Tissue acidosis does not mediate the hypoxia selectivity of [(64)Cu][Cu(ATSM)] in the isolated perfused rat heart

Copper-64-Diacetyl-bis(N(4)-methylthiosemicarbazone) [(64)Cu][Cu(ATSM)] is a hypoxia-targeting PET tracer with applications in oncology and cardiology. Upon entering a hypoxic cell, [(64)Cu][Cu(II)(ATSM)] is reduced to a putative [(64)Cu][Cu(I)(ATSM)](−) species which dissociates to deposit radiocopper, thereby providing hypoxic contrast. This process may be dependent upon protonation arising from intracellular acidosis. Since acidosis is a hallmark of ischemic tissue and tumors, the hypoxia specificity of [(64)Cu][Cu(ATSM)] may be confounded by changes in intracellular pH. We have therefore determined the influence of intracellular pH on [(64)Cu][Cu(ATSM)] pharmacokinetics. Using isolated perfused rat hearts, acidosis was induced using an ammonium pre-pulse method, with and without hypoxic buffer perfusion. Cardiac [(64)Cu][Cu(ATSM)] pharmacokinetics were determined using NaI detectors, with intracellular pH and cardiac energetics monitored in parallel by (31)P NMR. To distinguish direct acidotic effects on tracer pharmacokinetics from acidosis-induced hypocontractility, parallel studies used lidocaine perfusion to abolish cardiac contraction. Hypoxic myocardium trapped [(64)Cu][Cu(ATSM)] despite no evidence of it being acidotic when characterised by (31)P NMR. Independent induction of tissue acidosis had no direct effect on [(64)Cu][Cu(ATSM)] pharmacokinetics in either normoxic or hypoxic hearts, beyond decreasing cardiac oxygen consumption to alleviate hypoxia and decrease tracer retention, leading us to conclude that tissue acidosis does not mediate the hypoxia selectivity of [(64)Cu][Cu(ATSM)].