KDM2B promotes cell viability by enhancing DNA damage response in canine hemangiosarcoma

Abstract

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.

Hemangiosarcoma cells induce M2 polarization and PD-L1 expression in macrophages

“…Given that more than 80% of clinical HSA cases that we examined in our study expressed PD-L1 in tumor cells and/or macrophages, immunotherapy using anti PD-L1 antibody treatment could be useful as an alternative treatment for canine HSA. In our previous study, silencing of KDM2B resulted to increased interferon gamma and alpha responses 7 . This means that KDM2B inhibition induces immune reaction; therefore, combination therapy with anti-PD-L1 antibody and KDM2B inhibition might provide better outcomes than single treatment with anti-PD-L1 or KDM2B inhibitor treatment in canine HSA.…”

“…All tumors were examined for KDM2B expression and presence of lymphocytes and macrophages using immunohistochemistry (IHC). IHC was performed using antibodies to Iba1 (1:2000, Fujifilm Wako, Osaka, Japan, 019-19741), CD3 (1:1000; Agilent Technologies, CA, USA, IR503), KDM2B (1:50; Santa Cruz Biotechnology, Inc.,sc-293279), CD31 (1:250; Abcam, JC/70A), PD-L1 (1:100; clone 6C11-3A11), and CD204 (1:800; Medicinal Chemistry Pharmaceutical Co., Ltd., Sapporo, Japan, KT022) as described previously 7 . Nano Zoomer 2.0-RS (Hamamatsu Photonics, Hamamatsu, Japan) was used to scan histological slides, which were then processed in QuPath ver 0.2.133 34 .…”

“…Recently, we found that canine HSA highly expressed three histone demethylases (KDM1A, KDM2A and KDM2B) out of which KDM2B was found necessary for HSA cell survival by positively regulating the DNA damage response system in tumor cells 7 . KDM2B silencing not only dysregulated DNA damage response but also induced expressions of the genes related to inflammatory responses 7 . Inhibiting KDM2B could be an option to induce host immune responses against HSA tumor cells.…”

“…An effective treatment for HSA is difficult to develop since little is known about its molecular pathology. Recently, we found that canine HSA highly expressed three histone demethylases (KDM1A, KDM2A and KDM2B) out of which KDM2B was found necessary for HSA cell survival by positively regulating the DNA damage response system in tumor cells 7 . KDM2B silencing not only dysregulated DNA damage response but also induced expressions of the genes related to inflammatory responses 7 .…”

Epigenetics of Cutaneous Sarcoma

“…KDM2B regulates various physiological and pathological processes, such as proliferation and oncogenesis [ 52 ]. KDM2B is upregulated in angiosarcomas [ 53 ], where it suppresses p53 and activates mTOR signaling (reviewed in [ 54 ]). KDM2B silencing in angiosarcoma cells enhances cell death via inactivation of DNA repair.…”