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Structure-Activity Relationships for CYP4B1 Bioactivation of 4-Ipomeanol Congeners: Direct Correlation between Cytotoxicity and Trapped Reactive Intermediates.
Chem Res Toxicol 2019; 32(12):2488-2498CR

Abstract

Cytochrome P450 4B1 (CYP4B1) has been explored as a candidate enzyme in suicide gene systems for its ability to bioactivate the natural product 4-ipomeanol (IPO) to a reactive species that causes cytotoxicity. However, metabolic limitations of IPO necessitate discovery of new "pro-toxicant" substrates for CYP4B1. In the present study, we examined a series of synthetically facile N-alkyl-3-furancarboxamides for cytotoxicity in HepG2 cells expressing CYP4B1. This compound series maintains the furan warhead of IPO while replacing its alcohol group with alkyl chains of varying length (C1-C8). Compounds with C3-C6 carbon chain lengths showed similar potency to IPO (LD50 ≈ 5 μM). Short chain analogs (<3 carbons) and long chain analogs (>6 carbons) exhibited reduced toxicity, resulting in a parabolic relationship between alkyl chain length and cytotoxicity. A similar parabolic relationship was observed between alkyl chain length and reactive intermediate formation upon trapping of the putative enedial as a stable pyrrole adduct in incubations with purified recombinant rabbit CYP4B1 and common physiological nucleophiles. These parabolic relationships reflect the lower affinity of shorter chain compounds for CYP4B1 and increased ω-hydroxylation of the longer chain compounds by the enzyme. Furthermore, modest time-dependent inhibition of CYP4B1 by N-pentyl-3-furancarboxamide was completely abolished when trapping agents were added, demonstrating escape of reactive intermediates from the enzyme after bioactivation. An insulated CYP4B1 active site may explain the rarely observed direct correlation between adduct formation and cell toxicity reported here.

Authors+Show Affiliations

Department of Medicinal Chemistry, School of Pharmacy , University of Washington , Seattle , Washington 98105 , United States.Department of Medicinal Chemistry, School of Pharmacy , University of Washington , Seattle , Washington 98105 , United States.Department of Medicinal Chemistry, School of Pharmacy , University of Washington , Seattle , Washington 98105 , United States.Department of Medicinal Chemistry, School of Pharmacy , University of Washington , Seattle , Washington 98105 , United States.Department of Medicinal Chemistry, School of Pharmacy , University of Washington , Seattle , Washington 98105 , United States.Institute of Biochemistry , Heinrich-Heine University , 40225 Düsseldorf , Germany.Department of Pediatrics III, University Children's Hospital Essen , University of Duisburg-Essen , 45122 Essen , Germany.Department of Otorhinolaryngology and Head/Neck Surgery , Heinrich-Heine University , 40225 Düsseldorf , Germany.Department of Medicinal Chemistry, School of Pharmacy , University of Washington , Seattle , Washington 98105 , United States.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

31799839

Citation

Kowalski, John P., et al. "Structure-Activity Relationships for CYP4B1 Bioactivation of 4-Ipomeanol Congeners: Direct Correlation Between Cytotoxicity and Trapped Reactive Intermediates." Chemical Research in Toxicology, vol. 32, no. 12, 2019, pp. 2488-2498.
Kowalski JP, McDonald MG, Whittington D, et al. Structure-Activity Relationships for CYP4B1 Bioactivation of 4-Ipomeanol Congeners: Direct Correlation between Cytotoxicity and Trapped Reactive Intermediates. Chem Res Toxicol. 2019;32(12):2488-2498.
Kowalski, J. P., McDonald, M. G., Whittington, D., Guttman, M., Scian, M., Girhard, M., ... Rettie, A. E. (2019). Structure-Activity Relationships for CYP4B1 Bioactivation of 4-Ipomeanol Congeners: Direct Correlation between Cytotoxicity and Trapped Reactive Intermediates. Chemical Research in Toxicology, 32(12), pp. 2488-2498. doi:10.1021/acs.chemrestox.9b00330.
Kowalski JP, et al. Structure-Activity Relationships for CYP4B1 Bioactivation of 4-Ipomeanol Congeners: Direct Correlation Between Cytotoxicity and Trapped Reactive Intermediates. Chem Res Toxicol. 2019 Dec 16;32(12):2488-2498. PubMed PMID: 31799839.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Structure-Activity Relationships for CYP4B1 Bioactivation of 4-Ipomeanol Congeners: Direct Correlation between Cytotoxicity and Trapped Reactive Intermediates. AU - Kowalski,John P, AU - McDonald,Matthew G, AU - Whittington,Dale, AU - Guttman,Miklos, AU - Scian,Michele, AU - Girhard,Marco, AU - Hanenberg,Helmut, AU - Wiek,Constanze, AU - Rettie,Allan E, Y1 - 2019/12/04/ PY - 2019/12/5/pubmed PY - 2019/12/5/medline PY - 2019/12/5/entrez SP - 2488 EP - 2498 JF - Chemical research in toxicology JO - Chem. Res. Toxicol. VL - 32 IS - 12 N2 - Cytochrome P450 4B1 (CYP4B1) has been explored as a candidate enzyme in suicide gene systems for its ability to bioactivate the natural product 4-ipomeanol (IPO) to a reactive species that causes cytotoxicity. However, metabolic limitations of IPO necessitate discovery of new "pro-toxicant" substrates for CYP4B1. In the present study, we examined a series of synthetically facile N-alkyl-3-furancarboxamides for cytotoxicity in HepG2 cells expressing CYP4B1. This compound series maintains the furan warhead of IPO while replacing its alcohol group with alkyl chains of varying length (C1-C8). Compounds with C3-C6 carbon chain lengths showed similar potency to IPO (LD50 ≈ 5 μM). Short chain analogs (<3 carbons) and long chain analogs (>6 carbons) exhibited reduced toxicity, resulting in a parabolic relationship between alkyl chain length and cytotoxicity. A similar parabolic relationship was observed between alkyl chain length and reactive intermediate formation upon trapping of the putative enedial as a stable pyrrole adduct in incubations with purified recombinant rabbit CYP4B1 and common physiological nucleophiles. These parabolic relationships reflect the lower affinity of shorter chain compounds for CYP4B1 and increased ω-hydroxylation of the longer chain compounds by the enzyme. Furthermore, modest time-dependent inhibition of CYP4B1 by N-pentyl-3-furancarboxamide was completely abolished when trapping agents were added, demonstrating escape of reactive intermediates from the enzyme after bioactivation. An insulated CYP4B1 active site may explain the rarely observed direct correlation between adduct formation and cell toxicity reported here. SN - 1520-5010 UR - https://www.unboundmedicine.com/medline/citation/31799839/Structure-Activity_Relationships_for_CYP4B1_Bioactivation_of_4-Ipomeanol_Congeners:_Direct_Correlation_between_Cytotoxicity_and_Trapped_Reactive_Intermediates L2 - https://dx.doi.org/10.1021/acs.chemrestox.9b00330 DB - PRIME DP - Unbound Medicine ER -