RESEARCH PROGRAM

John W. Mellors, M.D.

Professor of Medicine Chief, Division of Infectious Diseases Director, HIV/AIDS Program, UPMC [email protected]

www.pitt.edu/~disease

Basic Research Translational Research Clinical Research Understand Drug Resistance Discover New Drugs Better Treatment and Prevention

Major Projects

• Discovery of novel nucleoside analogs with activity against drug-resistant HIV • • New mechanisms of HIV drug resistance Role of low frequency drug-resistant variants in treatment failure • Mechanisms of HIV persistence on antiretroviral therapy and strategies for eradication • HIV prevention with antiretrovirals

Doctoral Student Training Model

Important Clinically Relevant Question?

Genetic, Virologic, Biochemical, Structural Studies

Past/Current Doctoral Trainees

C. Shi, PhD 1991 1997 Development of recombinant retroviral systems for analysis of HIV-1 drug resistance J. Hammond, PhD U. Parikh, PhD E. Halvas, PhD 1996 2002 1999 2005 2000 Present Structural determinants of HIV resistance to nucleoside analogs Antagonistic evolutionary pathways of HIV-1 resistance to NRTI Role of low frequency drug-resistant variants in antiretroviral treatment failure S. Clark, PhD J. Brehm J. Meeter 2002 Present 2005 Present 2007 Present Genetic, biochemical, and structural mechanisms of NNRTI hypersusceptibility The influence of mutations in the polymerase and RNase H domains on resistance to reverse transcriptase inhibitors Structural basis of nucleoside activity, resistance and toxicity

Jessica Brehm Pre-doctoral Student IDM Department - GSPH And member of Mellors Lab since 2004 BS Biochemistry and Chemistry 2003 College of Charleston South Carolina PEER-REVIEWED PUBLICATION 1. Brehm JH

, Koontz D, Meteer JD, Pithak V, Sluis-Cremer N, Mellors JW. Selection of Mutations in the Connection and RNase H Domains of Human Immunodeficiency Virus Type 1 Reverse Transcriptase that Increase Resistance to 3'-Azido 3'-Dideoxythymidine.

J Virol

2007;81:7852-9

ABSTRACTS Presented at Conferences 2007-08

1. 14th Annual Conference on Retroviruses and Opportunistic Infections (CROI). Feb 2007, Los Angeles, CA. (Oral presentation) 2.

XVI International HIV Drug Resistance Workshop June 2007, Barbados.

3. 15 th Annual CROI conference, February 2008, 4.

Boston, MA (Oral presentation) XVII International HIV Drug Resistance Workshop, June 2008, Sitges, Spain (Oral)

Fellowship

: Pittsburgh AIDS Research Training Fellow (PART) – 2007-08

Jeff Meteer Pre-doctoral Student Molecular Virology and Microbiology Graduate Program MVM BS Biochemistry 2006 St. John Fisher College, Rochester NY Member of Mellors Lab since 2007 PEER-REVIEWED PUBLICATION

Brehm JH, Koontz D,

Meteer JD

, Pithak V, Sluis-Cremer N, Mellors JW. Selection of Mutations in the connection and RNase H domains of human immunodeficiency virus Type 1 reverse transcriptase that Increase resistance to 3’-azido-3’ Dideoxythymidine.

J Virol

2007;81: 7852-9.

ABSTRACT Presented at Conference: Meteer J,

Koontz D, Rapp KL, Detorio M, Ruckstuhl M, Schinazi RF, Mellors JW.

Novel resistance profile of the potent Nucleoside analogue reverse transcriptase Inhibitor 3’-azido-2’,3’-dideoxyguanosine. ORAL PRESENTATION at the XVII International HIV Drug Resistance Workshop. June 2008 Sitges, Spain.

Urvi Parikh, PhD University of Pittsburgh

Assistant Professor of Medicine

Associate Director Microbial Trials Network Lab Doctoral student in MELLORS LAB 1999-2005 PhD – IDM GSPH 2006 BS Biochemistry and Molecular Biology - 1998 Pennsylvania State University Urvi’s Carrer Path after Mellors Lab….(5 publications, 4 first author)

2006 to 2008 Emerging Infectious Disease Research Fellow Centers for Disease Control and Prevention Laboratory of Walid Heneine, PhD, Division of HIV/AIDS Prevention Laboratory Branch.

Conducted in vitro evaluations of nucleoside reverse transcriptase inhibitors for use as a microbicide gel for the prevention of HIV transmission. Also involved In design and implementation of candidate Microbicide safety and efficacy studies in non-human primates.

RECRUITED TO PITT FACULTY IN 2008

To serve as Associate Director of Microbial Trials Network and to enhance ID Division global health Initiatives

Nicolas Sluis-Cremer, PhD University of Pittsburgh Assistant Professor Director, Basic Education ID Division

Jessica Radzio Pre-doctoral Student IDM Department of GSPH Sluis-Cremer Lab since 2007 BS Biology - 2001 MS Life Sciences - 2003 Virginia Polytechnic & State University, Blacksburg, VA PEER-REVIEWED PUBLICATIONS 2007-08 1. Radzio J,

Sluis-Cremer N. Efavirenz accelerates HIV-1 reverse transcriptase ribonuclease H cleavage, leading to diminished zidovudine excision.

Mol Pharmacol

2008;73(2)601-6.

2. Zelina S, Sheen CW,

Radzio J

, Mellors JW and Sluis-Cremer N. Mechanisms by which the G333D Mutation in human immunodeficiency virus type 1 Reverse transcriptase facilitates dual resistance

Protein Sci

2007;16(8) 1728-37.

3. Xia Q,

Radzio J

and Sluis-Cremer N. Probing Non-nucleoside inhibitor-induced active-site distortion in HIV-1 reverse transcriptase by transient kinetic analyses.

Protein Sci

2007(16) (8);1728-37 4. Nissley DV,

Radzio J,

Anderson KS, Sluis Cremer N. Characterization of novel non Nucleoside inhibitor-induced active-site distortion In HIV-1 reverse transcriptase by RT.

Biochem J

2007;404(1) 151-7.

Brian Herman Pre-doctoral Student Molecular Virology and Microbiology Graduate Program BS Biology/Biochemistry Allegheny College 2005 Sluis-Cremer Lab 2007 PEER REVIEWED PUBLICATION:

Roberts A, Deming D, Paddock CD, Cheng A, Yount B, Vogel L,

Herman BD

, Sheahan T, Heise M, Genrich GL, Zaki SR, Baric R, Subbarao K. A Mouse-adapted SARS-coronavirus causes disease and mortality in BALB/cmice.

PLoS Pathog

2007;3(1):e5 T.

PhD Thesis:

Structure-activity-resistance Relationships and biochemical Pharmacology of novel nucleoside and Nucleotide HIV-1 reverse transcriptase inhibitors

Past MD Fellow Trainees

S. Riddler, MD H. Nguyen, MD 1991-1994 1991-1995 Antiretroviral activity of stavudine (2’,3’ didehydro’3’-depxythymiding, d4T) Molecular basis of HIV resistance to NNRTI N. Connolly, MD 2002-2005 Immune based therapies for HIV infection J. McKinnon, MD S. Patel, DO J. Chow, BS 2003-2005 Simplified maintenance antiretroviral therapy: Virologic predictors of outcome 2003-2005 Persistence of multidrug resistant HIV after structured treatment interruptions 2007-Present Role of minor drug-resistant variants in treatment failure

Laboratory Techniques

• • • Experimental design!

Cell culture and virology Antiviral susceptibility/cytotoxicity assays − drug assessments • • Biochemistry of RT and other enzymes Molecular biology − Cloning, PCR, site-specific mutagenesis, DNA sequencing, single genome sequencing • • Structural biology Application of above to human samples

Expectations

• • • • Hard work and commitment to science Weekly meetings with supervisor 2 or 3 first author publications (for Ph.D) Scientific presentations at national meetings

Mellors Lab University of Pittsburgh Division Infections Diseases