Summary: Lysosomes are the recycling centers of the cell where organelles and proteins are degraded during autophagy and macropinocytosis; they also serve as signaling hubs that control the activity of mTORC1. In this issue, Rebecca and colleagues report the development of a new type of lysosomal inhibitor for cancer therapy that can inhibit multiple lysosomal activities that are needed for tumor cell survival and growth. Cancer Discov; 7(11); 1218–20. ©2017 AACR.
See related article by Rebecca et al., p. 1266.
Lysosomes are important for both catabolic pathways such as autophagy and macropinocytosis and anabolic growth pathways driven by mTORC1. These pathways are all potential targets in cancer. Autophagy is the process by which cellular material is delivered to the lysosome for degradation, and it has multiple roles in cancer (1). Although autophagy can have both protumor and antitumor effects, current efforts to deliberately target autophagy in cancer treatment focus primarily on trying to inhibit autophagy due to its ability to promote growth of established tumors and mediate chemoresistance during cancer therapy (2). Macropinocytosis is a specialized type of endocytosis that functions to deliver extracellular proteins to lysosomes; this promotes tumor growth, especially in RAS-driven tumors (3). mTOR is a critical regulator of cell growth and metabolism (4), and mTOR inhibitors have been widely tested as anticancer agents in preclinical models and patients (5). Antimalarial drugs such as chloroquine, hydroxychloroquine, or quinacrine inhibit lysosomal function. However, these agents also have other effects and lower potency than we would like. Although chloroquine and hydroxychloroquine are currently the only drugs that are used clinically as autophagy inhibitors and are being tested in dozens of trials (2), they do not affect other lysosomal activities, such as mTORC1 regulation, and they also have autophagy-independent effects as anticancer agents (6). For example, it has been known for many years that chloroquine can bind to DNA (7). A new article by Rebecca and colleagues (8) takes a systematic approach to lysosome inhibition and, for the first time, describes an agent that can inhibit multiple cancer-driving functions of the lysosome.
In this new article, Rebecca and colleagues (8) extend a previous study from the same laboratories (9) that reported that dimerization of chloroquine could increase its potency as an autophagy inhibitor by more systematically testing dimerized antimalarials. Dimerization of the oldest antimalarial drug, quinacrine, was particularly potent at suppressing tumor growth, and the authors went on to optimize the linker sequence. A systematic medicinal chemistry effort allowed them to achieve three important advances. First, they were able to make an agent that, unlike the parent antimalarials, affects only the lysosome. Importantly, they also have versions that could not accumulate in lysosomes or inhibit lysosome function, and consequently have different biological effects as autophagy inducers versus inhibitors. Because these agents affect either lysosomes or DNA but not both, this allows more rigorous testing of the mechanism of action than is possible with previous lysosomal inhibitors like chloroquine. The second achievement focused on a lysosome-specific agent called DQ661. The high potency and lysosomal specificity of the drug allowed DQ661 to be used as a photo-affinity labeling probe to identify its molecular target as palmitoyl-protein thioesterase 1 (PPT1). PPT1 is required for depalmitoylation of proteins, and DQ661 has its effects on tumor cells by binding to and inhibiting this enzyme. A third advance showed that this drug has a very interesting multitarget activity profile. It can block lysosomal activity and the major catabolic functions of autophagy and macropinocytosis, but can also inhibit mTORC1; this is the first example of a molecule that can block both the signaling and degradative functions of lysosomes. Importantly, mTOR inhibition mediated by DQ661 involves a novel mechanism. DQ661 disrupts the complex that regulates mTORC1 at the lysosomal membrane. This ejects mTORC1 from the lysosome and prevents the amino acid–dependent regulation of mTORC1 kinase activity (Fig. 1). Potential benefits of this different approach to mTOR inhibition are increased potency compared with catalytic inhibitors, broader activity against tumor cells with different genotypes and resistance mechanisms, increased functionality in nutrient-rich conditions, and the ability to cooperate with catalytic inhibitors of mTORC1.
The net effect of all these advances was that DQ661 was shown to have in vivo antitumor activity against different tumor models in both immunocompetent animals and immunodeficient models. Importantly, the closely related drug DQ660, which does not target the lysosome but does cause tumor DNA damage, had little effect on tumor growth. This shows that the lysosome inhibition is critical for the antitumor effects of these molecules. Interestingly, in vivo activity was demonstrated in a tumor that is resistant to hydroxychloroquine and is not especially sensitive to genetic inhibition of either autophagy or mTOR alone. This is interesting because it has been argued that the key to success in targeting autophagy in cancer will be to focus on those tumors that are themselves especially autophagy dependent as defined by their high sensitivity to genetic interference with autophagy (2). However, the field currently lacks good biomarkers or oncogenic profiles to identify autophagy-dependent tumors, and even tumor cells with driver mutations such as mutant RAS, which has been associated with increased autophagy and sensitivity to autophagy inhibition (10), do not always respond to autophagy inhibition (11, 12). This raises the exciting notion that perhaps autophagy dependence may not be a necessary requirement for success when targeting autophagy using more potent multitargeting drugs like DQ661 and may broaden the applicability of autophagy inhibition as a therapeutic strategy.
As is often the case with important studies, the new findings also raise new questions. The authors' data suggest that inhibition of PPT1 affects the proper localization of the lysosomal v-ATPase, which generates and maintains the pH gradient that keeps lysosomes acidic. But the effects observed with DQ661 must be more complex, as other drugs that block acidification are not able to block mTORC1 in the presence of amino acids. Thus, DQ661 may be a useful tool to further understand exactly how the mTORC1 complex is regulated at the lysosome membrane. Why does blocking protein depalmitoylation in the lysosome inhibit autophagy? Is this due to the effect on lysosomal pH, or are there other specific palmitoylated target proteins that are needed for autophagy? Will tumor cells be able to develop resistance to DQ661-mediated inhibition of mTOR as easily as they circumvent the existing inhibitors? Why does interference with these processes kill tumor cells? And, will there be differences between tumors based on their susceptibility to inhibition of lysosome function with these agents? Finally, the identification of PPT1 as a new therapeutic target in cancer suggests that protein palmitoylation deserves further investigation from the perspective of understanding both cancer biology and cancer treatment. This article opens the way to addressing these and other questions and to further testing of the efficacy of lysosome inhibition for cancer treatment.
Disclosure of Potential Conflicts of Interest
No potential conflicts of interest were disclosed.
The authors are supported by NIH grants RO1 CA150925, RO1 CA190170, R21 CA197887, and T32 CA190216.