Retention of the Radiotracers ⁶⁴Cu-ATSM and ⁶⁴Cu-PTSM in Human and Murine Tumors Is Influenced by MDR1 Protein Expression

Abstract

Tumor hypoxia is often associated with resistance to chemotherapy. Multidrug resistance type 1 (MDR1) protein is a member of the adenosine triphosphate binding cassette (ABC) proteins, some of which are involved in the multidrug resistance (MDR) phenotype in tumors. Many studies have focused on the role of these proteins in modulating drug resistance, but their effect on retention of imaging agents is less well studied. To study the role of MDR1 expression on the accumulation of 64 Cu-diacetyl-bis(N4-methylthiosemicarbazone) ( 64 Cu-ATSM) and 64 Cu-pyruvaldehyde-bis(N4-methylthiosemicarbazone) ( 64 Cu-PTSM) in human tumors in vitro and in vivo, we used a model system composed of a low MDR1-expressing parent uterine sarcoma cell line and a daughter cell line selected for overexpression of MDR1. Aromatase knockout (ArKO) mice that spontaneously developed liver tumors were used as an additional in vivo model to study the effect of MDR expression on 64 Cu-ATSM and -PTSM retention. Methods: Biodistribution experiments after injection of 64 Cu-ATSM or -PTSM were performed in wild-type mice, ArKO mice, and ArKO mice bearing liver tumors (n 5 3-5/group), and in nude mice bearing human tumor xenografts for in vivo PET/CT. Liver expression of Abcb1a and Abcb1b, the MDR1 proteins in mouse liver, was determined by real-time polymerase chain reaction. 64 Cu-ATSM and -PTSM accumulation and efflux studies were conducted in tumor cell lines. The uptake experiments were repeated after knockdown of MDR1 protein expression using MDR1-specific small interfering RNAs. Results: In vivo, the hepatic tumors had a lower percentage injected dose per gram of 64 Cu-ATSM or -PTSM and more highly expressed Abcb1b than did wild-type liver or nontumor-bearing ArKO liver. High MDR1-expressing tumors showed lower tracer activity on PET/CT images. In vitro, cells highly expressing MDR1 had significantly decreased 64 Cu-ATSM and -PTSM retention and enhanced efflux. Knockdown of MDR1 expression significantly enhanced the 64 Cu-ATSM and -PTSM retention and decreased the efflux in MDR1-positive cells. Conclusion: The expression of MDR1 glycoprotein (or its equivalents in mice) affects the retention of 64 Cu-ATSM and -PTSM in the human and murine tumors tested. These results may have implications for clinical hypoxia imaging in tumors and the therapeutic efficacy of 64 Cu-ATSM.

A Comparison of the Behavior of 64Cu-Acetate and 64Cu-ATSM In Vitro and In Vivo

“…The correlation of intratumoral 64 Cu distribution with nitroimidazole-based markers was also variable and time-dependent (12)(13)(14). This suggests that the strong correlation between retention of radiocopper from Cu-ATSM and poor treatment outcome observed in clinical studies may be governed by additional mechanisms other than tumor pO 2 (11,(15)(16)(17).…”

Positron emission tomography to assess hypoxia and perfusion in lung cancer

“…This tracer shows favorable kinetics with rapid uptake in hypoxic tissue and fast clearance from normoxic tissues, enabling imaging within 30 min after injection [66,67] . However, the exact uptake mechanism of [Cu]ATSM is still under debate [63,68,69] and several preclinical studies have shown that [Cu]ATSM uptake depends on tumor type and other characteristics than hypoxia alone [70][71][72][73][74][75][76] .…”

64Cu-ATSM Reflects pO2 Levels in Human Head and Neck Cancer Xenografts but Not in Colorectal Cancer Xenografts: Comparison with 64CuCl2

“…In hypoxic cells, reoxidation occurs at a slower rate, leaving enough time for dissociation of the unstable [ 64 Cu-ATSM] 2 . The radioactive copper isotope then becomes part of the intracellular copper pool, and some studies have indicated that there also appears to be an efflux of either radiolabeled 64 Cu-ATSM or copper from cancer cells (30)(31)(32). Moreover, studies of copper metabolism using 64 CuCl 2 PET in tumor xenograft mouse models have reported high tumor accumulation in some tissue types (33,34).…”