Chronic myeloid leukemia progenitor cells require autophagy when leaving hypoxia-induced quiescence.

Angela Ianniciello,Philippe Brunet De La Grange,Zoran Ivanovic,Jean Chevaleyre,Persio Dello Sbarba,Arnaud Villacreces,Vanessa Desplat,Jean-Max Pasquet,Yan Peytour,François-Xavier Mahon,Isabelle Vigon,Amélie Guitart,Pierre-Yves Dumas,Muriel Priault,Claire Drullion

doi:10.18632/oncotarget.18904

Abstract

Albeit tyrosine kinase inhibitors anti-Abl used in Chronic Myeloid Leukemia (CML) block the deregulated activity of the Bcr-Abl tyrosine kinase and induce remission in 90% of patients, they do not eradicate immature hematopoietic compartments of leukemic stem cells. To elucidate if autophagy is important for stem cell survival and/or proliferation, we used culture in low oxygen concentration (0.1% O2 for 7 days) followed back by non-restricted O2 supply (normoxic culture) to mimic stem cell proliferation and commitment. Knockdown of Atg7 expression, a key player in autophagy, in K562 cell line inhibited autophagy compared to control cells. Upon 7 days at 0.1% O2 both K562 and K562 shATG7 cells stopped to proliferate and a similar amount of viable cells remained. Back to non-restricted O2 supply K562 cells proliferate whereas K562 shATG7 cells exhibited strong apoptosis. Using immunomagnetic sorted normal and CML CD34+ cells, we inhibited the autophagic process by lentiviral infection expressing shATG7 or using a Vps34 inhibitor. Both, normal and CML CD34+ cells either competent or deficient for autophagy stopped to proliferate in hypoxia. Surprisingly, while normal CD34+ cells proliferate back to non restricted O2 supply, the CML CD34+ cells deficient for autophagy failed to proliferate. All together, these results suggest that autophagy is required for CML CD34+ commitment while it is dispensable for normal CD34 cells.

Highlights

Chronic myeloid leukemia (CML) is a clonal malignant hematopoietic disorder characterized by the presence of a t(9;22)(q34;q11) reciprocal translocation [1, 2]
Inhibition of ATG7 expression was confirmed by western blotting and the consequent inhibition of autophagy in K562 shATG7 was verified by detecting the conversion of microtubule-associated light chain 3B-I in LC3B-II by western-blotting (Figure 1B)
According to the procedure described by Giuntoli et al [18] (Figure 1A), the two cell lines were cultured at 0.1% O2 for 7 days (LC1)

Summary

Introduction

Chronic myeloid leukemia (CML) is a clonal malignant hematopoietic disorder characterized by the presence of a t(9;22)(q34;q11) reciprocal translocation [1, 2]. TKI imatinib is the front-line therapy of CML in chronic phase, and competes with ATP for binding to the Abl kinase domain. Several studies reported that targeting other pathways in combination with TKI treatment could be efficient enough to target LSC. Among these interesting pathways, the inhibition of autophagy has been reported to be deleterious on leukemia and on the LSC reservoir [15]. The inhibition of autophagy has been reported to be deleterious on leukemia and on the LSC reservoir [15] This interesting result has led to propose preclinical and clinical trials in CML using autophagy inhibitors. It is triggered by stress conditions like nutriment starvation and used a complex machinery involving ATG proteins [16, 17]

Methods

Results

Conclusion