Functional analysis of apoptosis induction in acute myeloid leukaemia-relevance of karyotype and clinical treatment response

Abstract

Deficiencies or structural defects of the apoptotic machinery have been postulated as a potential mechanism for a broad resistance of acute myeloid leukaemia (AML) blasts towards cytotoxic therapy comprising chemotherapeutic agents with diverse pharmacodynamic principles but also cell-mediated cytotoxicity of the graft-versus-leukaemia effect, for example, in the setting of allogeneic transplantation. This hypothesis was systematically tested by functionally analysing the early, intermediate and late events of the apoptotic process in primary AML (n = 31) blasts following activation of the intrinsic and extrinsic pathway of apoptosis (etoposide and cytarabine as DNA damaging agents, FAS-ligand as an activator of the death receptor pathway). Activation of the extrinsic pathway by FAS-ligand did not induce apoptosis in primary AML, instead the proapoptotic signal was shown to 'fade', even in the early phase of the apoptotic sequence. However, activation of the intrinsic pathway induced severe cytotoxicity in all samples that showed the characteristic features of typical apoptosis, with a prominent apoptotic volume decrease (blebbing) in the early phase, significant increases in caspase 3 activity (intermediate or effector phase) and breakdown of cellular energy production in the late phase of apoptosis. These characteristics did not differ between prognostically favourable versus unfavourable AML karyotypes or between clinically responding versus refractory AML--indicating that a functional apoptotic apparatus is present even in the unfavourable AML subgroups. Our data indicate that the mechanism for a broad clinical resistance is not a dysfunctional apparatus per se but rather the consequence of anti-apoptotic regulation impeding otherwise functional apoptotic machinery.