An Enzyme‐Directed Imidazoquinoline for Cancer Immunotherapy

Hantho, Joseph D.; Strayer, Timothy A.; Nielsen, Amy E.; Mancini, Rock J.

doi:10.1002/cmdc.201600443

Cited by 20 publications

(19 citation statements)

References 35 publications

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“…Intermediates ( 8 ) and ( 11 ) were synthesized according to literature procedure [ 6 , 7 ]. EDIs ( 7 ), ( 10 ), and ( 13 ) were synthesized as detailed in Fig.…”

Section: Methodsmentioning

confidence: 99%

“…1a ). To this end, we developed an enzyme-directed immunostimulant (EDI) prodrug motif, which is metabolized by cancer cells, resulting in immunogenicity that is enhanced by drug efflux and the bystander effect [ 6 , 7 ]. In first-generation EDIs, the immunostimulant imiquimod [ 8 ] was chosen for its synthetic simplicity rather than potency, and the enzyme-directing groups were specifically matched to cancer cell model systems that overexpressed complementary enzyme and transport proteins required for BAIT.…”

Section: Introductionmentioning

confidence: 99%

“…Among these glycosidases, β-glu is unique because it is localized intracellularly in healthy cells but found extracellularly in tumor and necrotic tissues, although it remains unclear whether extracellular β-glu is derived from cancer cells themselves or introduced through other sources such as tumor-infiltrating lymphocytes [ 3 , 25 , 26 ]. Each glycosidase has been employed as an enzyme target, either in DEPT [ 27 , 28 ] or BAIT [ 6 , 7 ], but there have been few direct comparisons of different enzyme-directing groups in a single enzyme-directed prodrug system [ 29 ] and, with the exception of the present study, none that compare EDIs. As such, we were interested in comparing EDIs targeted to different glycosidases endogenously expressed across several cancer cell lines.…”

Section: Introductionmentioning

confidence: 99%

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Comparing the immunogenicity of glycosidase-directed resiquimod prodrugs mediated by cancer cell metabolism

et al. 2020

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We have recently developed an enzyme-directed immunostimulant (EDI) prodrug motif, which is metabolized to active immunostimulant by cancer cells and, following drug efflux, activates nearby immune cells, resulting in immunogenicity. In this study, we synthesized several EDI prodrugs featuring an imidazoquinoline immunostimulant resiquimod (a Toll-like receptor 7/8 agonist) covalently modified with glycosidase enzyme-directing groups selected from substrates of β-glucuronidase, α-mannosidase, or β-galactosidase. We compared the glycosidase-dependent immunogenicity elicited by each EDI in RAW-Blue macrophages following conversion to active immunostimulant by complementary glycosidase. At a cellular level, we examined EDI metabolism across three cancer cell lines (B16 melanoma, TC2 prostate, and 4T1 breast cancer). Comparing the relative immunogenicity elicited by each EDI/cancer cell combination, we found that B16 cells produced the highest EDI prodrug immunogenicity, achieving >95% of that elicited by unmodified resiquimod, followed by TC2 and 4T1 cells (40% and 30%, respectively). Immunogenicity elicited was comparable for a given cell type and independent of the glycosidase substrate in the EDIs or differences in functional glycosidase activity between cell lines. Measuring drug efflux of the immunostimulant payload and efflux protein expression revealed that EDI/cancer cell-mediated immunogenicity was governed by efflux potential of the cancer cells. We determined that, following EDI conversion, immunostimulant efflux occurred through both P-glycoprotein-dependent and P-glycoprotein-independent transport mechanisms. Overall, this study highlights the broad ability of EDIs to couple immunogenicity to the metabolism of many cancers that exhibit drug efflux and suggests that designing future generations of EDIs with immunostimulant payloads that are optimized for drug efflux could be particularly beneficial.

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“…Intermediates ( 8 ) and ( 11 ) were synthesized according to literature procedure [ 6 , 7 ]. EDIs ( 7 ), ( 10 ), and ( 13 ) were synthesized as detailed in Fig.…”

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Comparing the immunogenicity of glycosidase-directed resiquimod prodrugs mediated by cancer cell metabolism

et al. 2020

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“…The imidazoquinoline immunostimulant Imiquimod (IMQ) [21–24] was chosen as payload for our proof‐of‐concept LDNPC reagent because our previous work with enzyme‐directed IMQ allowed us to estimate its behavior as a prodrug [25,26] . Based on this, and the well‐defined structure activity relationship of the imidazoquinoline drug class, [27,28] we hypothesized that linkage at the aminoquinoline nitrogen would lead to abrogated immunostimulatory activity and therefore allow the LDNPC reagent, as well as the covalently labeled avidin complex to serve as a prodrug of IMQ.…”

Section: Figurementioning

confidence: 99%

A Ligand‐Directed Nitrophenol Carbonate for Transient in situ Bioconjugation and Drug Delivery

et al. 2020

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Here we report the first use of ligand‐directed proximity accelerated bioconjugation chemistry in the tandem delivery and release of a therapeutic payload. To do this, we designed a nitrophenol carbonate for ligand‐directed in situ bioconjugation of a prodrug payload to a protein. The transient nature of our conjugation chemistry renders the protein a depot for time‐dependent release of active drug following hydrolysis and self‐immolation. In our model system, using an immunostimulant prodrug, biotin ligand, and avidin protein, we observe release of bioavailable immunostimulant both spectroscopically and with an immune cell line over 48 h. Avidin co‐crystalized with the nitrophenolate directing group verified the binding pose of the ligand and offered insight into the mechanism of in situ bioconjugation. Overall, this scaffold warrants further investigation for the time‐dependent delivery of therapeutics and use in protein ligand pairs beyond biotin and avidin used for this work.

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“…The imidazoquinoline immunostimulant Imiquimod (IMQ) [21][22][23][24] was chosen as payload for our proof-ofconcept LDNPC reagent because our previous work with enzyme-directed imidazoquinolines allowed us to estimate imidazoquinoline behavior as prodrugs. [25,26] Due to the well-defined structure activity relationship of the imidazoquinoline drug class, [27,28] we hypothesized that linkage at the C4 aniline would lead to abrogated immunostimulatory activity and therefore allow the LDNPC reagent, as well as the covalently labeled avidin complex to serve as a prodrug of IMQ. Furthermore, the IMQ payload is an active immunostimulant for the RAW-Blue murine macrophage cell line (RB Cells) which links activation of the transcription factor NF-κB to secreted embryonic alkaline phosphatase which can be quantified with colorimetric phosphate substrates.…”

mentioning

confidence: 99%

A Ligand-Directed Nitrophenol Carbonate for Transient In Situ Bioconjugation and Drug Delivery

Burt¹,

Ahmadvand²,

Opp³

et al. 2020

Preprint

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Here we report the first use of ligand-directed proximity accelerated bioconjugation chemistry in the tandem delivery and release of a therapeutic payload. To do this we designed a nitrophenol carbonate for ligand-directed in situ bioconjugation of a prodrug payload to a protein. The transient nature of our conjugation chemistry renders the protein a depot for time-dependent release of active drug following hydrolysis and self-immolation. In our model system, using an immunostimulant prodrug, biotin ligand, and avidin protein, we observe time-dependent release of bioavailable immunostimulant both spectroscopically and with an immune cell line over 48 h. Avidin co-crystalized with the biotin directing group verified the binding pose of the ligand and offered insight into the mechanism of in situ bioconjugation. Overall, this scaffold warrants further investigation for the time-dependent delivery of therapeutics and use in protein ligand pairs beyond biotin and avidin used for this work.

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An Enzyme‐Directed Imidazoquinoline for Cancer Immunotherapy

Cited by 20 publications

References 35 publications

Comparing the immunogenicity of glycosidase-directed resiquimod prodrugs mediated by cancer cell metabolism

Comparing the immunogenicity of glycosidase-directed resiquimod prodrugs mediated by cancer cell metabolism

A Ligand‐Directed Nitrophenol Carbonate for Transient in situ Bioconjugation and Drug Delivery

A Ligand-Directed Nitrophenol Carbonate for Transient In Situ Bioconjugation and Drug Delivery

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