Heat shock protein-based therapy as a potential candidate for treating the sphingolipidoses.

Lysosomal storage diseases (LSDs) often manifest with severe systemic and central nervous system (CNS) symptoms. The existing treatment options are limited and have no or only modest efficacy against neurological manifestations of disease. We demonstrate that recombinant human heat shock protein 70 (HSP70) improves the binding of several sphingolipid-degrading enzymes to their essential cofactor bis(monoacyl)glycerophosphate in vitro. HSP70 treatment reversed lysosomal pathology in primary fibroblasts from 14 patients with eight different LSDs. HSP70 penetrated effectively into murine tissues including the CNS and inhibited glycosphingolipid accumulation in murine models of Fabry disease (Gla(-/-)), Sandhoff disease (Hexb(-/-)), and Niemann-Pick disease type C (Npc1(-/-)) and attenuated a wide spectrum of disease-associated neurological symptoms in Hexb(-/-) and Npc1(-/-) mice. Oral administration of arimoclomol, a small-molecule coinducer of HSPs that is currently in clinical trials for Niemann-Pick disease type C (NPC), recapitulated the effects of recombinant human HSP70, suggesting that heat shock protein-based therapies merit clinical evaluation for treating LSDs.

Connecting Hsp70, sphingolipid metabolism and lysosomal stability.

Heat shock protein 70 (Hsp70) is an evolutionary highly conserved molecular chaperone. Upon cancer-associated translocation to the lysosomal compartment, it promotes cell survival by inhibiting lysosomal membrane permeabilization, a hallmark of stress-induced death. We have recently shown that Hsp70 stabilizes lysosomes by binding to the endo-lysosomal lipid bis(monoacylglycero)phosphate (BMP), an essential co-factor for lysosomal sphingolipid catabolism. The Hsp70-BMP interaction enhances the activity of acid sphingomyelinase, an important enzyme that hydrolyzes sphingomyelin. Importantly, treatment with recombinant Hsp70 effectively reverts the dramatic increase in lysosomal volume and decrease in lysosomal stability in cells from patients with Niemann-Pick disease, a genetic disorder associated with reduced acid sphingomyelinase activity. These findings give new insight into the mechanisms controlling lysosomal stability and integrity, and open new exciting possibilities for the treatment of cancer as well as Niemann-Pick disease.

Hsp70 stabilizes lysosomes and reverts Niemann-Pick disease-associated lysosomal pathology.

Heat shock protein 70 (Hsp70) is an evolutionarily highly conserved molecular chaperone that promotes the survival of stressed cells by inhibiting lysosomal membrane permeabilization, a hallmark of stress-induced cell death. Clues to its molecular mechanism of action may lay in the recently reported stress- and cancer-associated translocation of a small portion of Hsp70 to the lysosomal compartment. Here we show that Hsp70 stabilizes lysosomes by binding to an endolysosomal anionic phospholipid bis(monoacylglycero)phosphate (BMP), an essential co-factor for lysosomal sphingomyelin metabolism. In acidic environments Hsp70 binds with high affinity and specificity to BMP, thereby facilitating the BMP binding and activity of acid sphingomyelinase (ASM). The inhibition of the Hsp70-BMP interaction by BMP antibodies or a point mutation in Hsp70 (Trp90Phe), as well as the pharmacological and genetic inhibition of ASM, effectively revert the Hsp70-mediated stabilization of lysosomes. Notably, the reduced ASM activity in cells from patients with Niemann-Pick disease (NPD) A and B-severe lysosomal storage disorders caused by mutations in the sphingomyelin phosphodiesterase 1 gene (SMPD1) encoding for ASM-is also associated with a marked decrease in lysosomal stability, and this phenotype can be effectively corrected by treatment with recombinant Hsp70. Taken together, these data open exciting possibilities for the development of new treatments for lysosomal storage disorders and cancer with compounds that enter the lysosomal lumen by the endocytic delivery pathway.

Anti-cancer agent siramesine is a lysosomotropic detergent that induces cytoprotective autophagosome accumulation.

A sigma-2 receptor ligand siramesine induces lysosomal leakage and cathepsin-dependent death of cancer cells in vitro and displays potent anti-cancer activity in vivo. The mechanism by which siramesine destabilizes lysosomes is, however, unknown. Here, we show that siramesine induces a rapid rise in the lysosomal pH that is followed by lysosomal leakage and dysfunction. The rapid accumulation of siramesine into cancer cell lysosomes, its ability to destabilize isolated lysosomes, and its chemical structure as an amphiphilic amine indicate that it is a lysosomotropic detergent. Notably, siramesine triggers also a substantial Atg6- and Atg7-dependent accumulation of autophagosomes that is associated with a rapid and sustained inhibition of mammalian target of rapamycin complex 1 (mTORC1; an inhibitor of autophagy). Siramesine fails, however, to increase the degradation rate of long-lived proteins. Thus, the massive accumulation of autophagosomes is likely to be due to a combined effect of activation of autophagy signaling and decreased autophagosome turnover. Importantly, pharmacological and RNA interference-based inhibition of autophagosome formation further sensitizes cancer cells to siramesine-induced cytotoxicity. These data identify siramesine as a lysosomotropic detergent that triggers cell death via a direct destabilization of lysosomes and cytoprotection by inducing the accumulation of autophagosomes. Threrefore, the combination of siramesine with inhibitors of autophagosome formation appears as a promising approach for future cancer therapy.