SBIR-STTR Award

We propose to develop rapid, cost-effective methods for detecting drug resistance in human immunodeficiency virus type l (HIV-l) isolates by analyzing the DNA sequences of patient samples. The health of HIV-l/AIDS patients may be better maintained through lower toxicity and more appropriate antiviral use. All currently approved HIV-l antivirals are targeted against reverse transcriptase (RT). Specific mutations in RT confer resistance to these antivirals. The aim of both phases of this SBIR proposal is to develop DNA chips to detect resistance conferring mutations in RT from clinical samples. In Phase I of this research, Affymetrix will demonstrate the feasibility of this approach by: * designing a set of overlapping DNA probes that corresponds to the most common Ziovudine (AZT) resistance conferring mutation site in RT, * synthesizing DNA chips containing arrays of these probes using light directed parallel chemical synthesis, * hybridizing fluorescently labeled target DNA to the chips, and * discriminating resistant and sensitive DNA sequences at this site. Completion of these Phase I goals will demonstrate the feasibility of using DNA chips to detect and monitor mutations in HIV-1 that confer resistance to approved and investigational therapeutics. In Phase II we will extend the DNA chip design to detect all known mutations in RT that confer resistance to approved HIV-l antivirals and investigate the sensitivity of the assay to mixtures of viral genotypes. In Phase Ill we will test the chips in a clinical laboratory setting and investigate DNA chips to resequence the entire pol gene. This will allow application of the chips to detecting mutations that arise in response to investigational therapeutic compounds. Successful completion of the proposed research will lead to an effective diagnostic for resistance conferring mutations to the pol gene and permit the application of the technology to other HIV-l genes.

We will develop rapid, cost-effective methods for detecting drug resistance in humanimmunodeficiency virus type l (HIV-l) isolates by analyzing the DNA sequences of patient samples. Thehealth of HIV-l/AIDS patients may be better maintained through lower toxicity and more appropriateantiviral use. All currently approved HIV-l antivirals are targeted against reverse transcriptase (RT).Specific mutations in RT confer resistance to these antivirals. The aim of both phases of this SBIRproposal is to develop DNA chips to detect resistance conferring mutations in RT from clinical samples.In Phase I, we will demonstrate the feasibility of this approach by 1) designing a set of overlapping DNAprobes that corresponds to the most common Ziovudine (AZT) resistance conferring mutation site in RT,2) synthesizing DNA chips containing arrays of these probes using light directed parallel chemicalsynthesis, 3) hybridizing fluorescently labeled target DNA to the chips, and 4) discriminating resistantand sensitive DNA sequences at this site. Completion of these Phase I goals will demonstrate thefeasibility of using DNA chips to detect and monitor mutations in HIV-1 that confer resistance toapproved and investigational therapeutics. In Phase II we will extend the DNA chip design to detect allknown mutations in RT that confer resistance to approved HIV-l antivirals and investigate the sensitivityof the assay to mixtures of viral genotypes. In Phase Ill, we will test the chips in a clinical laboratorysetting and investigate DNA chips to resequence the entire pol gene. This will allow application of thechips to detecting mutations that arise in response to investigational therapeutic compounds. Successfulcompletion of the research will lead to an effective diagnostic for resistance conferring mutations to thepol gene and permit the application of the technology to other HIV-l genes.