The first formal qualification of safety biomarkers for regulatory decision making marks a milestone in the application of biomarkers to drug development. Following submission of drug toxicity studies and analyses of biomarker performance to the Food and Drug Administration (FDA) and European Medicines Agency (EMEA) by the Predictive Safety Testing Consortium's (PSTC) Nephrotoxicity Working Group, seven renal safety biomarkers have been qualified for limited use in nonclinical and clinical drug development to help guide safety assessments. This was a pilot process, and the experience gained will both facilitate better understanding of how the qualification process will probably evolve and clarify the minimal requirements necessary to evaluate the performance of biomarkers of organ injury within specific contexts.

Organic anion-transporting polypeptides (OATP) 1B1 and 1B3 are widely acknowledged as important and rate-limiting to the hepatic uptake of many drugs in clinical use. Accordingly, to better understand the in vivo relevance of OATP1B transporters, targeted disruption of murine Slco1b2 gene was carried out. It is noteworthy that Slco1b2(Ϫ/Ϫ) mice were fertile, developed normally, and exhibited no overt phenotypic abnormalities. We confirmed the loss of Oatp1b2 expression in liver using real-time polymerase chain reaction, Western Blot analysis, and immunohistochemistry. Expression of Oatp1a4 and Oatp2b1 but not Oatp1a1 was greater in female Slco1b2(Ϫ/Ϫ) mice, but expression of other non-OATP transporters did not significantly differ between wild-type and Slco1b2(Ϫ/Ϫ) male mice. Total bilirubin level was elevated by 2-fold in the Slco1b2(Ϫ/Ϫ) mice despite the fact that liver enzymes ALT and AST were normal. Pharmacological characterization was carried out using two prototypical substrates of human OATP1B1 and -1B3, rifampin and pravastatin. After a single intravenous dose of rifampin (1 mg/kg), a 1.7-fold increase in plasma area under the concentration-time curve (AUC) was observed, whereas the liver-to-plasma ratio was reduced by 5-fold, and nearly 8-fold when assessed at steady-state conditions after 24 h of continuous subcutaneous infusion in Slco1b2(Ϫ/Ϫ) mice. Likewise, continuous subcutaneous infusion at low (8 g/h) or high (32 g/h) dose rates of pravastatin resulted in a 4-fold lower liver-plasma ratio in the in Slco1b2(Ϫ/Ϫ) mice. This is the first report of altered drug disposition profile in the Slco1b2 knockout mice and suggests the utility of this model for understanding the in vivo role of hepatic OATP transporters in drug disposition.

Macrophage colony-stimulating factor (M-CSF) is a hematopoietic growth factor that is responsible for the survival and proliferation of monocytes and the differentiation of monocytes into macrophages, including Kupffer cells (KCs) in the liver. KCs play an important role in the clearance of several serum enzymes, including aspartate aminotransferase and creatine kinase, that are typically elevated as a result of liver or skeletal muscle injury. We used three distinct animal models to investigate the hypothesis that increases in the levels of serum enzymes can be the result of decreases in KCs in the apparent absence of hepatic or skeletal muscle injury. Specifically, neutralizing M-CSF activity via a novel human monoclonal antibody reduced the CD14 ؉ CD16 ؉ monocyte population, depleted KCs, and increased aspartate aminotransferase and creatine kinase serum enzyme levels in cynomolgus macaques. In addition, the treatment of rats with clodronate liposomes depleted KCs and led to increased serum enzyme levels, again without evidence of tissue injury. Finally, in the osteopetrotic (Csf1 op /Csf1 op ) mice lacking functional M-CSF and having reduced levels of KCs, the levels of serum enzymes are higher than in wild-type littermates. Together, these findings support a mechanism for increases in serum enzyme levels through M-CSF regulation of tissue macrophage homeostasis without concomitant histopathological changes in either the hepatic or skeletal system.

Application of any new biomarker to support safety-related decisions during regulated phases of drug development requires provision of a substantial data set that critically assesses analytical and biological performance of that biomarker. Such an approach enables stakeholders from industry and regulatory bodies to objectively evaluate whether superior standards of performance have been met and whether specific claims of fit-for-purpose use are supported. It is therefore important during the biomarker evaluation process that stakeholders seek agreement on which critical experiments are needed to test that a biomarker meets specific performance claims, how new biomarker and traditional comparators will be measured and how the resulting data will be merged, analyzed and interpreted.

Development of feline immunodeficiency virus (FIV) infection in cats as a small animal model for lentiviral immunodeficiency disease has been hampered by the prolonged and variable disease course following experimental infection. To address this issue, we generated high-titer, unselected FIV stocks by pooling plasma from cats acutely infected with a subgroup C FIV isolate designated CABCpadyOOC (FIV-C-PGammer). Subsequent infection with this virus pool resulted in rapidly progressive, fatal disease in greater than 50% of infected cats. Accelerated FIV disease was characterized by rapid and progressive CD4 ؉ T-cell loss, lymphadenopathy, weight loss, lymphoid depletion, and severe thymic atrophy. Mortality and rate of disease progression were affected by the age of each cat at infection and whether the virus source animal was in the acute or chronic stage of infection. The rapid FIV disease syndrome was consistently associated with systemic lymphoid depletion, clinical disease, and susceptibility to opportunistic infections, analogous to accelerated and/or terminal HIV-1 infection. The results of this study demonstrate that FIV infection is a valid small animal model for lentiviral immunodeficiency disease.