News from SOLVO: Webinar Series, R&D News, Publications, Automated Transporter Screening Assays

July 16, 2020

MEET THE EXPERTS GOES ONLINE!

Like much of the world, we are limiting our travel and putting face-to-face meetings on hold as a result of the COVID-19 pandemic. The pace of scientific progress continues however, and with it the need for discourse and debate. Since taking the difficult decision to postpone our Meet the Experts Transporter Conference at the start of the year, we have received a significant level of interest from our colleagues in the transporter field for an alternative venue in which to showcase the latest in transporter research. We are therefore delighted to announce the start of the SOLVO Biotechnology Meet the Experts Transporter Webinar Series! The first event will take place on Thursday, July 16th at 5pm CEST (11am ET) and will feature Dr. Emmanuelle Reboul, Head of the Bioavailability of Micronutrients Research Axis at the Cardiovascular and Nutrition Center in Marseille, France, who will be speaking on the topic of ‘New Insights on Fat-Soluble Vitamin Efflux by the Intestine’. For further information and free registration please see the SOLVO website.

R&D NEWS – NEW ASSAYS FOR THTR1, THTR2, MDR1, NIS, OATP2B1 AND BCRP

Despite challenging external conditions, we are pleased to report that the progress of new assay development at SOLVO has not lost impetus in the first half of 2020. With our recently launched THTR1 and THTR2 assays we have entered the rapidly-expanding field of transporter-mediated drugnutrient interactions (Vora et al. (2020) Am. J. Clin. Nutr. 111(1):110). Since the Janus kinase 2 inhibitor fedratinib was shown to cause thiamine deficiencyrelated encephalopathy by potently inhibiting thiamine absorption through the human thiamine transporter 2 (THTR2) (Zhang et al. (2014) Drug Metab. Dispos. 42(10):1656), suppression of nutrient transport by drugs has become a new concern industry-wide. Whereas fedratinib is a niche drug indicated for myelofibrosis, a rare myeloproliferative neoplasm, the commonly prescribed antibiotic trimethoprim was also found to induce thiamine deficiency by inhibiting both THTR1 and THTR2 (Giacomini et al. (2017) Drug Metab. Dispos. 45(1):76). In a white paper released in 2018, the International Transporter Consortium listed THTRs among transporters of emerging clinical importance and underscored their relevance to drug-induced vitamin deficiency (Zamek-Gliszczynski et al. (2018) Clin. Pharmacol. Ther. 104(5):890).

In parallel, we have made another fresh entry into the area of chemical safety with the introduction of our sodium-iodine symporter (NIS) assay. Endocrine disruptors, man-made chemicals that as environmental pollutants interfere with hormonal regulation in humans and animals, have been in the focus of international organizations such as the WHO and the OECD for over two decades, but for long, most attention was directed to disruptors of reproductive health. In 2017, however, the OECD issued a scoping document on the identification of thyroid signaling modulators that lists NIS as a potential target of thyroid-disrupting substances. Compounds identified as inhibitors of active iodide uptake in our HEK293 cell-based NIS assay are liable to suppression of thyroid hormone synthesis in vivo. Importantly, increasing substitution of traditional animal chemical safety tests with in vitro systems such as the NIS inhibition assay is a crucial contribution to the ‘3Rs’ of animal welfare, and we are excited to add this new assay to the suite of in vitro endocrine disruptor testing services available from our parent company, Charles River.

We have continued to make improvements to our existing assays, and a recent major development in this area has been to our pre-existing MDCKII-MDR1 vectorial transport assay, which has been modified to knockout the endogenous canine P-glycoprotein (P-gp; Abcb1). Prior to this, the presence of endogenous canine P-gp found in MDCKII cells contributed to background efflux activity and potential ambiguity when interpreting transport data. In contrast, our new human MDR1 assay based on the Abcb1KOMDCKII cell host provides a superior assay with greater clarity in data interpretation. By genetically ablating canine Abcb1, the Abcb1-KO-MDCKII control cells display no directional transport of prototypic MDR1 substrates, and the human transporter is solely responsible for MDR1 substrate efflux in Abcb1-KO-MDCKII-MDR1 cells. The assay has been validated for use with digoxin and quinidine as probe substrates in both 24-well and 96-well formats.

In support of industry moves toward more physiologically-relevant transporter probes, we have updated our OATP2B1 uptake and BCRP vesicular transport assays to utilize rosuvastatin as a probe substrate. These assays join our OATP1B1 and OATP1B3 uptake assays, which we previously reported have been developed for use with rosuvastatin. In each case, rosuvastatin is accurately and sensitively quantified by LC-MS/MS

AUTOMATED TRANSPORTER SCREENING ASSAYS

As the transporter field evolves and our knowledge of the clinical significance of transporters deepens, we are finding that our industry partners are incorporating transporter testing earlier in development. Furthermore, this early transporter screening approach which was originally championed by scientists at large pharma is now being increasingly adopted by their counterparts at smaller biotechs. In order to support this demand, our team of transporter scientists and automation experts have been working to transfer our industryleading transporter assays onto an automated platform. As a pioneer in the field of drug transporter products and services, SOLVO is the only transporter-focused CRO to generate all our cell lines, membrane vesicle reagents, and assay methodologies inhouse. This depth of understanding of each experimental system involved has proved to be a unique advantage, allowing us to transfer assays quickly, cost effectively, and most importantly without any loss of the high level of quality our clients have come to expect from SOLVO. The first automated assay to be launched on this platform is bile salt export pump (BSEP; ABCB11) vesicular transport inhibition. Inhibition of BSEP is viewed as a potential risk factor for drug-induced liver injury (DILI), and this assay is being increasingly used as an early screen for potentially hepatotoxic compounds early in development (see Morgan et al. (2010) Toxicol. Sci. 118(2):485; Dawson et al. (2011) Drug Metab. Dispos. 40(1):130).

Contact our team today to discuss how early BSEP inhibition screening can be utilized as part of a strategy to de-risk your development pipeline, and stay in touch to learn more about other automated transporter assays that we will be launching in the near future!

SOLVO PUBLICATIONS IN 2019

Looking back, the past year was a productive one for SOLVO in which we authored or co-authored 13 papers.

Characterization, optimization and suggestions for assay selection have always been in the forefront of our scientific and business agenda. In collaboration with scientists from Novartis we showed that preincubation significantly affects IC50 values for several influx transporters leading to reassignment of clinical relevance of the interactions (Tatrai et al. (2019) Drug Metab. Dispos. 47:768).

Application of assays to address scientific questions was also an important field as in collaboration we have shown that testosterone-glucuronides are transported by MRP2 and MRP3 in human liver, intestine and kidney (Li et al. (2019) J Steroid Biochem. Mol. Biol. 191:105350). In addition, P-gp, BCRP and MRP2 were found to be involved in transplacental pharmacokinetics of maraviroc (Tupova et al. (2019) Drug Metab. Dispos. 47:954) and MATE1 reported to play an important role in renal excretion of flecainide (Doki et al. (2019) Biol. Pharm. Bull. 42:1226). In contrast, CNT2 does not transport tenofovir alafenamide and tenofovir disoproxil fumarate (Cusato et al. (2019) Diagn. Microbiol. Infect. Dis. 94:202). These studies were all collaborations with colleagues at the University of Washington (Seattle, USA; Li et al.), Charles University (Prague, Czech Republic; Tupova et al.), the University of Tsukuba (Tsukuba, Japan; Doki et al.) and the University of Turin (Torino, Italy; Cusato et al.).

Many natural products are occupying the same chemical space as drugs and they are substrates and inhibitors of transporters of xenobiotics. In collaboration with University of Szeged scientists, we reported that green tea inhibited gastrointestinal absorption of the antibiotic amoxicillin in rats pointing to a chance of a nutrient – drug interaction (Kiss et al. (2019) BMC Pharmacol. Toxicol. 20:54).

Last but by no means least, we contributed to enrichment of the transporter scientific knowledge-base by writing reviews on BCRP substrates and inhibitors (Safar et al. (2019) Exp. Opin. Drug Metab. Toxicol. 15:313) and, in collaboration with Péter Pázmány Catholic University, on age-related functional and expressional changes of efflux pathways in the blood-brain barrier (Erdő and Krajcsi (2019) Front. Aging Neurosci. 11:196).

As we continue to make progress towards improving our understanding of transporters in 2020 and beyond, we would especially thank all past and present collaborators for their invaluable support. If you have a transporter project that you would like to collaborate on, please contact your local SOLVO representative to arrange a meeting with our team of scientists!


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