While HIV/AIDS can be managed with antiretroviral drugs that block viral replication, these agents do not clear the virus and require life-long drug administration with associated risks of cumulative toxicity and drug resistance. Recently, we discovered a completely new class of compounds that interfere with the HIV-1 virulence factor, Nef. This viral protein is critical to HIV-1 replication in vivo, immune escape of HIV-infected cells, and AIDS progression. Nef antagonists have the potential to synergize with current antiretroviral drugs, thereby reducing toxicities and the risk of drug resistance. Nef inhibitors may also restore immune recognition of HIV-infected cells, potentially clearing the virus. Our lead Nef inhibitors are based on a diphenylpyrazolodiazene (DPP) scaffold that binds directly to the HIV-1 Nef protein and blocks its myriad functions in vitro. The goal of this Phase I STTR grant application, therefore, is to exploit our discovery of novel HIV Nef inhibitors through the design and preparation of new analogs that enable us to understand SAR, improve potency, and optimize ADME/PK properties. Successful completion of Phase I will form the basis of an expanded drug development program in the Phase II period. These goals will be accomplished by combining the pharmaceutical and medicinal chemistry expertise of scientists at the Fox Chase Chemical Diversity Center, Inc. with the expertise of the Smithgall group at the University of Pittsburgh in the preclinical aspects of HIV antiviral therapy targeting Nef. We will realize these goals through the following specific aims:
Aim 1 : Identify structure-activity relationships (SAR) governing interaction of the lead compounds with HIV Nef to improve potency and ADME/PK properties. Here we will increase Nef inhibitor potency as assessed by direct binding to Nef (via SPR), inhibition of Nef-activated Hck kinase (an essential host cell effector protein for Nef), as well a antiretroviral activity assays (Nef- dependent enhancement of HIV infectivity and replication). Our goal is to reduce the IC50 values in these assays into the nanomolar range, while removing unwanted functionalities present in the parent compound (i.e., diazo linker, NO2 and thiourea groups). We will synthesize ~50 new analogs based on the existing DPP scaffold, and expect to find 3-6 analogs that meet the criteria of this aim.
Aim 2 : Evaluate ADME/PK properties for Nef inhibitors meeting the criteria of Aim 1. We will explore potential drug properties by obtaining in vitro absorption, distribution, metabolism and excretion (ADME) data for up to 10 compounds. These compounds will be evaluated for metabolic stability (mouse and human microsomes), plasma protein binding (mouse and human), CYP inhibition (3A4-midazolam), solubility and permeability (Caco-2 cells). Promising compounds will be evaluated for pharmacokinetic (PK) parameters in mice. Successful completion of these studies will support an expanded Phase II effort with efficacy evaluation in a humanized mouse model of HIV/AIDS and lead us towards an IND application.
The proposed studies are focused on new approaches to HIV/AIDS drug discovery targeting an HIV-1 protein (Nef) essential for AIDS progression. Successful completion of the proposed work will identify small molecules that block Nef in HIV pathogenesis in animal models and ultimately in man. These Nef inhibitors represent a new class of anti-HIV therapeutics.