Modeling distinct osteosarcoma subtypes in vivo using Cre:lox and lineage-restricted transgenic shRNA

Abstract

Osteosarcoma is the most common primary cancer of bone and one that predominantly affects children and adolescents. Osteoblastic osteosarcoma represents the major subtype of this tumor, with approximately equal representation of fibroblastic and chondroblastic subtypes. We and others have previously described murine models of osteosarcoma based on osteoblast-restricted Cre:lox deletion of Trp53 (p53) and Rb1 (Rb), resulting in a phenotype most similar to fibroblastic osteosarcoma in humans. We now report a model of the most prevalent form of human osteosarcoma, the osteoblastic subtype. In contrast to other osteosarcoma models that have used Cre:lox mediated gene deletion, this model was generated through shRNA-based knockdown of p53. As is the case with the human disease the shRNA tumors most frequently present in the long bones and preferentially disseminate to the lungs; feature less consistently modeled using Cre:lox approaches. Our approach allowed direct comparison of the in vivo consequences of targeting the same genetic drivers using two different technologies, Cre:lox and shRNA. This demonstrated that the effects of Cre:lox and shRNA mediated knock-down are qualitatively different, at least in the context of osteosarcoma, and yielded distinct subtypes of osteosarcoma. Through the use of complementary genetic modification strategies we have established a model of the most common clinical subtype of osteosarcoma that was not previously represented and more fully recapitulated the clinical spectrum of this cancer.

Systematic Screening Identifies Dual PI3K and mTOR Inhibition as a Conserved Therapeutic Vulnerability in Osteosarcoma

“…2B). Analysis of the hits interacting with doxorubicin revealed a marked enrichment of candidates on chromosome 7 (P ¼ 4.77 Â 10 À11 , FDR ¼ 3.68 Â 10 À8 ; statistics derived from DAVID analysis), relevant as the loss of one to two copies of chromosome 7 from a tetraploid karyotype was recurrently noted in the murine osteosarcoma models (16). Using Ingenuity Pathway Analysis software, we tested whether the identified hits functioned in overlapping pathways/gene-expression signatures ( Fig.…”

“…To identify new therapeutic targets for osteosarcoma, we have made use of murine osteosarcoma models combined with primary human osteosarcoma xenograft derived cell cultures (13,16). These independent approaches converged to identify dual inhibition of the PI3K-mTOR pathways as a species conserved sensitivity in osteosarcoma.…”

“…We previously developed a GEMM of the fibroblastic osteosarcoma subtype, through deletion of p53 and pRb from the osteoblast lineage that has been independently validated (13)(14)(15). We recently described the first murine model of osteoblastic osteosarcoma, the most common subtype of human osteosarcoma (16).…”

“…Here, we report results from parallel screens using primary cell cultures derived from murine osteosarcoma models that faithfully replicate the human disease (13,16). First, a wholegenome siRNA screen for enhancers of cell death was performed.…”

Animal Models in Osteosarcoma

“…Although the greatest proportion of tumors was OS when P53 was conditionally deleted, neuroendocrine tumors and hibernomas were also reported to be generated in several mice (67, 68). However, Walkley et al enriched the C57BL/6 background of the mouse strain and the percentage of hibernomas was reduced, suggesting a possible impact of mouse strains in the phenotype observed (69). A recent study in mice that expressed SV40 T/t antigen (Tag) in mature osteoblasts under the Og2 (74) showed OS with complete penetrance (71) and 90% incidence of lung metastases.…”