Transgenic Expression of the Human MRP2 Transporter Reduces Cisplatin Accumulation and Nephrotoxicity in Mrp2-Null Mice

The chemotherapeutic drug cisplatin is actively transported into proximal tubules, leading to acute renal injury. Previous studies suggest that the multidrug resistance–associated protein 2 (Mrp2) transporter may efflux cisplatin conjugates from cells. We sought to determine whether the absence of Mrp2 alters the accumulation and toxicity of platinum in the kidneys of mice and whether transgenic expression of the human MRP2 gene could protect against cisplatin injury in vivo. Plasma, kidneys, and livers from vehicle- and cisplatin-treated wild-type and Mrp2-null mice were collected for quantification of platinum and toxicity. By 24 hours, twofold higher concentrations of platinum were detected in the kidneys and livers of Mrp2-null mice compared with wild types. Enhanced platinum concentrations in Mrp2-null mice were observed in DNA and cytosolic fractions of the kidneys. Four days after cisplatin treatment, more extensive proximal tubule injury was observed in Mrp2-null mice compared with wild-type mice. Kidneys from naive Mrp2-null mice had elevated glutathione S-transferase mRNA levels, which could increase the formation of cisplatin-glutathione conjugates that may be metabolized to toxic thiol intermediates. Transgenic expression of the human MRP2 gene in Mrp2-null mice reduced the accumulation and nephrotoxicity of cisplatin to levels observed in wild-type mice. These data suggest that deficiency in Mrp2 lowers platinum excretion and increases susceptibility to kidney injury, which can be rescued by the human MRP2 ortholog.Cisplatin is commonly used in chemotherapy regimens for the treatment of solid cancers. The success of cisplatin therapy is limited, in part, by kidney injury. Up to 37% of patients develop signs of nephrotoxicity after receiving a single dose of cisplatin despite strategies such as hydration to limit renal exposure.¹ This is problematic for patients because kidney injury can delay further treatment and limit the total number of chemotherapy cycles received, thereby reducing the overall efficacy of cisplatin-containing regimens.Previous studies have identified the contribution of uptake and efflux transporters to the renal secretion of cisplatin. The organic cation transporter 2 (Oct2; gene Slc22a2) is highly expressed in the kidneys and mediates the renal uptake of cisplatin.^2–5 Transgenic mice lacking Oct1 and Oct2 transporters exhibit decreased platinum (Pt) excretion and reduced nephrotoxicity.⁶ In addition, patients with cancer with a loss-of-function polymorphism in OCT2 (808G>T) are protected against cisplatin renal injury.^6,7 Additional data suggest that the copper transporter 1 (Ctr1; gene Slc31a1) participates in the renal uptake of cisplatin,^8–12 although recent work has found conflicting results.¹³ After uptake of cisplatin by Oct2 or Ctr1, the subsequent efflux of cisplatin into the renal lumen is mediated by the multidrug and toxin extrusion protein 1 (Mate1; gene Slc47a1).¹⁴ As expected, Mate1 knockout mice have increased renal concentrations of cisplatin and enhanced nephrotoxicity.¹⁴Once inside the renal cell, cisplatin is biotransformed by spontaneous hydrolysis to monoaquated and diaquated species. Formation of cisplatin aquated intermediates leads to electrophilic attack of cellular macromolecules, such as DNA and protein. The tripeptide glutathione (GSH) can also bind cisplatin and has been suggested to form a variety of nontoxic cisplatin conjugates, including monoplatinum-monoglutathione, diplatinum-monoglutathione, and bis-(glutathionato)-platinum.^15,16 It has been postulated that these conjugates have the potential for metabolism to reactive thiol intermediates. Previous studies suggest that the multidrug resistance–associated transporter 2 (Mrp2; gene Abcc2) can efflux GSH conjugates of cisplatin from cancer cells and confer resistance to cytotoxicity.^15,17,18 Overexpression of human (h)MRP2 increases cisplatin resistance by 10-fold in human embryonic kidney cells¹⁷ and reduces cisplatin accumulation by 30% in porcine kidney LLC-PK1 cells.¹⁸ Moreover, primary hepatocytes isolated from transport-deficient rats (Mrp2-deficient strain) exhibit increased binding of Pt to DNA and enhanced susceptibility to cisplatin cytotoxicity.¹⁹ However, the ability of Mrp2 to reduce the accumulation of cisplatin and/or its conjugates and limit renal injury in vivo is not well understood. Therefore, the purpose of this study was to determine whether loss of Mrp2 function enhances renal Pt accumulation and nephrotoxicity in mice and to evaluate the ability of the human MRP2 gene to rescue Mrp2-null mice from heightened cisplatin-induced nephrotoxicity in vivo.