title: Rational design of an AKR1C3-resistant analog of PR-104 for enzyme-prodrug therapy.
abstract: The clinical stage anti-cancer agent PR-104 has potential utility as a cytotoxic prodrug for exogenous bacterial nitroreductases expressed from replicating vector platforms. However substrate selectivity is compromised due to metabolism by the human one- and two-electron oxidoreductases cytochrome P450 oxidoreductase (POR) and aldo-keto reductase 1C3 (AKR1C3). Using rational drug design we developed a novel mono-nitro analog of PR-104A that is essentially free of this off-target activity in vitro and in vivo. Unlike PR-104A, there was no biologically relevant cytotoxicity in cells engineered to express AKR1C3 or POR, under aerobic or anoxic conditions, respectively. We screened this inert prodrug analog, SN34507, against a type I bacterial nitroreductase library and identified E. coli NfsA as an efficient bioactivator using a DNA damage response assay and recombinant enzyme kinetics. Expression of E. coli NfsA in human colorectal cancer cells led to selective cytotoxicity to SN34507 that was associated with cell cycle arrest and generated a robust 'bystander effect' at tissue-like cell densities when only 3% of cells were NfsA positive. Anti-tumor activity of SN35539, the phosphate pre-prodrug of SN34507, was established in 'mixed' tumors harboring a minority of NfsA-positive cells and demonstrated marked tumor control following heterogeneous suicide gene expression. These experiments demonstrate that off-target metabolism of PR-104 can be avoided and identify the suicide gene/prodrug partnership of E. coli NfsA/SN35539 as a promising combination for development in armed vectors.
authors.affiliation: Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Auckland 1023, New Zealand.
authors.forename: Alexandra M
authors.initials: AM
authors.lastname: Mowday
authors.affiliation: Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Auckland 1023, New Zealand.
authors.forename: Amir
authors.initials: A
authors.lastname: Ashoorzadeh
authors.affiliation: School of Biological Sciences, Victoria University of Wellington, Wellington 6140, New Zealand.
authors.forename: Elsie M
authors.initials: EM
authors.lastname: Williams
authors.affiliation: School of Biological Sciences, Victoria University of Wellington, Wellington 6140, New Zealand.
authors.forename: Janine N
authors.initials: JN
authors.lastname: Copp
authors.affiliation: Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Auckland 1023, New Zealand.
authors.forename: Shevan
authors.initials: S
authors.lastname: Silva
authors.affiliation: Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Auckland 1023, New Zealand.
authors.forename: Matthew R
authors.initials: MR
authors.lastname: Bull
authors.affiliation: Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Auckland 1023, New Zealand.
authors.forename: Maria R
authors.initials: MR
authors.lastname: Abbattista
authors.affiliation: Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Auckland 1023, New Zealand; Maurice Wilkins Centre for Molecular Biodiscovery, School of Biological Sciences, The University of Auckland, Auckland 1023, New Zealand.
authors.forename: Robert F
authors.initials: RF
authors.lastname: Anderson
authors.affiliation: Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Auckland 1023, New Zealand; Maurice Wilkins Centre for Molecular Biodiscovery, School of Biological Sciences, The University of Auckland, Auckland 1023, New Zealand.
authors.forename: Jack U
authors.initials: JU
authors.lastname: Flanagan
authors.affiliation: Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Auckland 1023, New Zealand; Maurice Wilkins Centre for Molecular Biodiscovery, School of Biological Sciences, The University of Auckland, Auckland 1023, New Zealand.
authors.forename: Christopher P
authors.initials: CP
authors.lastname: Guise
authors.affiliation: School of Biological Sciences, Victoria University of Wellington, Wellington 6140, New Zealand; Maurice Wilkins Centre for Molecular Biodiscovery, School of Biological Sciences, The University of Auckland, Auckland 1023, New Zealand.
authors.forename: David F
authors.initials: DF
authors.lastname: Ackerley
authors.affiliation: Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Auckland 1023, New Zealand; Maurice Wilkins Centre for Molecular Biodiscovery, School of Biological Sciences, The University of Auckland, Auckland 1023, New Zealand.
authors.forename: Jeff B
authors.initials: JB
authors.lastname: Smaill
authors.affiliation: Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Auckland 1023, New Zealand; Maurice Wilkins Centre for Molecular Biodiscovery, School of Biological Sciences, The University of Auckland, Auckland 1023, New Zealand. Electronic address: a.patterson@auckland.ac.nz.
authors.forename: Adam V
authors.initials: AV
authors.lastname: Patterson
mesh_terms: D015096:3-Hydroxysteroid Dehydrogenases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D065767:Activation, Metabolic / Q000187:drug effects; D000074425:Aldo-Keto Reductase Family 1 Member C3; D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology / Q000627:therapeutic use*; D001549:Benzamides / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology / Q000627:therapeutic use*; D002277:Carcinoma / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D049109:Cell Proliferation / Q000187:drug effects; D015179:Colorectal Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D015195:Drug Design; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D045325:HCT116 Cells; D006801:Humans; D006911:Hydroxyprostaglandin Dehydrogenases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008698:Mesylates / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology / Q000627:therapeutic use*; D008819:Mice, Nude; D008958:Models, Molecular; D062105:Molecular Docking Simulation; D009601:Nitroreductases / Q000235:genetics / Q000378:metabolism; D063065:Organophosphonates / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology / Q000627:therapeutic use*; D011355:Prodrugs / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology / Q000627:therapeutic use*; D011897:Random Allocation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013047:Specific Pathogen-Free Organisms; D013379:Substrate Specificity; D016019:Survival Analysis; D047368:Tumor Burden / Q000187:drug effects; D023041:Xenograft Model Antitumor Assays
chemical_list: D018906:Antineoplastic Agents, Alkylating; D001549:Benzamides; D004791:Enzyme Inhibitors; D029968:Escherichia coli Proteins; D008698:Mesylates; D063065:Organophosphonates; D011355:Prodrugs; D011994:Recombinant Proteins; C000618337:SN34507; C000618338:SN35539; D015096:3-Hydroxysteroid Dehydrogenases; D006911:Hydroxyprostaglandin Dehydrogenases; C487341:AKR1C3 protein, human; D000074425:Aldo-Keto Reductase Family 1 Member C3; C116053:NfsA protein, E coli; D009601:Nitroreductases
keywords: Aldo-keto reductase 1C3; Alkylation; Hypoxia; Oxidoreductase; PR-104 (PubChem CID: 11455973); PR-104A (PubChem CID: 9848786); Prodrug; SN34037 (PubChem CID: 73671441); SN34507 (PubChem CID: 90043967); SN35539 (PubChem CID: 90043246)