Establishment and characterization of a canine soft tissue sarcoma patient-derived xenograft model.

Spontaneously occurring soft tissue sarcoma (STS) is relatively common in canine cancer patients. Because of the similarities to human disease, canine STSs are a valuable and readily available resource for the study of new therapeutics. In this study, a canine patient-derived xenograft (PDX) model, CDX-STS2, was established. The CDX-STS2 model was engrafted and expanded for systemic administration studies with chemotherapeutic agents commonly used to treat STS, including doxorubicin, docetaxel and gemcitabine. Immunohistochemistry for drug-specific biomarkers and tumour growth measurement revealed tumour sensitivity to doxorubicin and docetaxel, whereas gemcitabine had no effect on tumour growth. Although many human PDX tumour models have been established, relatively few canine PDX models have been reported to date. CDX-STS2 represents a new STS PDX research model of canine origin that will be useful in bridging preclinical research with clinical studies of STS in pet dogs.

TGF-ß signaling alters the pattern of liver tumorigenesis induced by Pten inactivation.

Hepatocarcinogenesis results from the accumulation of genetic and epigenetic changes in liver cells. A common mechanism through which these alterations induce liver cancer is by deregulating signaling pathways. A number of signaling pathways, including the PI3K/PTEN/AKT and transforming growth factor ß (TGF-ß) pathways have been implicated in normal liver development as well as in cancer formation. In this study, we assessed the effect of the TGF-ß signaling pathway on liver tumors induced by phosphatase and tensin homolog (Pten) loss. Inactivation of only the TGF-ß receptor type II, Tgfbr2, in the mouse liver (Tgfbr2(LKO)) had no overt phenotype, while inactivation of Pten alone (Pten(LKO)), resulted in the formation of both hepatocellular carcinomas and cholangiocarcinomas (CC). Interestingly, deletion of both Pten and Tgfbr2 (Pten(LKO);Tgfbr2(LKO)) in the mouse liver resulted in a dramatic shift in tumor type to predominantly CC. Assessment of the PI3K/PTEN/AKT pathway revealed increased phosphorylation of AKT and glycogen synthase kinase 3 beta (GSK-3ß) in both the Pten(LKO) and Pten(LKO);Tgfbr2(LKO) mice, suggesting that this pathway is constitutively active regardless of the status of the TGF-ß signaling pathway. However, phosphorylation of p70 S6 kinase was observed in the liver of all three phenotypes (Tgfbr2(LKO), Pten(LKO), Pten(LKO);Tgfbr2(LKO)) indicating that the loss of Tgfbr2 and/or Pten leads to an increase in this signaling pathway. Analysis of markers of liver progenitor/stem cells revealed that the loss of TGF-ß signaling resulted in increased expression of c-Kit and CD133. Furthermore, in addition to increased c-Kit and CD133, Scf and EpCam expression were also increased in the double knock-out mice. These results suggest that the alteration in tumor types between the Pten(LKO) mice and Pten(LKO);Tgfbr2(LKO) mice is secondary to the altered regulation of stem-cell features induced by the loss of TGF-ß signaling.

Inactivation of TGF-ß signaling and loss of PTEN cooperate to induce colon cancer in vivo.

The accumulation of genetic and epigenetic alterations mediates colorectal cancer (CRC) formation by deregulating key signaling pathways in cancer cells. In CRC, one of the most commonly inactivated signaling pathways is the transforming growth factor-beta (TGF-ß) signaling pathway, which is often inactivated by mutations of TGF-ß type II receptor (TGFBR2). Another commonly deregulated pathway in CRC is the phosphoinositide-3-kinase (PI3K)-AKT pathway. Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is an important negative regulator of PI3K-AKT signaling and is silenced in ∼30% of CRC. The combination of TGFBR2 inactivation and loss of PTEN is particularly common in microsatellite-unstable CRCs. Consequently, we determined in vivo if deregulation of these two pathways cooperates to affect CRC formation by analyzing tumors arising in mice that lack Tgfbr2 and/or Pten specifically in the intestinal epithelium. We found that lack of Tgfbr2 (Tgfbr2(IEKO)) alone is not sufficient for intestinal tumor formation and lack of Pten (Pten(IEKO)) alone had a weak effect on intestinal tumor induction. However, the combination of Tgfbr2 inactivation with Pten loss (Pten(IEKO);Tgfbr2(IEKO)) led to malignant tumors in both the small intestine and colon in 86% of the mice and to metastases in 8% of the tumor-bearing mice. Moreover, these tumors arose via a ß-catenin-independent mechanism. Inactivation of TGF-ß signaling and loss of Pten in the tumors led to increased cell proliferation, decreased apoptosis and decreased expression of cyclin-dependent kinase inhibitors. Thus, inactivation of TGF-ß signaling and loss of PTEN cooperate to drive intestinal cancer formation and progression by suppressing cell cycle inhibitors.

Inhibition of PI-3K restores nuclear p27Kip1 expression in a mouse model of Kras-driven lung cancer.

Reduced expression of the CDK inhibitor p27(Kip1) (p27) in human lung cancer correlates with tumor aggressiveness and poor prognosis. However, the regulation of p27 expression and the role of p27 during lung cancer are poorly understood. Urethane-induced lung tumors in mice frequently harbor mutations in the Kras oncogene, and in this study, we use this model to address the regulation of p27 during tumorigenesis. The Ras effector Akt is known to regulate p27 mRNA abundance by phosphorylating and inactivating the FOXO transcription factors. Phosphorylated Akt and FOXO proteins were both increased in lung tumors, correlating with a reduction in p27 mRNA transcript. Akt also directly phosphorylates p27 and regulates its nuclear/cytoplasmic localization. Tumors showed a reduced nuclear/cytoplasmic ratio of p27 protein, together with an increase in phosphorylated Thr197 p27 in the cytoplasmic pool. Treatment of lung tumor-bearing mice with the phosphoinositol-3 kinase inhibitor LY294002 induced a rapid decrease in phosphorylated Akt and phosphorylated p27, concomitant with an increase in nuclear p27. Germline p27 deficiency accelerated both the growth and malignant progression of urethane-induced lung tumors, and did so in a cell autonomous manner, confirming a causal role of p27 in tumor suppression. These results show that p27 is a potent barrier to the growth and malignant progression of Kras-initiated lung tumors. Further, the reduction of nuclear p27 in tumors is mediated by oncogene signaling pathways, which can be reversed by pharmacological agents.