In vitro effect of chlorambucil on human glioma cell lines (SF767 and U87-MG), and human microvascular endothelial cell (HMVEC) and endothelial progenitor cells (ECFCs), in the context of plasma chlorambucil concentrations in tumor-bearing dogs.

Michael J Reese,Deborah W Knapp,Julie A Mund,Jamie Case,David R Jones,Kimberly M Anderson,Rebecca A Packer

doi:10.1371/journal.pone.0203517

Abstract

The objective of this study was to investigate a possible mechanism of action of metronomic chlorambucil on glioma by studying the in vitro cytotoxicity and anti-angiogenic effects on glioma and endothelial cells, respectively. The in vitro LD50 and IC50 of chlorambucil were determined using human SF767 and U87-MG glioma cell lines, human microvascular endothelial cells (HMVECs) and human endothelial colony forming cells (ECFCs). Results were analyzed in the context of chlorambucil concentrations measured in the plasma of tumor-bearing dogs receiving 4 mg m-2 metronomic chlorambucil. The LD50 and IC50 of chlorambucil were 270 μM and 114 μM for SF767, and 390 μM and 96 μM for U87-MG, respectively. The IC50 of chlorambucil was 0.53 μM and 145 μM for the HMVECs and ECFCs, respectively. In pharmacokinetic studies, the mean plasma Cmax of chlorambucil was 0.06 μM. Results suggest that metronomic chlorambucil in dogs does not achieve plasma concentrations high enough to cause direct cytotoxic or growth inhibitory effects on either glioma or endothelial cells.

Highlights

Gliomas are common primary brain tumors in both human and veterinary medicine
The concentration of chlorambucil necessary to inhibit human endothelial cell proliferation was generally lower than the concentration necessary to inhibit human glioma cell proliferation and cause direct cytotoxicity
The human microvascular endothelial cells (HMVECs) and endothelial colony forming cells (ECFCs) differed in their response to chlorambucil, which may be due to differences in maturity of endothelial cell types

Summary

Introduction

Gliomas are common primary brain tumors in both human and veterinary medicine. A diagnosis of a high-grade glioma carries a poor prognosis due to frequent disease recurrence [1,2]. One of the intriguing new approaches to cancer treatment is the use of relatively low, frequent dosing of chemotherapy agents, in a “metronomic” fashion to target tumor angiogenesis [5,6,7,8]. This contrasts to conventional chemotherapeutic protocols that utilize a maximum tolerated dose of drug given with prolonged breaks in between dosing to directly have an effect on tumor cells. The mechanism is likely multifactorial, including targeting tumor angiogenesis through a direct effect on activated vascular endothelial cells and circulating endothelial progenitor cells, but it may be mediated by prostaglandins, thrombospondin 1, growth factors, as well as other potential mediators of vasculogenesis [5,7,8,9]

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