RETROVIROLOGY ONCONOVIRUSES & LENTIVIRUSES Retrovirus Family / Taxonomy

1. RETROVIROLOGY ONCONOVIRUSES & LENTIVIRUSES Retrovirus Family / Taxonomy Retrovirus Structure & Morphology Retrovirus Genomic Organization Retrovirus Replication Cycle HIV-1 EPIDEMIOLOGY & THERAPY

2. FAMILY: RETROVIRIDAE (SUBFAMILY) GENUS PROTOTYPE DISEASE Oncovirinae Alpharetrovirus ALV, RSV Lymphoma, (Avian type C) Sarcoma Betaretrovirus MMTV Mammary Carcinoma, (type B) Lymphoma (type D)MPMV, SMRV, SRV-1 None;SAIDS Gammaretrovirus MLV, FeLV, GALV Leukemia, (type C) Lymphoma Deltaretrovirus HTLV-1, HTLV-2, STLV, BLVLeukemia, Lymphoma (type C)HTLV-3, HTLV-4 Lentivirinae Lentivirus HIV-1, HIV-2, SIV, FIV AIDS, Neurol. Dis. EIAV, CAEV, MVVPneumonia Spumavirinae Spumavirus HSRV, SFV, FeSFV, BFV None known

3. RETROVIRUS MORPHOLOGY RT SIZE INTRACYTOPLASMIC BUDDING MATURE VIRION RETROVIRUS CATION (nm) A-TYPE PARTICLE MORPHOLOGY CORE ( ICA ) ONCOVIRUS 80-100 Alpha/Gamma Mn+2- Crescent NC Icosahedral Type C Betaretrovirus Type B Mg+2+ ICA budding Icosahedral Type D Mg+2+ ICA budding Cylindrical/Conical Deltaretrovirus Mg+2- Crescent NC Icosahedral Type C LENTIVIRUS Mg+2 100-120- Crescent NC Cylindrical/Conical SPUMAVIRUS Mg+2 100-140 - ICA-like budding Icosahedral

4. LENTIVIRUS: LENTIVIRUS: SPUMAVIRUS ONCOVIRUS LENTIVIRUS FIV FeLV FeSFV Budding VIRUS MATURE VIRUS BUDDING VIRUS MATURE VIRUS HIV SIV MVV Yamamoto JK, et al. AJVR 49:1246-1258, 1988

5. LENTIVIRINAE Caprine Arthritis-Encephalitis Virus (1980) arthritis, encephalitis Maedi-Visna Virus (1953) pneumonia, encephalitis Bovine Immunodeficiency Virus (1972) lymphocytosis Feline Immunodeficiency Virus (1987)FAIDS: encephalitis, anemia Human Immunodeficiency Virus (1983)AIDS: arthritis, encephalitis, anemia Equine Infectious Anemia Virus (1905) fever, anemia Simian Immunodeficiency Virus (1985)SAIDS: encephalitis, anemia

6. HTLV & HIV VIROLOGY HTLV HIV MORPHOLOGY Oncovirus (deltaretrov.)/ no A particle Lentivirus GENOME SIZE (kb) 8.5 (lacks accessory genes) 9.8 GENETIC VARIATION 90-99% w/in geographic location Moderately high w/ in person (5%) Low between dif. Isolates High in & between subtypes VIRAL RECEPTOR GLUT1 (glucose transporter) CD4; CXCR4 (T cell); CCR5 (Mac) on many species (xenotropic) CELL TROPISM T cells ( CD4 / HTLV-1 ) T cells ( CD4) T cells ( CD8 / HTLV-2 ) Macrophages, Glial cells, B cells; Many cell types & Endothelial cells TRANSFORMATION T cells (Tax – viral promoter) None VIRAL REPLICATION Very low to negligible titers High titers TRANSMISSION Cell-associated; Milk; Sex (male to female) Body fluids; Cells; Blood; Blood transfusion (IV drug user, IDU) Milk; Sex; IDU. PATHOGENESIS VERY SLOW (>20 years) HIV-2 slow; HIV-1: Pediatric - acute CD4 / CD8 normal or Adult AIDS - acute to slow CD4 high w/ malignancy CD4 / CD8 & CD4 decrease DISEASES Adult T-cell Leukemia (CD4)-HTLV-1 CNS diseases; AIDS Tropical Spastic Paraparesis- HTLV-1 Kaposi’s sarcoma T-cell variant hairy-cell leukemia (CD8)HTLV-2

7. ACUTE ATL Adult T-cell Leukemia Skin lesions due to infiltration of leukemia cells (skin and/or lung infiltration) Polylobulated nucleus of two CD4+ T leukemic cells

8. GLOBAL DISTRIBUTION OF HTLV INFECTIONS STLV-1 HTLV-1 STLV-2 HTLV-2 STLV-3 HTLV-3 ? HTLV-4 Cameroon HTLV-3 (2005; 4 cases) HTLV-4 (2005; 1 case) (Discovered 1980; 15-20M cases in 2009) (Discovered 1982)

9. 33 MILLION LIVING WITH HIV-1 IN 2007 Haiti 1966 Subtype B HIV origin 1884-1924 Origin of Group M: Kinsasha 1959 (1959, subtype D; 1960, subtype A) Brazzaville 1983

10. Figure 1. RT PR IN SU TM RETROVIRUS STRUCTURE VIRAL CORE & CORE ASSOC. PROTEINS VIRAL ENZYMES Reverse Protease Transcriptase Integrase Matrix Protein Capsid Nucleoprotein p6 NC MA CA ACCESSORY PROTEINS (HIV) Vif Vpr Nef Cellular Plasma Membrane Surface Transmembrane Envelope Viral RNA Genome VIRAL ENVELOPE PROTEINS HIV, SIV, FIV

11. y TAR gag-pol FSR Viral mRNA RRE PB PAS SA SA SD PB = Primer Binding site FSR = Frame Shift Region SD = Splice Donor site SA = Splice Acceptor site 5’ CAP gag pro pol env AAA 3’ REGULATORY / ACCESSORY PROTEINS (genes) Tat - binds to TAR (trans-activating response element) on viral transcripts to enhance transcribing polymerase activity. Rev - binds to RRE (Rev response element) on viral transcripts to export unspliced mRNA out of nucleus. Rev rich in NES (nuclear export signal) which allows it to shuttle between nucleus & cytoplasm via interaction with nucleoporin-like protein (hRip/Rab) on nuclear pore. Nef - 1) Down regulate CD4 & MHC-I which affects immune response 2) Facilitate routing of CD4 from cell surface & golgi aparatus to lysosome; causes increase in Env incorporation into virion & promote particle release. 3) Enhance viral infectivity by binding to Src-family proteins & regulating their tyrosine kinase activities. Vif - 1) Functions before or during viral DNA synthesis hence affect viral production (block APOBEC). 2) Production of highly infectious mature virions by affecting assembly and/or maturation. Vpr - 1) Mediate transport of nucleoprotein complexes (RT, IN, MA, vRNA, partially reverse-transcribed DNA) to nucleus. 2) Has NLS that directs transport even in absence of mitotic nuclear envelop breakdown & therefore important for nuclear localization in nondividing cells (macrophages) 3) Cause G2 cell cycle arrests & kill T cells by apoptosis. 4) Influence mutation rates during viral DNA synthesis Vpu - 1) Down regulate CD4 & MHC-1 expression & affects immune response 2) Increase virion release

12. COMPARISON OF RETROVIRAL GENOMES ( kilobases ) 0 1 2 3 4 5 6 7 8 9 10

14. RETROVIRUS REPLICATION CYCLE 1. Virus attachment to specific cell-surface receptor. 2. Viral core penetration into cell. 3. Reverse transcription within core structure (RNA to DNA). 4. Viral DNA transit into nucleus. 5. Viral DNA integration into cell DNA to become provirus. 6. Viral RNA synthesis by cell RNA polymerase. 7. Processing viral transcripts to genome and mRNAs. 8. Viral protein synthesis. 9. Assembly and budding of virions. 10. Processing of capsid proteins.

15. Reverse Transcriptase Heterodimer - p66 & p51

16. VIRAL DNA SYNTHESIS ( Stage 1 - Cell Cytoplasm )

17. VIRAL DNA SYNTHESIS ( Stage 2 - Cell Nucleus ) Vpr –( RT / IN / MA / vDNA ) Vpr Transport to Nucleus Stage 2 Cell Nucleus ACTIVATION SIGNAL INTEGRASE

18. 0 1 2 3 4 5 6 7 8 9 10 Tat

19. 0 1 2 3 4 5 6 7 8 9 10 in

20. Viral mRNA transit to cytoplasm Vpr Rev Enhance release of virion Nef & Vpu

21. A CURE FOR HIV-1 USING HIGHLY ACTIVE ANTIRETROVIRAL THERAPY (HAART) IN COMBINATION WITH TOTAL BODY IRRADIATION (TBI) AND PERIPHERAL-BLOOD STEM-CELL TRANSPLANTATION (PSCT) WITHHOMOZYGOUS CCR5-Δ32/Δ32 STEM CELLS HAART to lower HIV-1 load (circulation) and to prevent infection of engrafting immune cells TBI to kill infected immune cells PSCT with CCRC-∆32/∆32 SCs to engraft resistant immune cells HIV-1 CURE ???????

22. QUESTIONS ( RETROVIROLOGY ) : • Name the three Retroviridae subfamilies and their major disease manifestations. • What is a provirus? • Can a retrovirus replicate in a cell using cellular reverse transcriptase (RT) and • without the use of viral RT? Describe how RT functions in HIV replication? • Is immune activation important in HIV replication? • What are the regulatory genes and their function(s)? NEXT SECTION IS ON : HIV-1 THERAPY

23. HIV-1 AND AIDS HIV THERAPY HIV EPIDEMIOLOGY & PHYLOGENY HIV PATHOGENESIS & HIV VACCINE

24. THERAPY • Anti-gp120 Abs • Anti-gp41 Abs • Anti-CD4 Abs • Soluble CD4 • 1A) CCR5 Inhibitor • 1B) Fusion Inhibitor • AZT, ddI, 3TC, dT4 • 5) Integrase Inhibitor • 8A) Protease Inhibitors • Interferon-Alpha • 9-10) DIBA • (Zinc-finger Inhibitor) HIV REPLICATION CYCLE STAGES TARGETED FOR THERAPY 1Virus adsorption to CD4 receptor Fusion Inhibitor 1B HIV gp41 fuses viral & cellular membranes CCR5 Inhibitor 1ACD4 binds to gp120 then CCR5 binds to gp120 CD4 CCR5 Nucleoside Analogs Non-nucleoside RT Inhibitors Protease Inhibitors Integrase Inhibitor

25. FDA APPROVED ANTI-HIV-1 DRUGS GENERIC NAME/ TRADE NAME FDA NOMENCLATURE APPROVAL (Company) NUCLEOSIDE ANALOGS ( NRTI ) : Zidovudine / ZDV / AZT Retrovir / AZT Mar. 19, 1987 (Glaxo Wellcome) Retrovir Didanosine / ddI Videx Oct. 9, 1991 (Bristol-Myers Squibb) Combinavir (Sept. 27, 1997) Х Zalcitabine / ddC Hivid June 19, 1992 Trizivir ( Nov. 14, 2000 ) ( Hoffmann-La Roche) Stavudine / d4T Zerit June 24, 1994 FDA APPROVED ANTI-HIV-1 DRUGS (Bristol-Myers Squibb) Lamivudine / 3TC Epivir Nov. 17, 1995 Epizicom (Aug. 2, 2004) (Glaxo Wellcome) Dec. 17, 1998 Ziagen Abacavir / ABC (Glaxo SmithKline) Oct. 26, 2001 Viread Tenofovir / TDF / PMPA (Gilead Sciences) Truvada (Aug. 2, 2004) Emtricitabine / FTC Emtriva July 2, 2003 (Gilead Sciences) NON-NUCLEOSIDE RT INHIBITORS ( NNRTI ) : Atripla ( July 12, 2006 ) Nevirapine / NVP Viramune June 24, 1996 (Roxane Laboratories) Delavirdine / DLV Rescriptor April 4, 1997 (Pharmacia & Upjohn) Efavirenz / EFV Sustiva Sept. 17, 1998 HIV-2 (DuPont Pharm. Co., Bristol-Myers Squibb) Effective Low effect No effect Etravirine / ETR Intelence Jan. 18, 2008 X X X X X X X (Tibotec Therapuetics, J&J) GENERIC NAME/ TRADE NAME FDA NOMENCLATURE APPROVAL (Company) PROTEASE INHIBITORS ( PI ) : Saquinavir / SQV-(HGC) Inverase Dec. 6, 1995 (Roche Pharmaceuticals) Ritonavir/ ABT-538/ RTV Norvir Mar. 1, 1996 (Abbott Laboratories) Indinavir / IDV Crixivan Mar. 13, 1996 (Merck & Co.) Nelfinavir / NFV Viracept Mar. 14, 1997 (Agouron Pharmaceuticals) Х Fortovase Saquinavir / SQV-(SGC) Nov. 7, 1997 (Roche Pharmaceuticals) Х April 15, 1999 Agenerase Amprenavir / APV / AMP (Glaxo SmithKline) Ritonavir+Lopinavir / RTV+LPV Kaletra Sept. 15, 2000 (Abbott Laboratories) Reyataz June 20, 2003 Atazanavir / ATV / ATZ (Bristol-Myers Squibb) Oct. 20, 2003 Lexiva Fosamprenavir / FPV (Glaxo SmithKline) Tipranavir / TPV Aptivus June 22, 2005 (Boehringer Ingelheim) Darunavir / DRV Prezista June 23, 2006 (Tibotec Therapeutics, J&J) FUSION INHIBITOR: Enfuvirtide / T20 Fuzeon Mar. 13, 2003 (Roche + Trimeris) CCR5 INHIBITOR: Maraviroc / MVC Selzentry Aug. 6, 2007 (Pfizer Inc.) INTEGRASE INHIBTOR: Raltegravir / RAL / MK-0518 Isentress Oct. 12, 2007 (Merck & Co.)

26. Cross-resistance occurs with many drugs that have similar mechanism(s) of drug action (i.e., within drug class). Highest frequency Lowest frequency of cross-resistance of cross-resistance NNRTIs PIs NRTIs > > Time for resistant strains or mutants (also cross-resistant strains) to develop: NNRTIs PIs NRTIs days - weeks 2-3 months 6 - 12 months < < DURATION OF DRUG EFFICACY: DEVELOPMENT OF RESISTANT MUTANTS

27. ( PMPA ) ( ABC ) ( 3TC ) ( d4T ) deoxyguanosine

28. INHIBITION OF HIV Reverse Transcriptase Heterodimer - p66 & p51 Functional Activities & Sites Polymerase – RdDp & DdDp RNase H AZT ( NRTI ) Nevirapine (NNRTI)

29. HIV PROTEASE INHIBITORS RT IN

31. F F H3C NH N N N N H3C CH3 Maraviroc or Selzentry ( CCR5 Inhibitor ) gp41 Blocks CCR5 co-receptor binding to gp120 NEW ANTIRETROVIRAL DRUGS FUNCTION AT CELL SURFACE 36 amino acid peptide C204 H301 O64 Enfuvirtide or Fuzeon or T-20 ( Fusion Inhibitor ) CD4 CCR5 Prevents viral gp41 from fusing the viral membrane to the cell plasma membrane Blocks CCR5 co-receptor binding to gp120

32. NRTI PMPA C19H30N5O10P· C4H4O4 ($5,733 / year) CH3CO-Tyr-Thr-Ser-Leu-Ile-His-Ser-Leu-Ile-Glu-Glu-Ser-Gln-Asn-Gln-Gln-Glu-Lys -Asn-Glu-Gln-Glu-Leu-Leu-Glu-Leu-Asp-Lys-Trp-Ala-Ser-Leu-Trp-Asn-Trp-Phe-NH2 PI Ritonavir C37H48N6O5S2 ($13,128 / year) Fusion-I Enfuvirtide C204H301N51O64 ($23,919 / year) SC injection Diester hydrolysis in intestine lumin & plasma CCR5-I Maraviroc C29H41F2N5O ($10,585 / year) Tenofovir diphosphate PMPAPP in cell INI Raltegravir C20H20FKN6O5 ($9,850 / year) NRTI 3TC C8H11N3O3S ($3,801 / year) NNRTI Nevarapine C15H14N4O ($4,796 / year) DRUG STRUCTURES & USA PRICING in 2007

33. CLINICAL OUTCOME: Delay in symptoms ( i.e., delay in AIDS manifestations ) Extended survival time - Reduction of HIV-related morbidity & mortality IMMUNOLOGIC OUTCOME: Preserve & restore immune functions ( increase CD4 counts & CD4 / CD8) Enhancement of protective immune functions VIROLOGIC OUTCOME: Decrease viral load for a long duration PBMC viral load ( HIV RNA, proviral DNA ) Plasma viral load ( HIV RNA by PCR; HIV p24 level ) Delay in viral evolution ( i.e., mutant emergence ) - Important in determining when to switch therapeutic regimen QUALITY OF LIFE - SAFETY & TOLERANCE: Improve quality of life - minimal-to-no drug toxicity and clinical AIDS Terotogenic effect (especially multiple drug combination) PARAMETERS FOR ASSESSING THERAPEUTIC EFFICACY GOALS OF HIV THERAPY X

34. RESISTANCE TO DRUG ( 70% ): Cause of high mutation is due to the fact that the REVERSE TRANSCRIPTASE is error-prone (i.e., makes a lot of mistakes & these mistakes lead to new mutants). LOW DRUG CONCENTRATIONS AT THE VIRAL REPLICATION SITE (>25%): 1) Low Drug Potency due to Patient Factors: a) Noncompliance b) Interpatient variability 2) Low Drug Potency due to Pharmacologic Factors: a) Drug-drug interactions b) Inefficient drug absorption c) Accelerated metabolism PROGRESSIVE DECLINE OF THE IMMUNE SYSTEM: CD4+ T cell loss CD8+ T cell loss Immune Network Destroyed Other immune cell loss MAJOR REASONS FOR TREATMENT FAILURE IN HIV PATIENTS

35. QUESTIONS ( HIV THERAPY ) : • What is HAART or ART? • An unsuccessful treatment with a protease inhibitor will often cause • the subsequent therapy with another protease inhibitor to be less • effective. Why? • Would a therapy with nucleoside analogs be more successful • in this patient?

36. ANTIRETROVIRAL(ARV)TREATMENT COVERAGE AS OF DECEMBER 2007 Estimated no. of Estimated no. of Geographical Region people receiving people needing ARV therapy ARV therapy ARV therapy coverage Sub-Saharan Africa 2.1 M 7 M 30% Latin America & Caribbean 390,000 630,000 62% East, South & SE Asia 420,000 1.7 M 25% Europe, Central Asia 54,000 320,000 17% Middle east, North Africa 7,000 100,000 7% Total 2.9 M 9.7 M 31% HIV-1 AND AIDS HIV EPIDEMIOLOGY & PHYLOGENY HIV PATHOGENESIS

37. HAART/TBI/PSCT WITH CCR5-Δ32/Δ32 STEM CELLS AS A CURE FOR HIV-1 ? HAART Chemo-ablation (4X) TBI (low dose, 1X) PSCT (2.3x106 SC/kg) 11mo AML relapse Chemo-ablation (1X) TBI (200 cGy, 1X) PSCT(2.1x106 SC/kg) 20mo - free of AML HIV-1 CURE ??????? HIV-1+ MAN with Acute Myeloid Leukemia (AML) (40 years of age; 415 CD4+ T cells): Allogeneic Stem Cells - important for AML CCR5-deletion Tissue-matched HAART until few weeks post-SCT 600mg/d Efavirenz (EFV; NNRTI) 200mg/d Emtricitabine (ETC; NRTI) 300mg/d Tonofovir (PMPA; NRTI) Anti-rejection drugs 1st: rabbit anti-thymocyte globulin 1st & 2nd: cyclosporine 1st & 2nd: mycophenolatemofetil No HIV-1 RNA in blood & BM HIV-1 DNA+ in blood 3.5 YEARS No HIV-1 RNA in blood and tissues Proviral DNA+ in tissues • 2.9% of his HIV-1 species was X4- or dual-tropic • Treat leukemia w/o increasing HIV-1 infection • HAART  CHEMO-ABLATION and/or TBI  PSCT • Control or treat HIV-1 infection • Stem cells from HLA-matched subject • Stem cells with homozygous CCR5 deletion (Δ32bp) • 85-90% of HIV-1 uses CCR5 coreceptor • 10-15% of HIV-1 uses CXCR4 coreceptor • 1% of European population has CCR5-Δ32/Δ32

38. 2008 850,000 59,000 850,000 240,000 310,000 1.5 million 2.0 million 3.8 million 1.4 million 22.4 million GLOBAL TOTAL ALIVE 33.4 million

39. 2008 25,000 13,000 12,000 77,000 59,000 2,000 20,000 87,000 270,000 1.4 million GLOBAL TOTAL DEATHS 2.0 million / Year (5,495 DEATH / DAY)

40. 2008 55,000 20,000 170,000 3,900 30,000 35,000 110,000 East Asia 75,000 280,000 1.9 million GLOBAL TOTAL NEWLY INFECTED 2.7 million / Year (7,418 NEWLY INFECTED / DAY)

41. Reported AIDS Cases (2007) Cumulative*Reported 2007 State # of Cases % of TotalRate/100,000** . 1. New York 181,461 18 % 24.9 (2nd) 2. California 148,949 15 % 13.5 (11th) 3. Florida 109,524 11 % 21.7 (4th) 4. Texas 72,828 7 % 12.4 (14th) 5. New Jersey 50,694 5 % 13.4 (12th) “Top 5 Total” 563,456 56 % TOTAL U.S. 998,255 100% 12.4 *Cumulative data are through 2007. Lowest cumulative number of cases in North Dakota with 153. **Reported AIDS cases in annual rates (per 100,000 population) in 2007: (1) District of Columbia 148.1; (3) Maryland 24.8; (5) Louisiana 20.5; (6) Delaware 19.8; (7) Georgia 19.7; (8) South Carolina 16.8; (9) Connecticut 15.1 Lowest rate in Vermont with 1.0.

42. Persons Newly Diagnosed with HIV-1 U.S.A. in 2007 and Florida in 2008* *Source: CDC, HIV/AIDS Surveillance Report, 2007; Estimate for Florida. M:F ratio: U.S.A., 3.2:1 Fla., 2.1:1

43. 54 1 2 12 2 5 7 2 32 2 43 1 1 0 287 19 4 0 4 8 1 N = 4,613 4 4 8 0 0 4 14 1 0 78 14 1 3 0 45 47 PTP Cases 0 to 10 11 to 50 51 to 100 > 101 13 8 5 31 16 423 29 32 36 71 161 375 13 18 2 1 67 6 2 19 8 0 3 460 42 12 31 698 9 1312 PERSONS TESTED NEWLY POSITIVE (PTP) FOR HIV IN 2008 90,968 Estimated Living HIV/AIDS Cases in Florida (2008) or 11% of Total USA Cases ( 822,055 Estimated Living HIV/AIDS Cases in USA, 2008 ) Univ. of Florida Florida State Univ. Univ. of Central Florida Univ. of South Florida H. Lee Moffitt Cancer Center Saint Lucie HIV/AIDS has spread out from 6 major urban epicenters into smaller cities, suburban and rural areas. 9

44. SIVcolCGU1 HIV-1 Group O SIVCPZ HIV-1 Group N HIV-1 Group M SIVSYK173 SIVMND2 SIVGSN99CM166 SIVRCMNg411 HIV-2 Group A SIVHOEST HIV-2 Group F HIV-2 Group G SIVSUN HIV-2 Group B SIVMND1 SIVAGM SIVSMM HIV ORIGIN LINKED TO SIV

45. HIV-1 GROUPS & G LBV271 HIV-1 SUBTYPES H G98 BUK3A 92RU131 V1536 ANT70 VAU CA13 92UG975 MPV5180 V1557 CAR4067 J SE7022 V1686 SE7887 CARSAS O A CA9 KENYA 92UG0378 92W0205 M 92BR025 U455 DJ373A C UC268A2 K 93MW965 CPZgab CAR4026 SIV SE364A 2UG021 NDK YBF-160 YBF-30 N MN JRFL Z2Z6 CPZant SF2 D NY5 ELI 93BR029 ALA1 M = Major Group N = Non-M, Non-O Group O = Outlier Group NL34 BZ163A BZ126A B F 93BR020 Former subtypes E (Ae) and I (A, G, H, K) were found to be subtype recombinants.

46. HIV-2 B, a, c, d, f, o HIV-2 B, a, d, o B, Ae B HIV-2 C, a, b, Ae B B C, Ae, b C b f, g, h, j, k recombinants Ae,b C, a, d B B As much as 4 subtype recombinants have been observe in a single isolate (former subtype I, A/G/H/K). 10-35% of circulating forms are recombinants. Recombinations are more common in gag & env. A, D,Ae GLOBAL DISTRIBUTION OF HIV-1 SUBTYPES 90% of Global Infections B C A Ae Ag d B, a, Ab, c, d, g, h B B B, C B, c,Ae, Bc C C, Ae A B HIV-2 A, O, Ag,d,g, h C B B, f,Bf, c B C B, Ae

47. SEXUAL CONTACT HIV enters via mucosal linings (61,461 cases/yr, 60%, USA 2007). ( vagina, vulva, penis, rectum, rarely via mouth ) Mucosal dendritic cells (MDC) & macrophages are first to be infected. Infected MDC & replicated HIV travel to germinal centers of the lymphoid organs where FDCs & T cells become exposed. RECEIPT OF INFECTED BLOOD, BLOOD PRODUCTS, OR TISSUES ContaminatedneedlessharedbyIV drug users (10,430/yr, 10%, USA 2007). Use of infected blood, blood products, or tissues (374/yr, 0.4%, USA 2007). 40 cases in 14 million blood units per year in USA. Since 1989, 4 cases infected with HIV-antibody negative blood. New HIV-p24 capture test detects HIV-1 earlier by 1 week. MOTHER-TO-CHILD TRANSMISSION (610cases/yr,0.6%,USA2007) 75-93% of cases occur in utero or during childbirth. Transmission also by nursing infected breast milk. Breast feeding strongly not recommended for HIV-positive mothers. ART during pregnancy decreases infection of the fetus and newborns. HIV-1 TRANSMISSION (76% ♂&24% ♀, 101,614casesinUSA2007)

48. SEXUAL TRANSMISSION HETEROSEXUAL TRANSMISSION: 2X - 5X Risk: male-to-female >> female-to-male HOMOSEXUAL TRANSMISSION: Receptive Partner >> Active Partner [ Infected insertive partner - to - Uninfected receptive partner > > Uninfected insertive partner - to - Infected receptive partner ]

49. HIV TRANSMISSION RATE POST-SINGLE EXPOSURE vs % TOTAL WITH CORRESPONDING EXPOSURE (GLOBAL vs USA) LIKELIHOOD OF INFECTION GLOBAL USA EXPOSURE TYPE AFTER SINGLE EXPOSURE (%) TOTAL (%)* TOTAL (%)* Sexual Transmission 0.01 - 1.0 70 - 80 78 Receptive vaginal 0.01 - 0.32 60 - 70 19 MSM 1.0 5 - 10 46 MSM + IDU NA NA 5 Contaminated Needle 0.5 - 1.0 5 - 10 18 ( IV drug users, IDU ) Maternal Transmission Pregnancy/delivery 12 - 50 5 - 10 1.1 Breast milk 12 NA NA (Continuous breast feeding) Blood (Product) Transfusion >90 3 - 5 1.2** Health Care Workers 0.1 - 1.0 <0.01 <0.005 (needlestick, etc.) * Global cumulative data were before 2000. USA cumulative data were through end of 2007. ** USA data include infection from contaminated blood, blood products, and tissues.

50. HIV VACCINES IN CLINICAL TRIALS Preclinical Testing – In vitro & In vivo animal trials Clinical Phase I - Dose-escalation, safety, & toxicity tests Clinical Phase II - Expanded safety & dose optimization & some immunogenicity Clinical Phase III - Efficacy trials HIV VACCINE CANDIDATES IN CLINICAL TRIAL HIV SUBTYPE CLINICAL VACCINE(S) (other / env) SPONSOR(S) PHASE LOCATION OF TRIAL(S) rgp120 BB VaxGen III USA Adenovirus 5-gag/pol/nef B Merck, HVTN IIb USA, Canada, Peru, Haiti, Porto Rico, Dominican Republic, Australia ALVAC (vCP gag/pol/env); rgp120 B / Be Aventis Pasteur, VaxGen III Thailand Lipopeptide antigens (Gag, Pol, Nef) B ANRS II France AAV-HIV (gag/PR/ΔRT) C IAVI, Targeted Genetics II South Africa, Uganda, Zambia DNA-HIV (B gag/pol/nef, ABC env); B / ABC HVTN, Vical II USA, Brazil, South Africa (Haiti, Jamaica) ADV5-HIV (B gag/pol, ABC env) DNA-HIV; Fowlpox (gag/po/tat/rev/env) A / Ae UNSW I-II Thailand DNA-HIV (AB gag, B RT/rev, ABC env) AB/ABCe MUCHS, SMI, USMHRP, I-II Tanzania MVA-HIV (A gag, B pol, E env) Karolinska Institute, Vecura Thirty other vaccine trials ABCe Many Companies I USA, France, Thailand, Brazil, Haiti, etc VaxGen (Brisbane,CA); Aventis Pasteur (Lyon, France); HVTN (HIV Vaccine Trials Network, DAIDS); ANRS (National Agency for AIDS Research, France); IAVI (Intl AIDS Vaccine Initiative); UNSW (University of New South Wales); MUCHS (Muhimbili University College of Health Sciences); SML (Swedish Institute of Infectious Dis Control); USMHRP (US Military HIV Research Program); ALVAC (canarypox vector); ADV5 (Adenovirus-5 vector); Lipopeptide antigens (LP5, LP6); AAV (Adeno-associated virus vector); DNA (plasmid DNA vaccine); MVA (modified vaccinia Ankara vector) FAILED TRIALS FAILED TRIALS