Background: Macroautophagy (autophagy) has been shown to be both a pro-survival and pro-death signalling pathway. The decision between life or death is dependent on the type and developmental stage of the cell and the duration and intensity of the signal. Autophagy is used by cancer cells as a mechanism to overcome cellular stresses such as starvation, reactive oxygen species (ROS) generation and misfolded protein accumulation and can result in tumorigenesis and resistance to chemotherapy. One of the pathways which regulate responses to ROS/ER stress is the unfolded protein response (UPR). Briefly, upon ER stress, the HSP70 family member BiP is sequestered away from IRE1 , PERK and ATF6, resulting in IRE1 and PERK phosphorylation and activation of downstream signalling pathways. These signals can result in cell survival by upregulating AKT or cell death via induction of CHOP and noxa. However this pathway has not been fully investigated in chronic lymphocytic leukaemia (CLL). Interestingly the UPR is also known to regulate autophagy. The lipidation of LC3 from LC3-I to LC3-II is a surrogate marker of autophagy, since LC3-II is essential for formation of the autophagasome and the initiation of autophagy. Beclin-1 is another essential protein involved in canonical autophagy and is sequestered by Bcl-2 preventing autophagy in a number of cell types. Its release from Bcl-2 enables Beclin-1 to interact with other autophagy specific proteins and initiates autophagasome formation. Methods: Twenty CLL samples were treated with the HSP70 inhibitor 2-phenylacetylenesulphonamide (PAS; 10-20 M), an activator of the UPR, in the presence of the pan-caspase inhibitor ZVAD. In addition 4 control samples (normal B cells) were treated with PAS for the indicated times. Immunoblotting was performed for BiP, HSP70, Phosphorylated eIF2 (eIF2 -P), JNK (JNK-P), Jun (Jun-P), p38MAPK, LC3-I/II, Beclin expression and its co-immunoprecipitation with Bcl-2. Ten CLL cell isolates were treated with 2 autophagy inhibitors bafilomycin (Baf) (50-200 nM) and 3-methyl adenine (3-MA) (2.5-10 mM) and their effects on cell killing evaluated using PARP cleavage, a surrogate marker of caspase 3 activation. Results: CLL cells expressed a greater basal level of LC3-II in a proportion of samples relative to normal B cells. In addition Beclin-1 did not coimmunoprecipitate with Bcl-2 above that shown with the isotype control in CLL cells. Treatment with PAS resulted in an increase in stress which resulted in the upregulation of BiP and HSP70 protein. This subsequently led to phosphorylation of eIF2 indicative of PERK activation, coincidental with an increase in the eIF2 targets LC3-I/II and noxa expression. Treatment with PAS also resulted in an increase in JNK, Jun and p38MAPK phosphorylation, 3 proteins involved in regulating autophagy in mammalian cells. Furthermore, CLL cells treated with the autophagy inhibitors 3-MA and Baf rapidly underwent apoptosis within 24 h. Conclusion: These data suggest that autophagy is active at a basal level in a proportion of CLL cells and is greater than that seen in control B cells. Activation of the UPR with PAS led to an upregulation of proteins known to regulate autophagy and resulted in an increase in LC3-II, a protein routinely used as an indicator of autophagy. Inhibitors of autophagy also rapidly induced apoptosis in CLL cells. These data suggested that autophagy may have a pro-survival role in the biology of CLL. However pro-survival autophagy is balanced on a knife-edge, such that perturbation by inducing or inhibiting the pathway results in CLL cell death.