological and phenotypic impacts on prostate cancer development.
Taken together, these models and their characterization will meet multiple goals of this grant opportunity.
Successful completion of these investigations will not only contribute greatly to the implementation of precision
oncology research but also enhance the applicability of animal models to translational research.

Grant Number: 5R01CA261949-04
NIH Institute/Center: NIH
Principal Investigator: MING CHEN