Epigenetic regulators have been implicated in tumorigenesis of many types of cancer; however, their roles in endothelial cell cancers such as canine hemangiosarcoma (HSA) have not been studied. In this study, we found that lysine-specific demethylase 2B (Kdm2b) was highly expressed in HSA cell lines compared to normal canine endothelial cells. Silencing of Kdm2b in HSA cells resulted to increased cell death in vitro compared to the scramble control by inducing apoptosis through the inactivation of the DNA repair pathways and accumulation of DNA damage. Similarly, doxycycline-induced Kdm2b silencing in tumor xenografts resulted to decreased tumor sizes compared to the scramble control. Furthermore, Kdm2b was also highly expressed in clinical cases of HSA, and its expression levels was higher than in hemangioma, a benign counterpart of HSA. Based on these results, we hypothesized that pharmacological Kdm2b inhibition can also induce HSA cell death and can be used as an alternative treatment for HSA. We treated HSA cells with GSK-J4, a histone demethylase inhibitor, and found that GSK-J4 treatment also induced apoptosis and cell death. On top of that, GSK-J4 treatment in HSA tumor-bearing mice decreased tumor sizes without any obvious side-effects. In this study, we demonstrated that Kdm2b acts as an oncogene in HSA by enhancing DNA damage response and can be used as a biomarker to differentiate HSA from hemangioma. Moreover, we indicated that histone demethylase inhibitor GSK-J4 can be used as a therapeutic alternative to doxorubicin for HSA treatment.

Canine hemangiosarcoma (HSA) is one of the most common mesenchymal tumors in dogs. Its high metastatic and growth rates are usually associated with poor prognosis. Neoplastic cells of HSA can show various levels of cellular atypia in the same mass and may consist of various populations at different differentiated stages. Up to present, however, there is no report analyzing their differentiation states by comparing cellular atypia with differentiation-related protein expressions. To evaluate whether cellular atypia can be used as a differentiation marker in HSA, we analyzed correlation between cellular atypia and intensities of CD31 and von Willebrand Factor (vWF) staining in HSA cases. We also compared cellular atypia and expression levels of CD31 and vWF in each growth patterns. Our results show that cellular atypia was negatively correlated to CD31 and vWF expression levels but no significant correlation was found between growth patterns and cellular atypia or CD31 and vWF expression levels. Our study suggests that cellular atypia is useful for identifying differentiation levels in HSA cases. This study also provides useful information to determine differentiation levels of cell populations within HSA based only on morphological analysis, which will aid further HSA research such as identifying undifferentiation markers of endothelial cells or finding undifferentiated cell population in tissue sections.

BackgroundHemangiosarcoma (HSA) is a malignant tumor derived from endothelial cells which usually shows poor prognosis due to its high invasiveness, metastatic rate and severe hemorrhage from tumor ruptures. Since the pathogenesis of HSA is not yet complete, further understanding of its molecular basis is required.ResultsHere, we identified Notch2 signal as a key factor in maintaining canine HSA cancer stem cell (CSC)-like cells. We first cultured HSA cell lines in adherent serum-free condition and confirmed their CSC-like characteristics. Notch signal was upregulated in the CSC-like cells and Notch signal inhibition by a γ-secretase inhibitor significantly repressed their growth. Notch2, a Notch receptor, was highly expressed in the CSC-like cells. Constitutive activation of Notch2 increased clonogenicity and number of cells which were able to survive in serum-free condition. In contrast, inhibition of Notch2 activity showed opposite effects. These results suggest that Notch2 is an important factor for maintaining HSA CSC-like cells. Neoplastic cells in clinical cases also express Notch2 higher than endothelial cells in the normal blood vessels in the same slides.ConclusionThis study provides foundation for further stem cell research in HSA and can provide a way to develop effective treatments to CSCs of endothelial tumors.Electronic supplementary materialThe online version of this article (10.1186/s12917-018-1624-8) contains supplementary material, which is available to authorized users.

Pancreatic ductal adenocarcinoma (PDAC) is a lethal cancer with a low survival rate. Recently, new drugs that target KRASG12D, a common mutation in PDAC, have been developed. We studied one of these compounds, MRTX1133, and found it was specific and effective at low nanomolar concentrations in patient-derived organoid models and cell lines harboring KRASG12D mutations. Treatment with MRTX1133 upregulated the expression and phosphorylation of EGFR and HER2, indicating that inhibition of ERBB signaling may potentiate MRTX1133 anti-tumor activity. Indeed, the irreversible pan-ERBB inhibitor, afatinib, potently synergized with MRTX1133 in vitro, and cancer cells with acquired resistance to MRTX1133 in vitro remained sensitive to this combination therapy. Finally, the combination of MRTX1133 and afatinib led to tumor regression and longer survival in orthotopic PDAC mouse models. These results suggest that dual inhibition of ERBB and KRAS signaling may be synergistic and circumvent the rapid development of acquired resistance in patients with KRAS mutant pancreatic cancer.

Epigenetic regulators have been implicated in tumorigenesis of many types of cancer; however, their roles in endothelial cell cancers such as canine hemangiosarcoma (HSA) have not been studied. In this study, we found that lysine-specific demethylase 2B (Kdm2b) was highly expressed in HSA cell lines compared to normal canine endothelial cells. Silencing of Kdm2b in HSA cells resulted to increased cell death in vitro compared to the scramble control by inducing apoptosis through the inactivation of the DNA repair pathways and accumulation of DNA damage. Similarly, doxycycline-induced Kdm2b silencing in tumor xenografts resulted to decreased tumor sizes compared to the scramble control. Furthermore, Kdm2b was also highly expressed in clinical cases of HSA, and its expression levels was higher than in hemangioma, a benign counterpart of HSA. Based on these results, we hypothesized that pharmacological Kdm2b inhibition can also induce HSA cell death and can be used as an alternative treatment for HSA. We treated HSA cells with GSK-J4, a histone demethylase inhibitor, and found that GSK-J4 treatment also induced apoptosis and cell death. On top of that, GSK-J4 treatment in HSA tumor-bearing mice decreased tumor sizes without any obvious side-effects. In this study, we demonstrated that Kdm2b acts as an oncogene in HSA by enhancing DNA damage response and can be used as a biomarker to differentiate HSA from hemangioma. Moreover, we indicated that histone demethylase inhibitor GSK-J4 can be used as a therapeutic alternative to doxorubicin for HSA treatment.