New Approaches to combat the HIV infection.

1. Dept. of Pharmaceutical chemistry
L. M. College of Pharmacy
Navrangpura, Ahmedabad-09.
Hardik Mistry
1

2. • Viruses are small infectious agents consisting essentially of nucleic acid
(either RNA or DNA) enclosed in a protein coat called capsid.
• The coat plus the nucleic acid core is termed as the nucleocapsid.
• Some viruses have, in addition, a lipoprotein envelope, which may contain
antigenic viral glycoproteins, as well as host phospholipids acquired when
the virus nucleocapsid buds through the nuclear membrane or plasma
membrane of the host cell.
• Certain viruses also contain enzymes that initiate their replication in the host
cell.
• The whole infective particle is termed as a virion. In different types of
viruses the genome may be double or single stranded.
2

3. • Viruses are intracellular parasites with no metabolic machinery of their own.
• In order to replicate they have to attach to and enter a living host cell-
animal, plant or bacterial and use its metabolic processes.
• The binding sites on the virus are polypeptides on the envelope or capsid.
• The receptors on the host cell, to which the virus attaches, are normal
membrane constituents like receptors for cytokines, neurotransmitters or
hormones, ion channels, integral membrane glycoproteins, etc.
Virus function and life historyVirus function and life history
3

4. • Viral DNA enters in the host cell nucleus, transcription of this viral DNA
into mRNA by host cell RNA polymerase followed by translation of the
mRNA into virus-specific proteins.
• Some of these proteins are enzymes that synthesize more viral DNA as
well as proteins of the coat and envelope.
• After assembly of coat proteins around the viral DNA, complete virions
are released by budding or after cell lysis.
Condit, R. C.: Principles of virology. In Knipe, D. M.; Howley, P. M. (eds): Fundamental Virology, Lipincott
Williams & Wilkins, 4th ed., 2001, pp. 19.
Replication in DNA virusesReplication in DNA viruses
4

5. • Enzymes in the virion synthesize its mRNA or the viral RNA serves as its
own mRNA.
• This is translated into various enzymes, including RNA polymerase (which
directs the synthesis of more viral RNA) and also into structural proteins of the
virion.
• Assembly and release of virions occurs as explained above.
Replication in RNA virusesReplication in RNA viruses
5

6. • The virion in retroviruses contains a reverse transcriptase (virus RNA-
dependent DNA polymerase), which makes a DNA copy of the viral RNA.
• This DNA copy is integrated into the genome of the host cell and it is
then termed a provirus.
• The proviral DNA is transcribed into both new genomic RNA and
mRNA for translation into viral proteins.
• The completed viruses are released by budding and many can replicate
without killing the host cell.
Replication in retrovirusesReplication in retroviruses
6

7.  HIV : ‘Human Immunodeficiency Virus‘ Member of the lentivirinae (lenti,
meaning “slow”) subfamily of retroviruses.
 Only three retroviruses are known to infect humans.
 HTLV I (Human T Leukemia Virus I)
 HTLV II (Human T Leukemia Virus II)
 HTLV III (Human T Lymphotropic Virus)
• HTLV III was confirmed to be the etiological virus for the AIDS
 Types of HIV:
• HIV-1: 3 groups - M (main) (9 subtypes : A-D,H-F,J,K),
- N (new or non-M, non-O),
- O (outlier).
• HIV-2: 6 distinct phylogenetic lineages – Subtypes/clades: A-F.
Rang, H. P.; Dale, M. M.; Ritter, J. M.; Moore, P. K., Antiviral Drugs., Text Book of Pharmacology, Elsevier
Science, 5th ed., 2003, pp. 657
Introduction of HIVIntroduction of HIV
7

8.  HIV-1 subtype B is primarily responsible for the AIDS in North America
and Western Europe.
• HIV-2 is less prevalent, less pathogenic & slow progression to AIDS and
cause lymphadenopathy.. Found in west Africa & India.
 Origin of HIV:
1. HIV-1 from cross-species transmission (zoonosis) of a chimpanzee virus to
human.
2. HIV-2 from cross-species transmission of a sooty mangabey virus.
• HIV discovered in 1984 by - Luc Montagneir - Pasteur Institute
• HIV is a retrovirus that infects cells of the human immune system (mainly
CD4 positive T cells and macrophages) & destroys or impairs their function
which results in the progressive depletion of the immune system, leading to
'immune deficiency‘ and AIDS.
8

9. p17
p24
Viral envelop
Proteases
Integrase
12. Pantaleo, G.; Graziosi, C.; Fuci, A.S. The immunopathogenesis of human immunodeficiency virus infection, N Engl J
Med, 1993, 328,327-335
HIV structure and molecular biologyHIV structure and molecular biology
9

10. • HIV has following types of cgenes encoding for structural proteins.
 The gag gene provides the physical infrastructure of the virus;
 The pol provides the basic enzymes by which retroviruses reproduce;
 The env gene supplies the proteins essential for viral attachment and entry
into a target cell.
• The accessory proteins tat, rev, nef, vif, vpr, and vpu enhance virus
production. tat and rev are essential for viral replication.
• A mutation causes the production of alternate accessory protein, from the
fusion of tat, rev, and env in some HIV.
The gp120 and gp41 proteins, both encoded by the env gene, enable the
virus to attach to and fuse with target cells to initiate the infectious cycle.
15. Mehannna, A. S. Rational of Design of Anti-HIV Drugs, Abraham, D. J.,Burger’s Text Book of Medicinal
Chemistry Drug Discovery, A John Wiley and Sons, Inc., 6th ed, 2003, 5, 461.
HIV genomeHIV genome
10

11. 1. Viral Attachment
2. Viral Penetration/Fusion
3. Uncoating
4. Reverse Transcription
5. Integration
6. Viral Latency and Protein Synthesis
7. Cleavage & Viral assembly
8. Budding
Mehannna, A. S. Rational of Design of Anti-HIV Drugs, Abraham, D. J., Burger’s Text Book of Medicinal Chemistry Drug
Discovery, A John Wiley and Sons, Inc., 2003, 6th ed, 5, 461-464.
Important steps of HIV life cycleImportant steps of HIV life cycle
11

12. Virus binding to the host cell membraneVirus binding to the host cell membrane
12

13. Viral Penetration/FusionViral Penetration/Fusion
13
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14. 14
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15. Viral DNA formation by the reverse transcriptaseViral DNA formation by the reverse transcriptase
15
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16. Integration of viral DNA into the host genomeIntegration of viral DNA into the host genome
16
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17. Integration of viral DNA into the host genomeIntegration of viral DNA into the host genome
17
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18. Integration of viral DNA into the host genomeIntegration of viral DNA into the host genome
18
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19. Assembly of viral proteins to form the virionAssembly of viral proteins to form the virion
19
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20. Viral budding out of the host cellViral budding out of the host cell
20
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21. • Mode of viral transmission
I. Sexual (primarily receptive anorectal and vaginal intercourse)
II. Parenteral (sharing of blood-contaminated needles by injection-drug
users, needlesticks, intravenous injection with used needles, receipt of
blood products, or organ transplants)
III.Perinatal/Vertical transmission (mother-to-child transmission-15-30%
risk during pregnancy, labor or breastfeeding: prolonged rupture of
membranes, chorioamnionitis, genital infections during pregnancy, preterm
delivery, vaginal delivery, illicit drug use during pregnancy, high maternal
viral load and Breast-feeding)
IV.Infected body fluids (semen and cervical secretions)
HIV infection and its pathological effectsHIV infection and its pathological effects
21

22. • It targets T4-lymphocyte, a white blood cell that has a central role
specifically the CD-4 helper T-cell.
• HIV starts its replication cycle in the host cell with the help of the
enzyme Reverse Transcriptase
• Fall from a normal value of 800-1300 cells/cm2 of blood to below 200
which may give threatening illness.
• The suppressed immune system leaves the patient vulnerable to the so-
called opportunistic infections by agents that would not harm a healthy
person.
• The most common of such infections is pneumonia caused by
Pneumocystis carinii which cause most of the clinical complications
Fauci, A. S. The human immunodeficiency virus: infectivity and mechanisms of pathogenesis. Science, 1988, 239,
617-622.
Cellular picture of the infection
Clinical picture of the infection
Immunosuppressive effect
22

23. • It is observed in the brain , independent of the immunodeficiency are an
abnormal proliferation of the glial cells that surround the neurons and lesions
resulting from loss of white matter.
• This can give rise to a neurological symptoms such as dementia and multiple
sclerosis.
• With HIV virus there is an increased risk of at least three types of human
tumors.
 Kaposi's sarcoma a rare tumor of blood vessel tissue in the skin or internal
organs.
 carcinoma including skin cancer which are often seen in the mouth or rectum
of infected homosexuals.
 B-lymphomas (tumors originating in B-lymphocytes).
Neurological effect
Carcinogenic effects
23

24. 1
2
3
4
5
6
7
8
9
10
T
A
R
G
E
T
S
T
A
R
G
E
T
S
24

25. N N
N
SS
O
O
O
O
CH3
H3C
Cyclotriazadisulffonamide (CADA)
• Cyclotriazadisulfonamide (CADA) (1) was recently shown to inhibit
HIV (as well as human herpesvirus type 7) infection by down-modulation
of the cellular CD4 receptor.
• CADA derivatives not alter the expression of any other cellular receptor
(i.e., CXCR4, CCR5).
1. CD4 as the primary cell receptor1. CD4 as the primary cell receptor
25

26. • CADA is assumed to down-regulate CD4 expression at the post translational
level.
• CADA ,it proved synergistic in its anti-HIV activity when combined with
 NRTIs (i.e., zidovudine, lamivudine, zalcitabine, abacavir),
 NtRTIs (i.e., tenofovir), NNRTIs (i.e., nevirapine, delavirdine),
 PIs (protease inhibitors lopinavir, saquinavir, indinavir, nelfinavir,
amprenavir. and ritonavir,
 The gp41 fusion inhibitor T-20 (enfuvirtide),
 The CXCR4 antagonist AMD3100, and the
 Mannose-specific plant lectins from Galanthus nivalis (GNA) and
Hippeastrum hybrid (HHA).4
Mechanism of action
26

27. • Galanthus nivalis (GNA) and Hippeastrum hybrid (HHA) ,the plant lectins
derived from GNA (Snowdrop) and HHA (Amaryllis) were shown to
interrupt the viral entry process by interfering with the viral envelope
glycoprotein gp120.
• When HIV-1 was made resistant to GNA and HHA, several amino acid
changes were noted in gp120 but not gp41; occurred at the N-glycosylation
sites (at the S or T residues).
• They are used as topical microbicides for the prevention of the sexual
transmission of HIV infection.
2. Virus Attachment Inhibitors2. Virus Attachment Inhibitors
27

28. • Cyanovirin
A potential microbicide to prevent the transmission of HIV and AIDS ,
isolated from the cyanobacterium Nostoc ellipsosporum.
• Cyanovirin-N has a uniquely high affinity for gp120:
 it impairs both CD4-dependent and -independent binding of gp120 to the
target cells,
 it blocks CD4-induced binding of gp120 with CXCR4, and it dissociates
bound gp120 from target cells.
• The aglycons of the glycopeptide antibiotics vancomycin, teicoplanin, and
eremomycin, were found to display activity against HIV-1, HIV-2, and
Moloney murine sarcoma virus at the lower concentration range.
28

29. • The Teicoplanin aglycon , interferes with viral entry, probably at the virus
adsorption step, because their anti-HIV activity was lost if added at 1-2 h
after infection.
• Glycopeptide antibiotic aglycon derivatives could be envisaged as
potential lead compounds for application as microbicides against sexual
HIV transmission.
29

30. • BMS- 378806 : A new class of HIV-1 attachment inhibitors [prototype: 4-
benzoyl-1-[4-methoxy-1H-pyrrolo[2,3-b]- pyridin-3-yl)oxoacetyl]-2-(R)-
methylpiperazine (BMS- 378806) (4)] that interferes with CD4-gp120
interactions.
• It binds directly to gp120 at a stoichiometry of approximately 1:1, with a
binding affinity similar to that of soluble CD4.
N
N
H
O
N
N
O
OCH3
CH3
O
BMS-378806
4
30

31. • The potential target site of BMS- 378806 was localized to a specific region
within the CD4 binding pocket of gp120.
• The 4-resistant variants have been isolated with amino acid substitutions
(M426L and M475I) in the CD4-binding pocket of gp120, confirming that
BMS- 378806 targets gp120.
• BMS- 378806 displays many favorable pharmacological traits such as
 Low protein binding,
 Minimal human serum effect on anti- HIV-1 potency,
 Good oral bioavailability in animal species, and
 A clean safety profile in initial animal toxicology studies.
31

32. • BMS-488043 : This Virus attachment inhibitors may be effective in vivo
in HIV-1-infected patients (5), which is structurally closely related to
BMS- 378806 .
• BMS- 488043 is assumed to bind to the HIV-1 glycoprotein gp120 and to
induce or block conformational changes in the gp120.
• The compound was well tolerated, and no serious adverse events were
noted.
N
N
H
O
N
N
O
OCH3
O
BMS-488043
5OCH3
32

33. • To enter the cells following binding with the CD4 receptor, the HIV
particles must interact, again through the viral envelope glycoprotein
gp120, with the CXCR4 or CCR5 coreceptor.
• CXCR4 is the coreceptor for Tlymphotropic (or X4) HIV strains, whereas
CCR5 is the coreceptor for macrophage (M)-tropic (or X5) HIV strains.
• CXCR4 normally functions as the receptor for the chemokine SDF-1
(stromal cell derived factor), and CCR5 does so for RANTES (regulated
upon activation, normal T-cell expressed, and secreted) and MIP-1R and
-1‚ (macrophage inflammatory proteins), and accordingly, these
chemokines inhibit the infectivity of X4 and X5 HIV strains, respectively.
3. CXCR4 and CCR5 Antagonists3. CXCR4 and CCR5 Antagonists
33

34. • Bicyclam AMD3100 (6) is low-molecular-weight CXCR4 and CCR5
antagonists
• Compound 6 is not truly specific for CXCR4. It does interact with any
other CXCR or CCR receptor and blocks X4 HIV-1 replication through
CXCR4 antagonization.
N N HNNH
HNNH NH HN
AMD3100
6
34

35. • During phase I clinical studies AMD3100 caused a significant
enhancement of the white blood cells (WBC) counts in human volunteers.
• It mobilize hematopoietic stem cells from the bone marrow into the
bloodstream, and because this effect proved synergistic with the action of
G-CSF (granulocyte-colony stimulating factor), so AMD3100 is now being
pursued (in phase II clinical studies) for stem cell mobilization and
transplantation in patients with multiple myeloma or non-Hodgkin
lymphoma.
• CXCR4 antagonists such as AMD3100 are able to suppress in vivo
replication of X4 or dualtropic X4/R5 HIV strains.
35

36. • Structure-activity relationship (SAR) studies revealed that the bis-
macrocyclic structure is not an absolute prerequisite for anti-HIV activity.
• For example: AMD3465 (7), an AMD3100 derivative ,proved to be as
potent, as AMD3100 as both a CXCR4 antagonist and inhibitor of X4 HIV
strains.
• Also, KRH-1636 (8), another CXCR4 antagonist, with anti-HIV activity
similar to that of AMD3100.
• KRH-1636 appears to be duodenally absorbable, at least in rats, which
means that it may be orally bioavailable.
N N
H
NH
HNNH
AMD3465
7
N
N
H
O
H
N
O
N
H
CH3
NH
H2N NH
N
KRH-1636
8
36

37. • TAK-779 (9): The CCR5 antagonists, the quaternary ammonium derivative
was the first nonpeptidic molecule which block the replication of the M-
tropic R5 HIV-1 strains (in the nanomolar concentration range) by
interaction with CCR5.
• The binding site for TAK-779 has been identified within the
transmembrane (TM) helices 1, 2, 3, and 7 of CCR5.23
• TAK-779 is not orally bioavailable and provokes irritation at the injection
site.
H3C
O
H
N
N
CH3
CH3
O
Cl-
TAK-779
9
37

38. • SCH 351125 (SCH-C) (11) was the first CCR5 antagonist to be advanced
to clinical studies.
• SCH 351125 has potent activity in vitro against primary HIV-1 isolates
using CCR5 as their cell entry coreceptor.
• SCH-351125 bind to overlapping but nonidentical sites within a putative
ligand-binding cavity formed by the transmembrane (TM) helices 1, 2, 3,
and 7.
N
N
N O
N
Br
O
OSCH-C(SCH 351125)
11 38

39. • Spirodiketopiperazine E913 (12).
• E913 specifically blocks the binding of MIP-1R to CCR5, the MIP-1R-
elicited Ca2+ flux, and the replication of both laboratory and primary R5
HIV-1 strains, as well as various multidrug-resistant monocyte/ M-tropic
R5 HIV-1 strains.
• From the spirodiketopiperazine class of compounds, E913 was recently
quoted as binding only partially to CCR5 while exhibiting much greater
anti-HIV activity.
O
O
N
NH
N
O
O
39

40. • UK-427,857 (13): CCR5 antagonist, has recently been selected as a
clinical development candidate drug for the treatment of HIV infection.
• UK-427,857 was reported to display excellent potency against a isolates
utilizing CCR5 for cell entry (at IC90 < 10 nM) while being inactive
against CXCR4-tropic viral isolates.
F
F
O
H
N
N
H
H
N N
N
UK-427857
13
40

41. • 1,3,4-trisubstituted pyrrolidine : CCR5 receptor antagonists have possess
oral bioavailability and/or potent anti-HIV activity.
• A representative congener of this 1,3,4- trisubstituted pyrrolidine series is
14. The compound’s site of interaction with CCR5 has been mapped to a
cavity, near the extracellular surface, formed by the TM helices 2, 3, 6, and
7.
N
N
O
2
1
3
4
5
1,3,4-Trisubstituted pyrrolidine
14
41

42. • CMPD 167, CCR5 antagonists are efficacious in vivo against CCR5-using
virus strains, designated previously as MRK-1 (15).
• This compound caused a rapid and substantial (4- to 200-fold) decrease in
plasma viremia in rhesus macaques chronically infected with simian
immunodeficiency virus (SIV).
• It is used as a topical microbicide to prevent HIV-1 sexual transmission
because viral replication could be partially inhibited by it.
H3C
CH3H3C
O
CH3
N
N
N
CH3
F
MRK-1
CMPD 167
15
42

43. • The interaction of the X4 or R5 HIV-1 envelope gp120 with the
coreceptors CXCR4 or CCR5, respectively, is followed by a spring-loaded
action of gp41 that then anchors through its amino terminus into the target
cell membrane and, in doing so, initiates the fusion of the viral envelope
with the cellular plasma membrane.
• During the process, hydrophobic grooves on the surface of the coiled coil
gp41 ectodomain become available for binding with extraneous inhibitors,
that are virus cell fusion inhibitors.
4. Virus-Cell Fusion Inhibitors4. Virus-Cell Fusion Inhibitors
43

44. • For example: Enfuvirtide (T-20, DP-178, pentafuside, Fuzeon) (16), a
synthetic, 36-amino-acid peptide corresponding to residues 127-162 of
gp41 or residues 643-678 of the gp160 precursor.
44

45. • Drawbacks of Enffuvirtide
 Enfuvirtide has to be administered twice daily by subcutaneous injection.
This inevitably leads to injection- site reactions including erythema,
induration, and nodules, and cysts.
 Another problem is the production cost for a 5000 Da molecular mass
peptide such as enfuvirtide.51
 Even enfuvirtide-insensitive HIV-1 variants may exist in an enfuvirtide-
naive population, and this may allow virus escape from drug.
45

46. • Tris-functionalized 3,2’,2’’-terphenyl derivatives (17) could serve as
effective mimics of the exposed N-helical regions of the transient gp41
intermediate and thus potentially trap this structure prior to the six-helix
bundle formation, which is required for virus-cell fusion.
• Compound 17 was found to inhibit HIV-1 mediated cell-to cell fusion but
only at a relatively high concentration (IC50 15 μg/mL).
O
OH
OCH3
CH3
CH3H3C
CH3
H3C
HO
O
Terphenyl derivative
17
46

47. • Zidovudine (AZT, Retrovir),
• Didanosine (ddI, Videx),
• Zalcitabine (ddC, Hivid),
• Stavudine (d4T, Zerit),
• Lamivudine (3TC, Epivir) (also marketed in combination with
zidovudine under the trademark Combivir and with abacavir under the
trademark Epzicom), and
• Abacavir (ABC, Ziagen) (also marketed in combination with zidovudine
and lamivudine under the trademark Trizivir),
• Emtricitabine (2’,3’-dideoxy-3’-thia-5-fluorocytidine, (-)-FTC,
previously referred to as Coviracil and now marketed as Emtriva) is the
seventh 2’,3’- dideoxynucleoside analogue that has been officially
approved (for the treatment of HIV infections.
5. Nucleoside Reverse Transcriptase Inhibitors
(NRTIs)
5. Nucleoside Reverse Transcriptase Inhibitors
(NRTIs)
47

48. N
N
NH2
O
O
HO
Zalcitabine
N
N
NH2
O
O
S
HO
Lamivudine
HN
N
O
O
O
HO
CH3
Stavudine
HN
N
O
O
O
HO
CH3
N3
Zidovudine
HN
N
N
O
O
HO
Didanosine
48

49. • Emtricitabine can be conveniently administered as a once-daily dose of
200 mg for long term clinical use in HIV-1-infected individuals.
• In studies through 60 weeks, once daily emtricitabine, combined with
once-daily didanosine and efavirenz, demonstrated durable and superior
virologic efficacy and tolerability, compared to twice-daily stavudine and
once-daily didanosine and efavirenz.
S
O
HO
N
N
NH2
F
O
Emtricitabine [(-)FTC]
18 49

50. • Emtricitabine has been considered an “ideal drug candidate” because
 It shows synergism with other antiretrovirals,
 Excellent tolerability,
 A long intracellular half-life (supporting the once-daily dosing), and,
 In comparison with lamivudine, 4- to 10-fold higher in vitro potency
against HIV.
50

51. • Amdoxovir ((-)-‚-D-2,6- diaminopurine dioxolane, DAPD, 21):
New purine -based 2’,3’-dideoxynucleoside analogues
• DAPD is converted by adenosine deaminase to dioxolane guanine (DXG)
which is then phosphorylated intracellularly to DXG 5’-triphosphate, the
active metabolite.
• DXG 5’-triphosphate (DXG-TP) acts as an alternative substrate/ inhibitor
of the HIV-1 reverse transcriptase.
• DAPD/DXG has proven active against HIV-1 mutants resistant to
zidovudine (M41L/D67N/K70R/T215Y/K219Q) and lamivudine
(M184V)73 but has decreased activity against viruses carrying the K65R
and Q151M mutations.
N
N
N
N
NH2
O
O
HO
Amdoxovir (DAPD)
21
NH2
51

52. • The nucleotide reverse transcriptase inhibitors (NtRTIs) such as
 Adefovir [9-(2-phosphonylmethoxyethyl) adenine (PMEA)] and
 Tenofovir [(R)-9-(2- phosphonylmethoxypropyl)adenine (PMPA)]
are already equipped with a phosphonate group and, therefore, only need
two phosphorylation steps to be converted to the active metabolites
(PMEApp and PMPApp, respectively).
• The metabolites then serve as alternative substrates in the reverse
transcriptase reaction, where upon their incorporation they act as chain
terminators.
6. Nucleotide Reverse Transcriptase Inhibitors
(NtRTIs)
6. Nucleotide Reverse Transcriptase Inhibitors
(NtRTIs)
52

53. • The NtRTIs Adefovir and Tenofovir are active not only against HIV but
also against hepatitis B virus (HBV), because HBV uses for its replication
a reverse transcriptase that is quite similar to that of HIV.
• They have been officially approved, in their oral prodrug forms Adefovir
dipivoxil [bis( pivaloyloxymethyl )-PMEA, Hepsera] (22) and Tenofovir
disoproxil [bis( isopropyloxycarbonyloxymethyl )- PMPA] fumarate
(Viread) (23), for the treatment of HBV and HIV infections, respectively.
N
N
N
N
OP
O
O
O
NH2
OH2C
OH2C
C
C
O
O
(H3C)3C
(H3C)3C
Adefovir dipivoxil
22
N
N N
N
OP
O
O
O
NH2
OH2C
OH2C
C
C
O
O
O
O
Tenofovir disoproxil fumarate
23
H3C
(H3C)3C
(H3C)3C
HOOC
COOH
53

54. • Both the d4T (Stavudine) and TDF (Tenofovir) arms showed a similar,
high virological response. However, lipid abnormalities (increase in
triglyceride and cholesterol levels) were significantly lower in the TDF arm
than in the d4T arm.
• Also, the toxicities (peripheral neuropathy, lipodystrophy, lactic acidosis,
pancreatitis) associated with mitochondrial dysfunction through week 96
were markedly lower in the TDF arm than in the d4T arm, while both arms
showed a similar renal safety profile.
• Although in an animal model for HIV transmission through breast-feeding
topical administration of lowdose TDF did not protect infant macaques
against multiple oral exposures of simian immunodeficiency virus (SIV),
54

55. • Another indication for the clinical use of tenofovir disoproxil fumarate is
(lamivudine-resistant) chronic hepatitis B in HIV/HBV- coinfected
patients.
• Several studies have demonstrated that TDF is very effective in reducing
HBV DNA levels in HIV/HBV- coinfected patients carrying either wild-
type or 3TC-resistant [YMDD variant (rt M204I/V)] HBV.
• Noteworthy is that TDF treatment for 12 months in patients coinfected
with HIV and 3TC-resistant HBV was not associated with the emergence
of HIV- or HBV-specific resistance mutations.
55

56. • More than 30 structurally different classes of compounds have been
identified as NNRTIs, that are targeted at a specific, allosteric (i.e.,
nonsubstrate binding) site of the reverse transcriptase.
• The “first-generation” NNRTIs are notorious for rapidly eliciting virus
drug resistance, especially when used singly.
• The most common mutations occurring in the clinical setting following the
use of NNRTIs are K103N and Y181C.
• Therefore, attempts have been made to develop “second-generation”
NNRTIs that are resilient to such drug resistance mutations.
• “first-generation” NNRTIs are Nevirapine and Delavirdine while
“Second-generation” NNRTIs are Efavirenz .
7. Non-Nucleoside Reverse Transcriptase Inhibitors
(NNRTIs)
7. Non-Nucleoside Reverse Transcriptase Inhibitors
(NNRTIs)
56

57. • DAPY derivatives
1. TMC 125 (Etravirine) (27) is an investigational NNRTI with potent
activity against HIV-1 strains resistant to the currently available NNRTIs.
• It is under the advance clinical development.
N
O
N
N NH
N
Br
NH2
Etravirine (Tmc125)
27
57

58. 2. Rilpivirine
more active than NVP, EFV and dapivirine against wild-type HIV-1
and all single and double mutants tested .
• E-form was superior in terms of antiviral activity with respect to
the corresponding Z-isomer.
• Rilpivirine was not mutagenic,no relevant side effects, had no
effect on cardiovascular, pulmonary, electrophysiological and
behavioral parameters in dog.
• It is also under the advance clinical development.
HN
N
N NH
N
Rilpivirine
CN
58

59. • Dihydropyrazinones
• The cyano substituent in the 4-position of the aniline moiety was the
optimal choice,
• The presence of a 6-methyl group in the dihydropyrazinone ring
cause a favorable effect on activity against mutants.
• Introduction of methyl substituents on the benzene ring and at
position- 6 of the dihydropyrazinone ring on the left wing increased
the activity against both HIV wild-type and drug-resistant mutants.
• Compounds with X = SO2 were slightly superior to those with X = S,
which was only confirmed for the Y181C mutation.
N
N
N
H
X
R
R1
OR2
R3
35
2
1
6
4
59

60. • Benzophenones
• These compounds proved to be much more potent than NVP and
DLV, and equivalent to EFV against both wild-type HIV-1 and the
Y181C mutant strain.
R1
R2
O
Cl
O
O
H
N
SO2NH
23 GW4511 R1=CF3, R2 =F
24 GW4751 R1=H, R2=CN
F3C
F
O
Cl
O
O
H
N
25 GW3011
O SO2NH
60

61. • 2-Quinolones
• The quinolones were demonstrated to be potent inhibitors of wild-
type (HIV-1 IIIB) and NVP-resistant strains in MT-4 cell assays.
• R = F; Cl as substituents were optimal for antiviral activity in MT-4
cells.
• Replacement of R with CH3 and OCH3, led to much less active
compounds.
• Introduction of a cyclopropylethynyl group as the R2 substituent led
to the more potent compounds
N
H
X
Cl R
O
61

62. • Indolyl aryl sulfone
• The iodole derivatives bearing the 3-[(3,5-dimethylphenyl) sulfonyl]
moiety (52) displayed high activity and selectivity not only against
the HIV-1 wild-type strain, but also against the Y181C and K103N-
Y181C viral variants and the EFV-resistant (EFVR) mutant K103R-
V179D-P225H.
HN
Cl
O
NH2
SO2
R1
R2R3
62

63. • Capravirine analogues
• Capravirine retained activity against HIV-1 strains carrying the K103N
mutation in their reverse transcriptase.
• Replacement of ethyl in position-4 of the imidazole nucleus with an
iso-propyl group led to more potent derivatives.
• The activity of the compounds was also affected by the group at the
5-position of the imidazole ring in the order 3,5-(CH3)2-phenyl >
cyclohexyl > phenyl group
• The alkoxymethyl substituent at the 1-position of imidazole ring
showed higher activity than compounds containing a 4-pyridylmethyl
group at the same position.
S
N
N
O
O
H2N
CH3
H3C
Cl
Cl
N
Capravirine (S-1153,AG1549)
25
1
2
3
4
5
63

64. • Pyridones
• in vitro activity profile of these new molecules, and in particular of
compound 118, was better than that of EFV.
HN
X
O
N
111 X= H2
112 X = O
HN
X
O
N
117 X= O
118 X = H2
CN
64

65. • Integration of the proviral DNA into host cell chromosomal DNA is an
essential step in the viral replication cycle.
• This process is mediated by the viral integrase, and because there is no
cellular homologue for this enzyme, it has been considered an attractive
target for HIV therapeutics.
• Numerous small-molecule HIV-1 integrase inhibitors have been described,
the two most predominant classes of inhibitors being the catechol
containing hydroxylated aromatics and the diketo acid containing
aromatics.
8. HIV Integrase Inhibitors8. HIV Integrase Inhibitors
65

66. • Catechol containing hydroxylated aromatics
• An example is L- chicoric acid (28).
• Integrase was identified as the molecular target for the action of L- chicoric
acid, since a single amino acid substitution (G140S) in the integrase
rendered the corresponding HIV-1 mutant resistant to L- chicoric acid.
• It was later shown that the G140S mutation confers resistance not only to
L- chicoric acid but also to the diketo acid L-731,988 and furthermore
attenuates the catalytic activity of the enzyme.
HO
HO
O
O
O
OH
O
O
O
OH
OH
L-Chicoric acid
28
HO
66

67. • In our hands, however, HIV-1 integrase carrying the G140S mutation
appeared to be as sensitive to the inhibitory effect of L- chicoric acid as the
wild-type integrase.
• Upon repeated passages of the virus in the presence of L- chicoric acid,
mutations were found in the viral envelope glycoprotein gp120 but not in
the integrase.
• HIV strains resistant to polyanionic compounds showed cross-resistance to
L- chicoric acid, and time-of addition experiments further confirmed that
the primary site of interaction for L- chicoric acid is the virus adsorption
stage rather than the integrase.
67

68. • The structure of the HIV-1 integrase core domain complexed with the
inhibitor 5CITEP [1-(5-chloroindol- 3-yl)-3-hydroxy-3-(2H-tetrazol-5-yl)
propenone] has been described as a platform for the structure-based design
of novel HIV-1 integrase inhibitors.
C
O
N
H
N
N
N
HO NH
Cl
5CITEP
• Diketo acid containing aromatics.
68

69. F
N
COOH
O O
L-731,988
O
COOH
O
O O
L-708,906
O
O
OH
O
OH
O
O
HO
Granulatine
H
N
H
N
O O
OH HO
Salicylhydrazine
COOHCHO
O
O
Integric acid
69

70. • There are number of diketo acids such a L-708,906 (29) and L-731,988
(30) as inhibitors of the integrase-mediated strand transfer reaction (which
is responsible for the covalent linkage of the viral DNA 3’ ends to the
cellular DNA).
• The diketo acid derivative 29 and 30 were also found to inhibit HIV-1
replication in cell culture .
• The proviral DNA integration is indeed the target of action for the diketo
acids in cell culture.
N
F
O O
O
OH
L-731,988
30
O
O O
O
OHO
L-708,906
29 70

71. • S-1360 (31) actually represents the first integrase inhibitor to reach clinical
studies.
• S-1360 would inhibit the HIV-1 integrase at an IC50 of 20 nM and the
HIV-1 replication at an EC50 of 140 nM while its CC50 would be 110 μM,
thus achieving a therapeutic index of almost 1000, the highest selectivity.
• The mechanism of action of the diketo acids may be based on an
interaction between the carboxylate group of the diketo acids or the
isosteric heterocycle in the two other compounds and metal ion(s) in the
active site of the integrase, resulting in a functional sequestration of these
critical metal cofactors.
O
O OH
F
N
NHN
S-1360
31
71

72. • Diketo acids could be classified into two groups:
 Those similarly potent in the presence of magnesium and
 Those potent in manganese and relatively ineffective in the presence of
magnesium.
• Both the aromatic and the carboxylic or tetrazole functions of the diketo
acids determine their metal-chelating selectivity.
• Starting from 30, several additional 4-aryl-2,4-dioxobutanoic acid
derivatives have been described as HIV-1 integrase inhibitors.
• Also, new azido-containing aryl diketo acids have been described as HIV-1
integrase inhibitors capable of conferring antiviral protection in HIV-
infected cells.
72

73. • 8-hydroxy[1,6]-naphthyridines.
Cause Inhibition of the strand transfer reaction of the integration process.
• The naphthyridine shown (32) inhibits strand transfer catalyzed by
integrase with an IC50 of 10 nM and inhibits 95% of the spread of HIV-1
infection in cell culture at 0.39 μM.
• While no cytotoxicity is exhibited in cell culture at e12.5 μ M, and a good
pharmacokinetic profile was displayed when dosed orally to rats.
N
O
N
N
OH
S
O
O
8-Hydroxy-[1,6]naphthyridine
32 73

74. • Recently, an entirely new class of HIV integrase inhibitors was 5H-
pyrano- [2,3-d:-6,5-d¢]dipyrimidines (PDPs).
• The most potent congener of this series, 5-(4-nitrophenyl)-2,8-dithiol-4,6-
dihydroxy-5H-pyrano[2,3-d:-6,5-d’]dipyrimidine (V-165) (33), inhibited
the replication of HIV-1 at an EC50 of 8.9 μM, which is 14-fold below the
cytotoxicity threshold.
N
N O N
N
OHOH
HS
SH
NO2
Pyranodipyrimidine (PDP) (V-165)
33
74

75. • At the transcriptional level, HIV gene expression may be inhibited by
compounds that interact with cellular factors (such as NF-B) that bind to
the LTR promotor and that are needed for basal-level transcription.
• These compounds specifically inhibit the transactivation of the HIV LTR
promotor by the viral Tat (trans-activating) protein.
• These compounds inhibit HIV-1 replication in both acutely and chronically
infected cells, through interference with the transcription process that
attributed to inhibition of Tat (or other transactivators).
9. Transcription (Transactivation) Inhibitors9. Transcription (Transactivation) Inhibitors
75

76.  K37 [7-(3,4-dehydro-4-phenyl-1-piperidinyl)- 1,4-dihydro-6-fluoro-1-
methyl-8-trifluoromethyl- 4-oxoquinoline-3-carboxylic acid;
 Temacrazine [1,4-bis(3-(6-oxo-6H-v-triazolo[4,5,1-de]acridin-5-
ylaminopropyl) piperazine];
 Flavopiridol, a cyclin-dependent kinase (CdK) inhibitor;
 EM2487, a natural product from Streptomyces; and
 CGP 64222, a 9-mer peptoid
structurally reminiscent of the amino acid 48- 56 sequence RKKRRQRRR
of Tat. peptoid CGP 64222 owes its anti-HIV activity in cell culture
primarily to an interaction with CXCR4, the coreceptor for X4 HIV strains.
76

77. • A 6-aminoquinolone, WM5 (34), was recently shown to inhibit HIV-1
replication in acutely infected as well as chronically infected cells.
• This aminoquinoline was found to efficiently bind to TAR RNA and to
inhibit Tat mediated LTR-driven transcription.
N
N
N N
H2N
CH3
O O
OH
Aminoquinolone WM5
34
77

78. • Recently prooved that the co transcriptional capping of HIV mRNA is
stimulated by Tat and consequently its binding to the capping enzyme.
• These findings implicate capping as an elongation checkpoint critical to
HIV gene expression and thus corroborate that S-adenosylmethionine-
dependent methylations play an important role in the Tat dependent
transactivation of transcription from LTR.
• They also explain the inhibitory effects of S-adenosylhomocysteine
hydrolase inhibitors, such as neplanocin A and 3-deazaneplanocin A (35),
on Tat dependent transactivation and HIV replication.
N
X
N
N
NH2
HO
OH OH
X= N2 Neplanocin A
X= CH: 3-Deazaneplanocin A
35
78

79. • After transcription the unspliced (or partially spliced) HIV mRNA has to
be transported from the nucleus into the cytoplasm in order to be translated
to viral proteins.
• This export is promoted by the HIV-1 Rev (regulator of expression of viral
proteins).
• Nuclear export of Rev is mediated by its leucine-rich nuclear export signal
(NES). NES uses the export factor CRM1 to export viral mRNA from the
nucleus to the cytoplasm.This process can be blocked by a small
molecular- weight molecule, PKF 050-638 (36), that specifically inhibits
CRM1-NES complex formation and, hence, Rev-mediated nuclear export.
N
N
N
Cl
NH2
O
CH3
O
PKF 050-638
36
79

80. • N-aminoimidazole derivatives (NAIMS), structurally analogous to
Capravirine (S-1153, AG1549) have been found to inhibit HIV replication
through one of the following types of interaction.
 (i) the classical NNRTI type of action.
 (ii) an as yet unidentified target of action; and
 (iii) combination of type i and type ii actions.
N
N
HN
R
Y
X2
X1
N-aminoimidazoles (NAIMS)
37
X1
,X2
; Halogen ,Alkyl
R :Alkyl , Aryl
Y: SH , H
80

81. • Pyridine oxide derivatives (prototypes JPL-32, JPL-88, and JPL-133) (38)
like the NAIMS, can exhibit same behavior.
• Some of the pyridine oxides behave as typical NNRTIs.
S
R1
R2
N Y1
Y2
Y3
Y4
X3
X2X4
X5 X1
+
O -
Pyridine oxides JPL-32,-88 and-133
JPL-32 JPL-88 JPL-133
X1 Cl CH3 CH3
X2 Cl H H
X3 Cl CH3 H
X4 Cl H CH3
X5 Cl CH3 H
Z H H H
R1 O - -
R2 O O -
Y1 H H H
Y2 H H H
Y3 H H H
Y4 H H Cl
81

82. • The most active congener from this series, JPL-133, has an EC50 of 0.05
μg/mL against HIV-1 and a selectivity index of approximately 760 in cell
culture.
• For other pyridine oxides such as JPL-32 and JPL-88, experiments
revealed a postintegration step in the HIV replicative cycle as the most
likely target of action.
• The mode of action of JPL-32 appeared to be reminiscent of that of K-37
because JPL-32, like K-37, was active in both acutely and chronically HIV-
1-infected cells.
• JPL- 32 also inhibited the Tat-mediated HIV-1 mRNA transcription from
HIV-1 LTR, thus clearly indicating that its target of action is located at the
transcription transactivation level.
82

83. • The HIV protease is responsible for the cleavage of the gag and gag-pol
precursor polyproteins to the structural proteins (p17, p24, p7, p6, p2, p1)
and functional proteins [protease (p11), reverse transcriptase (p66/p51),
and integrase (p32)], thereby securing maturation and infectivity of the
progeny virions.
• HIV protease inhibitors will interfere with this late stage of the viral
replication cycle and prevent formation of infectious virus particles.
10. HIV Protease Inhibitors10. HIV Protease Inhibitors
83

84. • All protease inhibitors (PIs) that have been licensed for the treatment of
HIV infections, namely,
 Saquinavir (Fortovase, Invirase),
 Ritonavir (Norvir),
 Indinavir (Crixivan),
 Nelfinavir (Viracept),
 Amprenavir (Agenerase) (and its phosphate derivative, Lexiva),
 Lopinavir (Kaletra, lopinavir with ritonavir at a 4:1 ratio), and
 Atazanavir (Reyataz),
N
N
N
H
H
N O
O
H
N
N
H
O
O
O
O OH
Atazanavir (BMS-232632)
39
84

85. • The Aza-dipeptide analogue Atazanavir (39), the latest and seventh PI
 To be approved for clinical use,
 Combines a favorable resistance profile distinct from that of the other PIs,
 A favorable pharmacokinetic profile allowing once-daily dosing.
• Nelfinavir-, Saquinavir-, and Amprenavir-resistant HIV-1 strains
remained sensitive to Atazanavir, while Indinavir- and Ritonavir-
resistant viruses showed 6- to 9-fold changes in sensitivity to atazanavir.
• Conversely, Atazanavir-resistant (N88S, I84V) virus, selected upon
repeated passage of the virus in the presence of the compound, remained
sensitive to Saquinavir but showed various levels of cross-resistance to
Nelfinavir, Nndinavir, Ritonavir, and Amprenavir.
85

86. • TMC 126 (UIC-94003) ,Compound 40 is a peptidomimetic PI containing a
bis-tetrahydrofuranyl urethane and 4-methoxybenzenesulfonamide (and
thus structurally related to amprenavir).
• It was reported to be extremely potent against a wide spectrum of HIV-1
strains (EC50 ) 0.3-0.5 nM), whether resistant to other PIs or not.
N
H
N
S
OH CH3
CH3
O
O
OCH3
O
O
O
O
UIC-94003 (TMC-126)
40 86

87. • Upon passage of HIV-1 in the presence of 40, mutants carrying a novel
active-site mutation, A28S, emerged.
• Modeling studies revealed close contact of 40 with the main chain of the
protease active-site amino acid residues D29 and D30 different from that of
the other PIs, and this may be important for its potency, particularly
against drug-resistant HIV-1 variants.
• TMC 114, like TMC 126, has an excellent activity profile against HIV
variants that are highly resistant to current PIs.
87

88. • Tipranavir (PNU-140690) (41) is a nonpeptidomimetic inhibitor of the
HIV protease,which, shows little cross-resistance to the peptidomimetic
PIs.
• Tipranavir retained marked activity against HIV-1 isolates derived from
patients with multidrug resistance to other Pis.
• Tipranavir drug levels can be markedly increased (“boosted”) by
coadministration with ritonavir.
O O
OH
N
H
S
O O
N F
F
F
Tipranavir (PNU-140694)
41
88

89. • During the reverse transcription process that converts the single-stranded
viral RNA into double-stranded (pro)viral DNA, a RNA‚DNA
heteroduplex (hybrid) is formed.
• The RNA strand of this heteroduplex must be cleaved by the ribonuclease
H (RNase H) component before the remaining (-)- DNA strand can be
duplicated to give the doublestranded (pro)viral DNA that is then
integrated into the host cell genome.
• RNase H has been considered as an attractive target for the development
of new antiretroviral agents.
HIV Ribonuclease H InhibitorsHIV Ribonuclease H Inhibitors
89

90. • It is feasible to specifically inhibit HIV-1 Rnase H by, for example, novel
Diketo acids, such as 4-[5- (benzoylamino)thien-2-yl]-2,4-dioxobutanoic
acid (42), and N-hydroxyimides such as 2-hydroxy-4H-isoquinolone- 1,3-
dione (43).
• In both cases, binding of the compound was (much) stronger in the
presence of Mn2+ compared to Mg2+.
N
O
OH
O
2-hydroxyisoquinoline-1,3(2H,4H)-dione
O
H
N S
O O
O
OH
4-(5-benzamidothiophen-2-yl)-2,4-dioxobutanoic acid
42
90

91. Inhibitors of viral entry
• It is most ideal approach to inhibit HIV binding to the T-cell
• It can neutralize the virus in circulation.
• The initial work on vaccine development focused on iso-typic variants of
the HIV envelope glycoprotein gp120 obtained by recombinant DNA
techniques due to concern about the safety of live/attenuated vaccines.
• The gp120 glycoprotein is a coat protein, and if great care is taken, a virus-
free vaccine is obtainable.
• Moreover, glycoprotein gp120 is a primary target for neutralizing antibody
associated with the first step in HIV infection.
HIV vaccinesHIV vaccines
91

92. • Today it is facilitated the design of vaccination regimens that elicit specific
immune responses and effector mechanisms.
• The importance of CD8+ CTL responses in controlling HIV and SIV
viremia has led to a series of vaccine candidates that effectively induce
these responses.
• gp120-derived vaccines induced little cell-mediated immunity and strong
antibody response in T-cell lines, but failed to neutralize virus derived from
peripheral blood mononuclear cells.
• Recombinant attenuated vaccinia virus expresses key viral envelope
protein followed by a booster of soluble envelope protein derived from
HIV.
• It produced a good humoral and CMI response and IgA antibodies in
animal models.
92

93. • AIDSVAX25
• AIDSVAX from VaxGen Inc. is a preventive vaccine made up of synthetic
gp120. Two Phase III clinical trials were initiated. Failed due to lack of
adequate protection.
• DNA VACCINE26
• In recent years a new type of vaccine, created from an infectious agent's
DNA called DNA vaccination.
• It works by insertion (and expression, triggering immune system
recognition) into human or animal cells, of viral or bacterial DNA.
• As of 2006, DNA vaccination is still experimental, but shows some
promising results.
• However, the nature of the disease people infected with HIV develop only
low titers of neutralizing antibody and that presents problem for vaccine
development.
93