Exploiting Drug Metabolism to Activate Immunity Against Cancer

Dr Rock J. Mancini EXPLOITING DRUG METABOLISM TO ACTIVATE IMMUNITY AGAINST CANCER

Multidrug resistance is one of the main culprits underlying the failure of chemotherapy as a cancer treatment. Whilst many therapies are initially effective, a considerable proportion of patients eventually incur a poor prognosis and recurrence of malignant spread due to developing drug resistance at a later stage. Dr Rock J. Mancini, from Washington State University, has devised an approach that exploits proteins over-expressed in drug- resistant cancers to convert inactive prodrug substrates into active drugs that initiate an immune response targeted at cancer cells.

Fighting Multidrug Resistance in To address this, Dr Rock J. Mancini, Cancer Assistant Professor of Organic Chemistry at Washington State University, is In recent years, there have been pioneering methods that exploit drug significant advances in the treatment efflux to link the immune system, at of several types of cancer with a chemical level, to drug-resistant chemotherapy and immunotherapy cancers. Dr Mancini is working to create alike. Many cancers are either resistant an array of smart immunostimulant to specific chemotherapeutic drugs or drugs that are finely modulated by acquire drug resistance at a later stage, stimuli including enzyme activity, despite initially responding favourably temperature, and disease-specific to therapy. However, immunotherapy biomarkers that are present in or on has also emerged as a new class of cancer cells. cancer treatment with efficacy that is fundamentally orthogonal to acquired Dr Mancini’s team of researchers have chemotherapeutic drug resistance. devised an ingenious strategy that Molecular structure of a smart exploits drug efflux to raise an immune immunostimulant developed by the There are several mechanisms through response against cancer cells, an Mancini Laboratory. which cancer cells develop drug approach that could provide clinical resistance. One commonality across advantage to patients with multidrug- many drug-resistant cancers is that they resistant cancers. The approach uses cells to escape their fate. The hope is use molecular transporters that derive smart immunostimulants that are that, by selectively activating immune energy from intracellular adenosine metabolised by cancer cells, which then cells in close proximity to cancer cells, triphosphate to actively pump cancer secrete immunostimulant drugs via drug this technology will enable a new way drugs out of the cells; a series of efflux, effectively alerting the immune in which drug-resistant cancers can be biochemical reactions resulting in the system to the cancer. By coupling cancer eradicated, and tumour growth and mechanisms of drug resistance known cell metabolism and drug efflux to spread can be halted. collectively as ‘drug efflux’. activate the immune system, the team hopes to affect the ability of cancer

WWW.SCIENTIA.GLOBAL A Trojan Horse to Trigger Immunity prodrug into an active anti-tumour The advantage of engaging immune Against Cancer Cells chemotherapeutic. With this approach, cells to target tumours is that, once there is the potential for diffusion of they are trained to recognise cancer Over the past decades, medicinal the active drug away from the target cells, they have the ability to destroy chemists have contributed to the cell, where it can interact with other them with high specificity, leaving rational design and production nearby cells via a phenomenon known healthy cells nearby intact. Given that of synthetic immunostimulants, as the ‘bystander effect’. The bystander enzyme-directed immunostimulants compounds that interact with specific effect can be beneficial, resulting in the are generated within the tumour receptors on immune cells, triggering a permanent damage of nearby cancer microenvironment, they are perfectly proinflammatory response. However, cells, or unwanted, resulting in overall placed to target tumour cells with high immunostimulants are susceptible to toxicity from the damage of nearby non- efficacy and low toxicity. diffusion, off-target activity, and other cancerous cells. drug resistance mechanisms, including Cancer Takes the Bait the drug efflux pathway described Importantly, drug efflux enhances above. the bystander effect. Dr Mancini and In two different papers, published in his team created a way to harness 2016 and 2018, Dr Mancini and his Dr Mancini’s group intentionally uses the bystander effect for potential team reported the creation of the first immunostimulants as drug efflux therapeutic advantage by replacing enzyme-directed proimmunostimulants substrates to hijack the drug resistance the chemotherapeutics typically used in the form of imidazoquinoline- mechanism to produce molecules that in DEPT with immunostimulants. This pyranosides, substrates for hydrolase activate immune cells. Specifically, modification makes the bystander effect enzymes, such as the alpha- his research group is recognised as a desirable outcome, causing multidrug- mannosidases expressed by cancer the first to develop enzyme-directed resistant cancer cells to activate cell metabolism. Effectively acting like immunostimulants (immunostimulants bystander immune cells. a piece of cheese in a mousetrap, the activated by enzymes present in cancer BAIT prodrug is harmless and inert cell metabolism) and demonstrate Dr Mancini and his team named until it is eaten (metabolised) by cancer that they undergo drug efflux from their approach ‘bystander-assisted cells, ultimately initiating an anti-cancer multidrug-resistant cancers. immunotherapy’ (BAIT). In BAIT, an inert immune response. prodrug is first metabolised by enzymes Cocking the Molecular Guns within multidrug-resistant cancer cells In the system developed by Dr Mancini’s to produce an active immunostimulant laboratory, alpha-mannosidase One approach to addressing drug metabolite. Following this conversion, converts the prodrug substrate into the resistance is directed enzyme prodrug the immunostimulant is transported to active immunostimulant imiquimod. therapy (DEPT), which attempts to the extracellular space, via drug efflux. The studies were conducted in vitro outcompete drug efflux by increasing Once in the extracellular space, the on cultured prostate cancer cells, and the local concentration of therapeutic immunostimulant activates bystander the enzyme-mediated production of drugs in the area surrounding tumour immune cells, resulting in an immune imiquimod resulted in the activation cells. In DEPT, enzymes within a solid response initiated at the site of the of macrophages and dendritic cells, tumour convert an enzyme-directed cancer. providing robust evidence that the

WWW.SCIENTIA.GLOBAL studies demonstrate that enzyme- glycosidase activity and drug efflux B)

κ directed proimmunostimulants are across different cancer cell lines will - well-tolerated and do not cause provide insight into which factors Melanoma systemic inflammatory toxicity in affect the cancer-mediated activation

Prostate Cancer otherwise healthy mice. The team of immune cells, serving as a powerful is currently testing the efficacy on a diagnostic tool to predict the efficacy of Breast Cancer model of drug-resistant breast cancer BAIT in specific cancers. in mice, providing evidence that immunostimulants produced by the Optimizing BAIT Drug Design and enzyme-directed BAIT method are more Other Future Developments effective when compared to directly administered parent immunostimulant. The team is currently optimising a lead enzyme-directed immunostimulant Immune Response (Macrophage NF (Macrophage Response Immune BAIT Drug BAIT Drug Comparing the Immunostimulant for further studies in vivo. Dr Mancini Alone (+) Cancer Cells Effect Across Different Cancer Cell and his group also hope to confirm Lines the observation that cancer cell drug Data taken from Ryan et al., Comparing efflux, a trait linked to drug resistance, the immunogenicity of glycosidase- In optimizing the BAIT approach, is the most important factor that directed resiquimod prodrugs mediated by cancer cell metabolism, Acta Mancini and his group compared the drives efficacy of the prodrugs. To test Pharmacologica Sinica, 2020, doi: immunogenicity of several enzyme- this hypothesis, his lab has spent the 10.1038/s41401-020-0432-4 directed prodrugs that all have the past months growing cells that are imidazoquinoline immunostimulant resistant to chemotherapy, a process resiquimod embedded in their structure. that involves adding chemotherapeutic Like imiquimod, resiquimod is a ligand drugs to the cell culture and selecting for immunogenic toll-like receptors. This the resistant cancer cells that survive. compound was chosen for its potency Overall, the findings suggest that BAIT at nanomolar concentration, rather than is best suited for cancer cells that simplicity, as was the case in earlier overexpress proteins associated with iterations. The cancer types studied drug efflux, in particular multidrug- were skin cancer, prostate cancer, and resistant cancers that do not respond to breast cancer, because they are among chemotherapy. However, to understand the most frequently diagnosed cancers the process at a molecular level, more in the USA. studies will be needed to characterise all of the possible routes of efflux, as the The group published their latest immunostimulant might be a substrate mechanisms of drug efflux in cancer results in 2020 which confirm that for many different transport proteins cells can be exploited via BAIT. Like most immunogenicity across different involved in drug efflux. cancer immunotherapies, BAIT offers cancer cell lines is due to the enzymatic the advantage of specifically targeting conversion of substrate prodrug into Given that the efficacy of traditional one undesired type of cell, unlike other the immunostimulant resiquimod, chemotherapeutics is attenuated by chemotherapeutic agents that cause liberated following drug efflux. The mechanisms of drug resistance, Dr high levels of toxicity due to their lack of team compared cancer cell metabolism Mancini’s research efforts will continue specificity. of the prodrug to adding the enzymes to provide tools to interrogate the responsible for metabolism separately, immune system, shedding light on Dr Mancini and his team used cell both of which resulted in increased how immune cells can be activated cultures to demonstrate that enzyme- production of active immunostimulant and initiate a targeted response directed immunostimulants only target and the activation of immune cells. It against cancer. In the long term, the those cultures that overexpressed the was established that melanoma cells team has the ambition of contributing corresponding proteins associated in vitro showed the greatest extent of significantly to the field of medicinal with multidrug-resistant cancer cells. immunogenicity, followed by prostate chemistry by designing the next This indicates that this method triggers cancer cells and breast cancer cells. generation of immunotherapeutics and a cancer-specific immune response cancer vaccines. without causing inflammatory toxicity, Not surprisingly, the group established and could be used alongside blockers that different cell lines displayed of immunosuppression to improve different levels of enzyme expression efficacy. Preliminary in vivo mouse and activity. Those differences in

WWW.SCIENTIA.GLOBAL Meet the researcher Dr Rock J. Mancini Department of Chemistry Washington State University Pullman, WA USA

Dr Rock J. Mancini is Assistant Professor of Organic Chemistry KEY COLLABORATORS at Washington State University. Dr Mancini obtained his PhD in Organic Chemistry from the University of California, Los Dr Amy E. Nielsen, Clinical Assistant Professor of Chemistry, Angeles, in 2012. He subsequently trained as a synthetic Washington State University immunologist from 2012 to 2015 while pursuing his Dr Maryam Davaritouchaee (Postdoctoral Scholar, Mancini postdoctoral studies at the University of California, Irvine. Since Group) starting his tenure track position in 2015, his research interests Austin Ryan and Anthony Burt (Graduate Student Researchers, have evolved to incorporate organic chemistry, chemical Mancini Group) biology, and polymer chemistry. The research focus of his laboratory is the development of methods to both control and FUNDING interrogate the immune system, aimed at creating the next generation of immunotherapeutics and vaccines. Dr Mancini The research reported in this publication was supported by the mentors several junior researchers in his team and encourages National Cancer Institute of the National Institutes of Health them to reach their full potential as scientists by acquiring a under Award Number R01CA234115. The content is solely deep understanding of basic and applied research processes, the responsibility of the researchers and does not necessarily from hypothesis generation to its proof of concept and ultimate represent the official views of the National Institutes of Health. dissemination to the scientific community. FURTHER READING CONTACT AT Ryan, AJ Pulukuri, M Davaritouchaee, et al, Comparing the E: [email protected] Cancer Cell Metabolism and Immunogenicity of Glycosidase- W: https://chem.wsu.edu/faculty/mancini-rock/ Directed Resiquimod Prodrugs Mediated by Cancer Cell @ManciniLab_WSU Metabolism, Acta Pharmacologica Sinica, 2020, 41, 9915–1004.

AJ Burt, JD Hantho, AE Nielsen, RJ Mancini, An Enzyme- Directed Imidazoquinoline Activated by Drug Resistance, Biochemistry, 2018, 57, 2184–2188.

JD Hantho, TA Strayer, AE Nielsen, RJ Mancini, An Enzyme- Directed Imidazoquinoline for Cancer Immunotherapy, ChemMedChem, 2016, 11, 2496–2500.