Effects of 2-Chlorodeoxyadenosine (Cladribine) on Primary Rat Microglia

Abstract

2-chlorodeoxyadenosine (CdA, Cladribine) is an immunosuppressant that has recently been shown to be effective in the treatment of multiple sclerosis (MS). There is extensive clinical experience with CdA for the treatment of neoplastic diseases, especially hematologic malignancies, due to its apoptotic effects on leukemic and several other neoplastic cells. Furthermore, CdA crosses the blood-brain-barrier and thus may also exert its effects directly on cells of the central nervous system (CNS). Therefore, we have studied the effects of CdA on cultured primary rat microglia, the resident macrophage in the CNS, which is also thought to be involved in the pathogenesis of MS. Treatment of microglia with CdA inhibited their proliferation and induced apoptosis. Phosphorylation of CdA to CdATP was required for both effects and was inhibited by deoxycytidine. Furthermore, activation of caspase-3 and -9 revealed the involvement of the intrinsic mitochondrial mediated apoptotic pathway. However, the absence of caspase-8 activation specified independency from the extrinsic death receptor mediated apoptosis. The mitochondrial membrane potential was significantly reduced after CdA exposure and was not conserved with Bax or caspase-3 inhibition. Assessment of DNA fragmentation by TUNEL and DNA-release-assay showed microglia with fragmented nuclei. Other functions of microglia like phagocytosis and LPS-induced NO and TNF-α release were not affected by CdA. These data suggest a potential of CdA treatment to induce not only leukopenia but also apoptosis in microglia in the CNS. These results help to understand the mechanism of action of CdA in CNS diseases and may open the possibility to target microglia.