Engineering Macaca fascicularis cytochrome P450 2C20 to reduce animal testing for new drugs

Rua, Francesco; Sadeghi, Sheila J.; Castrignanò, Silvia; Nardo, Giovanna Di; Gilardi, Gianfranco

doi:10.1016/j.jinorgbio.2012.05.017

Cited by 16 publications

(15 citation statements)

References 42 publications

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“…The surface coverage value on GC/PDDA was calculated to be 1.1X10 12 molecules/cm 2 in the same range previously reported for the Macaca fascicularis P450 2C20 immobilized in the same manner [25]. As expected the same calculation of the surface coverage in the presence of AuNps resulted in a value of 4.6x10 12 molecules/cm 2 , around 4 times higher than using PDDA alone.…”

Section: Direct Electrochemistry Of P450 2d15 On Glassy Carbon Electrsupporting

confidence: 84%

“…Canis familiaris is one of the most widely studied animal models used in safety determination of new pharmaceuticals [11][12] and, although the major isoforms of the human cytochromes P450 2D6, 3A4, 2E1, 2C19 and 1A2 have been identified in C. familiaris [13][14][15][16], there is still a lack of knowledge on their pharmacogenomic/metabolic diversity [9]. Electrochemical techniques already developed in our lab for human hepatic monooxygenases including cytochromes P450 [17][18][19][20][21][22] represent the ideal approach for a sensitive, accurate and rapid evaluation of animal P450-drug interactions obviating both the requirement for a redox partner and the addition of NADPH cofactor as already reported for some animal P450 enzymes recombinantly expressed in a soluble form [23][24][25]. In this work, Canis familiaris P450 2D15 was chosen as a model for the investigation of canine cytochromes P450 by adopting electrochemical approaches to provide a method for the screening of the safety of new chemical entities/drugs.…”

Section: Introductionmentioning

confidence: 99%

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Electrochemistry of Canis familiaris cytochrome P450 2D15 with gold nanoparticles: An alternative to animal testing in drug discovery

Rua

Sadeghi

Castrignanò

et al. 2015

Bioelectrochemistry

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This work reports for the first time the direct electron transfer of the Canis familiaris cytochrome P450 2D15 on glassy carbon electrodes to provide an analytical tool as an alternative to P450 animal testing in the drug discovery process. Cytochrome P450 2D15, that corresponds to the human homologue P450 2D6, was recombinantly expressed in Escherichia coli and entrapped on glassy carbon electrodes (GC) either with the cationic polymer polydiallyldimethylammonium chloride (PDDA) or in the presence of gold nanoparticles (AuNPs). Reversible electrochemical signals of P450 2D15 were observed with calculated midpoint potentials (E1/2) of −191 ± 5 and −233 ± 4 mV vs. Ag/AgCl for GC/PDDA/2D15 and GC/AuNPs/2D15, respectively. These experiments were then followed by the electro-catalytic activity of the immobilized enzyme in the presence of metoprolol. The latter drug is a beta-blocker used for the treatment of hypertension and is a specific marker of the human P450 2D6 activity. Electrocatalysis data showed that only in the presence of AuNps the expected α-hydroxy-metoprolol product was present as shown by HPLC. The successful immobilization of the electroactive C. familiaris cytochrome P450 2D15 on electrode surfaces addresses the ever increasing demand of developing alternative in vitromethods for amore detailed study of animal P450 enzymes' metabolism, reducing the number of animals sacrificed in preclinical tests.

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Section: Direct Electrochemistry Of P450 2d15 On Glassy Carbon Electrsupporting

confidence: 84%

Section: Introductionmentioning

confidence: 99%

Electrochemistry of Canis familiaris cytochrome P450 2D15 with gold nanoparticles: An alternative to animal testing in drug discovery

Rua

Sadeghi

Castrignanò

et al. 2015

Bioelectrochemistry

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“…Owing to P450 BM3 characteristics, multiple protein engineering studies have been performed on this enzyme to widen its catalytic abilities . Moreover, several constructions have been already reported including different fusion human P450 enzymes, engineered by connecting the P450 BM3 reductase domain with human cytochromes P450 3A4, 2C9, 2C19 , 2A6, CYP2C6, and CYP4F11 , monkey 2C20 , and dog CYP2D15 . Also the catalytic performance of one of the created chimeric proteins was improved in terms of coupling efficiency and enzyme turnover by engineering the loop connecting the two domains .…”

Section: Biopharmaceuticalsmentioning

confidence: 99%

Progress in biopharmaceutical development

Kęsik-Brodacka

2017

Biotech and App Biochem

267

126

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Since its introduction in 1982, biopharmaceutical drugs have revolutionized the treatment of a broad spectrum of diseases and are increasingly used in nearly all branches of medicine. In recent years, the biopharmaceuticals market has developed much faster than the market for all drugs and is believed to have great potential for further dynamic growth because of the tremendous demand for these drugs. Biobetters, which contain altered active pharmaceutical ingredients with enhanced efficacy, will play an important role in the development of biopharmaceuticals. Another significant group of biopharmaceuticals are biosimilars. Their introduction in the European Union and, recently, the Unites States markets will reduce the costs of biopharmaceutical treatment. This review highlights recent progress in the field of biopharmaceutical development and issues concerning the registration of innovative biopharmaceuticals and biosimilars. The leading class of biopharmaceuticals, the current biopharmaceuticals market, and forecasts are also discussed.

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“…For this reason we already reported the construction of different fusion human P450 enzymes, engineered by connecting the reductase domain BMR with human cytochromes P450 2E1 (Fairhead et al, 2005), 3A4, 2C9, 2C19 (Dodhia et al, 2006), 2A6, CYP2C6, and CYP4F11 (Ortolani, 2012;Rua, 2012a;Castrignanò et al, 2014), monkey 2C20 (Rua et al, 2012b) and dog CYP2D15 (Sadeghi and Gilardi, 2013;Rua et al, 2015).…”

Section: Cytochrome C Reduction Activitiesmentioning

confidence: 99%

Role of Protein-Protein Interactions in Metabolism: Genetics, Structure, Function

Pandey¹,

Henderson²,

Ishii³

et al. 2018

Frontiers Research Topics

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Engineering Macaca fascicularis cytochrome P450 2C20 to reduce animal testing for new drugs

Cited by 16 publications

References 42 publications

Electrochemistry of Canis familiaris cytochrome P450 2D15 with gold nanoparticles: An alternative to animal testing in drug discovery

Electrochemistry of Canis familiaris cytochrome P450 2D15 with gold nanoparticles: An alternative to animal testing in drug discovery

Progress in biopharmaceutical development

Role of Protein-Protein Interactions in Metabolism: Genetics, Structure, Function

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