1. HIV INFECTION AND THE ACQUIRED IMMUNODEFICIENCY SYNDROME (AIDS)

2. HISTORICAL PERSPECTIVES OF AIDS
1981
1982
1983
1984
1986
Recognition of Pneumocystis carinii pneumonia (PCP) and Kaposi’s sarcoma (KS) in young healthy men in NYC and Los Angeles
GRID to AIDS by CDC
Isolation of Lymphadenopathy-Associated Virus (LAV) by Pasteur Institute (Luc Montagnier)
Isolation of Human T-Lymphotrophic Virus , Type III (HTLV-III) by NCI/NIH (Robert Gallo)
Recommendation of the name Human Immunodeficiency Virus (HIV) by an international subcommittee on virus taxonomy

7. HUMAN IMMUNODEFICIENCY VIRUSES (HIV)
Classification
Retroviridae (family)
Lentivirus (genus)
Characteristics
100 nm in diameter
Genome of 2 single strands of RNA
Nine genes
Reverse transcriptase
RNA-dependent DNA polymerase
Transcribes RNA into DNA

11. GENOME OF HIV Contains 6 regulatory genes Contains 3 structural genes
Env (Envelope glycoproteins)
gp120 and gp41
Gag (Core and matrix proteins)
p55, p40 and p24
Pol (Enzymes)
Reverse transcriptase (p66, p51)
Protease (p11)
Integrase (p32)

13. HUMAN RETROVIRIDAE (EXOGENOUS RETROVIRUSES)
Seven genera
Alpha, Beta, Gamma, Delta, Epsilon, Lenti and Spuma
Deltavirus
Human T-lymphotropic virus, type I (HTLV-1)
Human T-lymphotropic virus, type II (HTLV-II)
Lentivirus
Human immunodeficiency virus, type 1 (HIV-1)
Human immunodeficiency virus, type 2 (HIV-2)

14. CLASSIFICATION OF THE HUMAN IMMUNODEFICIENCY VIRUSES (HIV)
Types
Human immunodeficiency virus, type 1 (HIV-1)
Human immunodeficiency virus, type 2 (HIV-2)
HIV-1 is divided into groups
M (Major)
N (New)
O (Outlier)
Group M is divided into
Subtypes (Clades)
Circulating recombinant forms (CRF)

15. CLASSIFICATION OF HIV

16. ORIGIN OF HUMAN IMMUNODEFICIENCY VIRUSES
Existed as monkey virus in equatorial Africa
HIV-1
Chimpanzee (Pan troglodytes troglodytes)
HIV-2
Sooty Mangabey (Cercocebus atys)
Transition from monkeys to humans
When - Circa 1908
Molecular phylogenetics
How – Hunter theory

19. MECHANISM OF PATHOGENICITY OF HIV
Envelope protein (gp120) of HIV binds with CD-4 receptor on surface of
T-lymphocytes
Macrophages
Dendritic cells
Microglial cells
Coreceptors for attachment of HIV
CCR5 (T-cells, macrophages, dendritic cells, microglial
cells)
CXCR4 (T-cells)

20. MECHANISM OF PATHOGENICITY OF HIV
Early infection
CCR5 coreceptor is used (R5 strains)
Growth equal in monocytes and lymphocytes
Non syncytium-inducing (NSI)
Late infection
CXCR4 coreceptor is used (X4 strains)
Growth in T cells
Syncytium-inducing (SI)
Emergence of X4 strains associated with accelerated decline in CD4 T cells
Cause or consequence?

21. MECHANISM OF PATHOGENICITY OF HIV
Following attachment, virus enters cells and removes protein coat
Viral RNA is transcribed into DNA by
Reverse transcriptase
Viral DNA then integrated into host cell DNA
Integrase
Integrated viral DNA
Referred to as “provirus”
Production of active infection

24. EPIDEMIOLOGY OF HIV INFECTION AND AIDS
Since 1981, 65 million people worldwide have contracted HIV
> 25 million deaths
87% of HIV cases in developing nations
64% in sub-Saharan Africa
23% in southern and Southeast Asia
Since 1981, 1.5 million people in the U.S. have contracted HIV
Approximately 576,000 deaths
In 2009, 56K new cases in the U.S.

27. TRANSMISSION OF HIV INFECTION AND AIDS
Sexual intercourse with infected person
Homosexual (MSM)
Heterosexual
Bisexual
Children born to infected mothers
Perinatal
IV drug addicts sharing contaminated syringes/needles
Transfusion of blood and blood products
Transfusion recipients
Hemophiliacs
Occupational exposure in health-care setting

28. CDC CLASSIFICATION OF HIV INFECTION AND DISEASE IN ADULTS AND ADOLESCENTS
Latest revision in 1993
Clinical Categories A B C
CD4 T Cell Categories (Absolute number or %)
> 500/uL or > 29% of total lymphocytes
200 – 499/uL or 14-28% of total lymphocytes
< 200/uL or < 14% of total lymphocytes

29. CDC CLASSIFICATION SYSTEM
CD4 Cell
Categories
Clinical Categories A
Asymptomatic, Acute HIV, or PGL B
Symptomatic Conditions, not A or C C
AIDS-Indicator Conditions
(1) > 500 cells/µL A1 B1 C1
(2) cells/µL A2 B2 C2
(3) < 200 cells/µL A3 B3 C3

32. CDC CLASSIFICATION SYSTEM (CLINICAL CATEGORY A)
Following initial infection
Asymptomatic
Acute Retroviral Syndrome
Infectious mononucleosis-like or flu-like illness
2 days to 4 weeks following infection
Clinical manifestations
Fever, headache, lethargy, pharyngitis, myalgias, photophobia, lymphadenopathy and a faint maculopapular rash
Resolution within 30 days
Persistent generalized lymphadenopathy (PGL)

35. CDC CLASSIFICATION SYSTEM (CLINICAL CATEGORY B)
Symptomatic conditions not meeting conditions of clinical categories A or C
Herpes zoster (shingles)
Oropharyngeal Candidiasis (thrush)
Candida albicans
Vulvovaginal candidiasis
Bacillary angiomatosis
Bartonella henselae
Peripheral neuropathy
Idiopathic thrombocytopenic purpura (ITP)
Hairy leukoplakia (oral)

40. CDC CLASSIFICATION SYSTEM (CLINICAL CATEGORY C)
Acquired Immunodeficiency Syndrome (AIDS) Defining Conditions
Esophageal Candidiasis
Cryptosporidiosis
Pneumocystis jiroveci (carinii) pneumonia
Tuberculosis (pulmonary or extrapulmonary)
Disseminated Mycobacterium avium complex (MAC) disease
Histoplasmosis (disseminated or extrapulmonary)

41. HIV INFECTION IN ADULTS (CLINICAL CATEGORY C)
Acquired Immunodeficiency Syndrome (AIDS) Defining Conditions
HIV wasting syndrome
Cryptococcal meningitis
Cytomegalovirus retinitis
Cerebral Toxoplasmosis
Progressive multifocal leukoencephalopathy (PML)
JC virus
Kaposi’s sarcoma
Human herpesvirus type 8 (HHV-8)

52. PROGNOSIS AND MONITORING OF HIV TREATMENT AND DISEASE
CD4 T cell count (Immunological response)
Absolute number
Best indicator for patients with counts < 200 cells/uL
Percent
Best indicator for patients with counts > 200 cells/uL
HIV-1 RNA (Viral load) (Virological response)
Discordant immunological and virological responses exist

53. TREATMENT OF HIV INFECTION AND DISEASE
Anti-retroviral drugs do not cure HIV infection or disease
Suppression of virus to undetectable levels
Suppression of virus
Drugs must be taken continuously
Patients remain infectious
Mutation rate in HIV is high and resistance develops
Recommendation for combination therapy
Combination of drugs from two or more classes
Highly Active Anti-Retroviral Therapy (HAART)

54. TREATMENT OF HIV INECTION AND DISEASE
Classes of anti-retroviral drugs
Reverse Transcriptase Inhibitors (RTI’s)
Nucleoside
Nucleotide
Non-nucleoside
Protease Inhibitors (PIs)
Fusion or Entry Inhibitors
Act on gp41 or CCR5 coreceptor
Integrase Inhibitors
Fixed dose combinations
Drugs from two or more classes into a single product

55. TREATMENT OF HIV INECTION AND DISEASE
Reverse transcriptase inhibitors
Nucleoside analog (NARTI, NRTI)
Converted into nucleotide
Incorporated into and stops viral DNA synthesis
Zidovudine (Retrovir)
Nucleotide analog (NtARTI, NtRTI)
Tenofir (Viread)
Non-nucleoside (NNRTI)
Not incorporated into viral DNA
Binds to enzyme and inhibits function
Nevirapine (Viramune)

56. TREATMENT OF HIV INECTION AND DISEASE
Goals of HAART
Suppression of HIV
Decrease viral load
Reduce potential for resistance to anti-viral agents
Immune system reconstitution
Restore CD4 T cell population
Most successful with high baseline CD4 count at HAART initiation
Increase of 50 to 150 cells per year

57. TREATMENT OF HIV INECTION AND DISEASE
HAART negatives
High cost, medication fatigue, adherence to complicated drug regimens, adverse events, names for anti-retroviral drugs
HAART Interruption
Minimize negatives using structured treatment interruption (STI)
6 months of IL-2 without HAART
Safety (unclear) and efficacy (inferior)
HAART associated with
Immune reconstitution syndrome

58. IMMUNE RECONSTITUTION SYNDROME (IRS)
Immune reconstitution inflammatory syndrome (IRIS)
Strong response by recovering immune system to latent or active infections
Risk factors for IRIS following HAART
CD4 percent of < 15%
CD4 count of < 100 cell/uL
High rate of increase of CD 4 count
Most commonly associated with
Pneumocystis pneumonia
Cytomegalovirus disease
Herpes zoster
Mycobacterium avium complex (MAC) disease
Tuberculosis

59. IMMUNE RECONSTITUTION SYNDROME
Important to distinguish between IRIS and clinical failure
Clinical failure
Disease progression with development of OI or malignancy when drugs given for sufficient time
IRIS
Seen within first several weeks of therapy when a latent or active infection is present

60. IMMUNE RECONSTITUTION SYNDROME
Management options
Inflammatory reaction treated with
Steroids
Non-steroidal anti-inflammatory drugs (NSAIDS)
Antimicrobial agents directed at the infectious agent
Antiretroviral therapy
Withhold or continue (?)

61. NAMING ANTI-RETROVIRAL DRUGS
Anti-retroviral drugs have at least 3 names
Abbreviation
Research or chemical name
Generic name
Trade name
Example
Abbreviation (Research/Chemical) AZT
Abbreviation (Generic name) ZDV
Generic Zidovudine
Trade Retrovir

62. MARAVORIC (MVC / SELZENTRY)
First in new class anti-retroviral drug
CCR5 co-receptor antagonist (entry inhibitor)
Indicated for CCR5 tropic HIV-1 showing resistance to multiple anti-retroviral drugs
Black box warning
Hepatotoxicity
Systemic allergic reaction
Pruritic rash, eosinophilia, elevated IgE
FDA approval on August 8, 2007
Requires tropism testing

63. HIV CO-RECEPTOR TROPISM ASSAY
Trofile ™ (Monogram Bioscience)
FDA approval on August 6, 2007
In vitro diagnostic assay
Determines tropism of patient’s HIV
CCR5
CXCR4
D/M (dual / mixed)
Trofile ™ assay
Specimen is EDTA plasma
Viral load of 1,000 copies/mL
TAT of 14 days
Cost $$$$$

66. LABORATORY DIAGNOSIS OF HIV INFECTION
Standard algorithm consists of using two tests for the detection of antibody to HIV-1/2
Screening
Enzyme immunoassay (EIA) or
Enzyme-linked Immunosorbent Assay (ELISA)
High sensitivity
Confirmation
Western blot (WB)
High specificity
Sensitivity is “positivity in disease”
Specificity is “negativity in disease”

67. LABORATORY DIAGNOSIS OF HIV INFECTION (STANDARD ALGORITHM)
Specimens “initially reactive” by EIA / ELISA are retested in duplicate
One or both repeat tests positive, specimens are considered “repeatedly reactive” for antibody
Specimens “repeatedly reactive” by EIA / ELISA then tested by Western Blot (WB) assay
Specimens “reactive” for both EIA / ELISA and WB are considered “positive” for HIV infection
Seroconversion
From infection to antibody

69. LABORATORY DIAGNOSIS OF HIV INFECTION
Rapid detection of HIV-1/2 antibody
OraQuick ® ADVANCE ™ Rapid HIV-1/2 Antibody
OraSure Technologies, Inc., PA
Immunochromatographic assay (ICA)
Analytical time of 25 minutes
Sensitivity of 99.5% and specificity of 99.9%
Specimens of Choice
Whole blood
Fingerstick
Venipuncture (EDTA)
EDTA plasma
Oral fluid (Oral mucosal transudate)

72. CLINICAL USE OF ORAQUICK ® RAPID HIV-1/2 ASSAY
Rapid screening
HCW with potential HIV exposure
Pregnant females with unknown HIV status at time of delivery
New HIV clinic patients
Same day screening
All other patients
Reporting of Results
Negative for HIV-1/2 Antibodies
Preliminary Positive for HIV-1/2 Antibodies. Confirmation by Western Blot testing to follow.

78. LABORATORY DIAGNOSIS OF HIV INFECTION
Detection of HIV Core Antigen (p24)
Serum or CSF
Methods
EIA or ELISA (Non-ICD)
EIA or ELISA (immune complex dissociation)
Positives confirmed by neutralization
Clinical Use
Early diagnosis before antibody response
Monitor effectiveness of therapy
Marker of disease progression

79. LABORATORY DIAGNOSIS OF HIV INFECTION
Detection of proviral DNA
EDTA whole blood
Method
Polymerase chain reaction (PCR)
Clinical Use
Diagnosis of infection in neonates of HIV positive mothers
Early diagnosis before antibody response

80. LABORATORY PROGNOSIS OF HIV INFECTION
Quantitation of HIV-1 RNA (Viral load)
EDTA plasma
Methods
Reverse Transcriptase – PCR (RT-PCR)
Branched chain DNA (bDNA)
Clinical Use
Determination of amount of free virus (Viral load)
Predicting progression and outcome of infection
Assessing efficacy of antiviral therapy

82. LABORATORY DIAGNOSIS OF HIV INFECTION BY ORAL FLUID TESTING
OraQuick ® ADVANCE ™ Rapid HIV-1/2 Antibody Test
Pad on test device used to swab between upper and lower outer gums and cheek
Pad is stored in preservative vial and sent for ICA testing
Advantages
Reduces occupational exposure
Patient appeal

84. LABORATORY DIAGNOSIS OF HIV INFECTION BY URINE TESTING
Calypte HIV-1 Urine EIA (Calypte Biomedical, Berkeley, CA)
FDA approval for EIA (1996) and Western Blot (1998)
Sensitivity and specificity
Lower compared to blood and oral fluid
Question
IgG in urine
Calypte ® Aware ™ HIV-1/2 Urine Rapid Test
Available outside US
Advantages
Reduces occupational exposure
Patient appeal

85. THE IMMUNOLOGY OF HIV INFECTION
Interactions between HIV and human immune system are extremely complex
HIV subverts immune system by
Infecting CD4 T cells and inducing quantitative and qualitative dysfunction
Hyperactivating B cells with resulting hypergammaglobulinemia
Inducing cytokine system to own replicative advantage
There are no known correlates of protective immunity

86. MECHANISMS OF CD4 T-CELL DEPLETION
Direct killing of infected T cells
Increased rate of apoptosis in infected T cells
Molecule associated with apoptosis (PD-1) is over-expressed in chronic viremia
Syncytia formation
Fusion of infected and non-infected T cells
Killing of infected CD4 cells by CD8 cells

87. KILLING OF INFECTED CD4 CELLS BY CD8 CELLS – ALTERNATIVE VIEW
Mechanism that keeps HIV in check in long term non-progressors (LTNPs)
Long term non-progressors (LTNPs)
Carry the virus but do not get AIDS
Have 20 times more CD8 T cells than progressors
Function of CD8 T cell surplus
Up-regulate production (2X rate of progressors) of 2 killer proteins
Perforin
Granzyme B

89. IMMUNE DYSFUNCTION DURING HIV INFECTION - SUMMARY
HIV infection is multifactorial process capable of disarming immune system by direct and indirect mechanisms
Certain chemokine receptors function as necessary coreceptors for entry of HIV into cells
Central Paradox
Progression of HIV disease in setting of vigorous immune response
Lack of correlates of protective immunity are major obstacle to immunotherapy and vaccine development