Objective To evaluate the potentially improved therapeutic efficacy and safety of nephrotropic macromolecular prodrugs of glucocorticoids (GC) in the treatment of lupus nephritis. Methods Monthly injection of N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer-based dexamethasone prodrug (P-Dex) and daily injection of dexamethasone phosphate sodium (Dex, overall dose equivalent to P-Dex) were given to lupus-prone (NZB×NZW)F1 female mice for two months. The animals were monitored for albuminuria, mean arterial pressure and serum autoantibody levels during the treatment. Nephritis, renal immune complexes and macrophage infiltration were evaluated histologically. The bone quality was analyzed with pDEXA and μ-CT. Optical imaging, immunohistochemistry (IHC) and fluorescence-activated cell sorting (FACS) were used to understand the in vivo distribution of P-Dex. The anti-inflammatory effect of P-Dex was validated using LPS-activated human proximal tubule epithelial cells (HK-2). Results Monthly P-Dex injection completely abolished albuminuria in the (NZB×NZW)F1 mice, which is significantly (P < 0.001) more efficacious than daily Dex treatment. P-Dex did not reduce serum levels of anti-dsDNA antibodies or renal immune complexes, but did reduce macrophage infiltration, a marker of chronic inflammation. IHC and FACS analyses revealed that P-Dex was primarily sequestered by proximal tubule epithelial cells and it could attenuate the inflammatory response in HK-2 cell culture. Different from Dex treatment, P-Dex did not lead to any significant bone quality deterioration or total serum IgG reduction. Conclusion Macromolecularization of GCs renders them nephrotropic. The protracted retention, subcellular processing and activation of GC prodrugs by kidney cells would potentiate nephritis resolution with reduced risk of systemic toxicities.

We evaluated the ability of a macromolecular prodrug of dexamethasone (P-Dex) to treat lupus nephritis in (NZB × NZW)F1 mice. We also explored the mechanism underlying the anti-inflammatory effects of this prodrug. P-Dex eliminated albuminuria in most (NZB × NZW)F1 mice. Furthermore, P-Dex reduced the incidence of severe nephritis and extended lifespan in these mice. P-Dex treatment also prevented the development of lupus-associated hypertension and vasculitis. Although P-Dex did not reduce serum levels of anti-dsDNA antibodies or glomerular immune complexes, P-Dex reduced macrophage recruitment to the kidney and attenuated tubulointerstitial injury. In contrast to what was observed with free dexamethasone, P-Dex did not induce any deterioration of bone quality. However, P-Dex did lead to reduced peripheral white blood cell counts and adrenal gland atrophy. These results suggest that P-Dex is more effective and less toxic than free dexamethasone for the treatment of lupus nephritis in (NZB × NZW)F1 mice. Furthermore, the data suggest that P-Dex may treat nephritis by attenuating the renal inflammatory response to immune complexes, leading to decreased immune cell infiltration and diminished renal inflammation and injury.

To characterize the autoimmune phenomena in patients receiving procainamide, we studied the prevalence of positive direct antiglobulin (Coombs') tests and immune hemolytic anemia in 100 such patients and compared them with 100 age-matched and sex-matched controls. There was a significant increase in the frequency of positive direct antiglobulin tests in patients receiving procainamide (21 vs. 10 per cent, P = 0.05). The mechanism of red-cell sensitization in patients receiving procainamide was the production of red-cell autoantibody, which was serologically indistinguishable from that seen in warm autoimmune hemolytic anemia. In contrast, positive direct antiglobulin tests in control patients were due to the presence of complement components. Red-cell autoantibody production secondary to procainamide was not correlated with a higher-than-expected frequency of antinuclear antibodies or the clinical syndrome of drug-induced lupus erythematosus. In the series of 100 patients receiving procainamide, we identified three cases of immune hemolytic anemia. In two of the three cases, the anemia resolved after the medication was discontinued and did not require steroid therapy. We conclude that procainamide often results in the production of red-cell autoimmune phenomena.

Lupus is an autoimmune disease characterized by the development of antinuclear autoantibodies and immune complex-mediated tissue damage. T cells in lupus patients appear to undergo apoptosis at an increased rate, and this enhanced T cell apoptosis has been postulated to contribute to lupus pathogenesis by increasing autoantigen load. However, there is no direct evidence to support this hypothesis. In this study, we show that an Lck-cre transgene, which increases T cell apoptosis as a result of T cell specific expression of cre recombinase, accelerates the development of autoantibodies and nephritis in lupus-prone (NZB×NZW)F1 mice. Although the enhanced T cell apoptosis in Lck-cre transgenic mice resulted in an overall decrease in the relative abundance of splenic CD4+ and CD8+ T cells, the proportion of activated CD4+ T cells was increased and no significant change was observed in the relative abundance of suppressive T cells. We postulate that the Lck-cre transgene promoted lupus by enhancing T cells apoptosis, which, in conjunction with the impaired clearance of apoptotic cells in lupus-prone mice, increased the nuclear antigen load and accelerated the development of anti-nuclear autoantibodies. Furthermore, our results also underscore the importance of including cre-only controls in studies using the cre-lox system.

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