posted on 2024-01-03, 18:11authored byColten
D. Eberhard, Eric P. Mosher, Namandjé
N. Bumpus, Benjamin C. Orsburn
Tenofovir (TFV) is a nucleotide reverse
transcriptase
inhibitor
prescribed for the treatment and prevention of human immunodeficiency
virus infection and the treatment of chronic hepatitis B virus infection.
Here, we demonstrate that creatine kinase brain-type (CKB) can form
tenofovir-diphosphate (TFV-DP), the pharmacologically active metabolite,
in vitro and identify nine missense mutations (C74S, R96P, S128R,
R132H, R172P, R236Q, C283S, R292Q, and H296R) that diminish this activity.
Additional characterization of these mutations reveals that five (R96P,
R132H, R236Q, C283S, and R292Q) have ATP dephosphorylation catalytic
efficiencies less than 20% of those of the wild type (WT), and seven
(C74S, R96P, R132H, R172P, R236Q, C283S, and H296P) induce thermal
instabilities. To determine the extent CKB contributes to TFV activation
in vivo, we generated a CKB knockout mouse strain, Ckbtm1Nnb. Using an in vitro assay, we show
that brain lysates of Ckbtm1Nnb male and female mice form 70.5 and 77.4% less TFV-DP than
wild-type brain lysates of the same sex, respectively. Additionally,
we observe that Ckbtm1Nnb male mice treated with tenofovir disoproxil fumarate for 14 days
exhibit a 22.8% reduction in TFV activation in the liver compared
to wild-type male mice. Lastly, we utilize mass spectrometry-based
proteomics to elucidate the impact of the knockout on the abundance
of nucleotide and small molecule kinases in the brain and liver, adding
to our understanding of how the loss of CKB may be impacting tenofovir
activation in these tissues. Together, our data suggest that disruptions
in CKB may lower levels of active drugs in the brain and liver.