Multimodal analysis of drug transporter expression in gastrointestinal tissue

Corbin G. Thompson, John K. Fallon, Michelle Mathews, Paige Charlins, Leila Mulder, Martina Kovarova, Lourdes Adamson, Nithya Srinivas, Amanda Schauer, Craig Sykes, Paul A Luciw, Victor V. Garcia, Ramesh Akkina, Philip C. Smith, Angela D.M. Kashuba

Research output: Contribution to journalArticlepeer-review

8 Scopus citations

Abstract

OBJECTIVES:: Drug transporters affect ART tissue disposition, but quantitative measures of drug transporter protein expression across pre-clinical species are not available. Our objective was to use proteomics to obtain absolute transporter concentrations and assess agreement with corresponding gene and immunometric protein data. DESIGN:: In order to make interspecies comparisons, two humanized mouse (hu-HSC-Rag (n?=?41); BLT (n?=?13)) and one primate (rhesus macaque, (NHP, n?=?12)) models were dosed to steady-state with combination ART. Ileum and rectum were collected at necropsy and snap frozen for analysis. METHODS:: Tissues were analyzed for gene (qPCR) and protein (LC-MS proteomics and Western blot) expression and localization (immunohistochemistry) of ART efflux and uptake transporters. Drug concentrations were measured by LC-MS/MS. Multivariable regression was used to determine the ability of transporter data to predict tissue ART penetration. RESULTS:: Analytical methods did not agree, with different trends observed for gene and protein expression. For example, qPCR analysis showed a 2-fold increase in permeability glycoprotein (Pgp) expression in NHPs versus mice, however proteomics showed a 200-fold difference in the opposite direction. Proteomics results were supported by IHC staining showing extensive efflux transporter localization on the luminal surface of these tissues. ART tissue concentration was variable between species, and multivariable regression showed poor predictive power of transporter data. CONCLUSIONS:: Lack of agreement between analytical techniques suggests that resources should be focused on generating downstream measures of protein expression to predict drug exposure. Taken together, these data inform the use of pre-clinical models for studying ART distribution and the design of targeted therapies for HIV eradication.

Original languageEnglish (US)
JournalAIDS
DOIs
StateAccepted/In press - Jun 5 2017

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology
  • Infectious Diseases

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