Anti HIV agents

Some facts about HIV:

• HIV – the Human Immunodeficiency Virus is the retrovirus that causes AIDS

• Discovered independently by Luc Montagnier of France and Robert Gallo of the US in 1983-84

• HIV belongs to the retrovirus subfamily lentivirus.

• Former names of the virus include:– Human T cell lymphotrophic virus (HTLV-III)– Lymphadenopathy associated virus (LAV)– AIDS associated retrovirus (ARV)

• One million people infected in US, 30 million worldwide are infected.

• Leading cause of death of men aged 25-44 and 4th leading cause of death of women in this age group in the US.

Methods of transmission:

– Sexual transmission, presence of STD increases likelihood of transmission.

– Exposure to infected blood or blood products.– Use of contaminated clotting factors by

hemophiliacs.– Sharing contaminated needles (IV drug users).– Transplantation of infected tissues or organs.– Mother to fetus, perinatal transmission variable,

dependent on viral load and mother’s CD4 count.

Transmission

Characteristics of the virus

• Icosahedral (20 sided), enveloped virus of the lentivirus subfamily of retroviruses.

• Retroviruses transcribe RNA to DNA.• Two viral strands of RNA found in core surrounded by

protein outer coat.– Outer envelope contains a lipid matrix within which

specific viral glycoproteins are embedded.– These knob-like structures responsible for binding to target

cell.

Characteristics of the virus:

HIV• The outer shell of the virus is

known as the Viral enevlope. Embedded in the viral envelope is a complex protein known as env which consists of an outer protruding cap glycoprotein (gp) 120, and a stem gp14. Within the viral envelope is an HIV protein called p17(matrix), and within this is the viral core or capsid, which is made of another viral protein p24(core antigen).

Viral Replication

• First step, HIV attaches to susceptible host cell.– Site of attachment is the CD4 antigen found on a variety of

cells• helper T cells• macrophages• monocytes• B cells• microglial brain cells• intestinal cells

– T cells infected later on.

Life Cycle:• (a) HIV (red) attaches to two cell-surface receptors

(the CD4 antigen and a specific chemokine receptor). • (b) The virus and cell membrane fuse, and the virion

core enters the cell. • (c) The viral RNA and core proteins are released from

the virion core and are then actively transported to the nucleus.

• (d) The viral RNA genome is converted into double-stranded DNA through an enzyme unique to viruses, reverse transcriptase (red dot).

• (e) The double-stranded viral DNA moves into the cell nucleus.

• (f) Using a unique viral enzyme called integrase, the viral DNA is integrated into the cellular DNA.

• (g) Viral RNA is synthesized by the cellular enzyme RNA polymerase II using integrated viral DNA as a template. Two types of RNA transcripts shorter spliced RNA (h) and full-length genomic RNA (j) are produced.

• (h) Shorter spliced RNAs are transported to the cytoplasm and used for the production of several viral proteins that are then modified in the Golgi apparatus of the cell (i).

• (j) Full-length genomic RNAs are transported to the cytoplasm (k).

• (l) New virion is assembled and then buds off. • (m) Mature virus is released.

Latency and manifestation of symptoms:

• After a period of latency lasting up to 10 years viral replication is triggered and occurs at high rate.

• CD4 cell may be destroyed in the process, body attempts to replace lost CD4 cells, but over the course of many years body is unable to keep the count at a safe level.

• Destruction of large numbers of CD4 cause symptoms of HIV to appear with increased susceptibility to opportunistic infections, disease and malignancy.

Clinical Latency Period:• HIV continues to reproduce, CD4 count gradually

declines from its normal value of 500-1200.• Once CD4 count drops below 500, HIV infected

person at risk for opportunistic infections.• The following diseases are predictive of the

progression to AIDS:– persistent herpes-zoster infection (shingles)– oral candidiasis (thrush)– oral hairy leukoplakia– Kaposi’s sarcoma (KS)

AIDS• CD4 count drops below 200 person is considered to have

advanced HIV disease• If preventative medications not started the HIV infected

person is now at risk for:– Pneumocystis carinii pneumonia (PCP)– cryptococcal meningitis– toxoplasmosis

• If CD4 count drops below 50:– Mycobacterium avium– Cytomegalovirus infections– lymphoma– dementia– Most deaths occur with CD4 counts below 50.

Anti- HIV Drug Targets

Three types of drugs are currently in clinical use:

1. nucleoside and nucleotide reverse transcriptase (RT) inhibitors

2. non-nucleoside reverse transcriptase inhibitors

3. protease inhibitors (PIs)

Nucleoside and Nucleotide Analogs

• Nucleoside analogs (NRTI) act as chain terminators or inhibitors at the substrate binding site of RT

• NRTI’s must be phosphorylated (three steps) to their 5’-triphosphate form to become active inhibitors.

• Nucleotide analogs (NtRTI) already contain a phosphate group and only go through 2 steps to become active.

• The 5’-triphosphate of the NRTI’s compete with the 2’-deoxynucleoside’s 5’-triphosphate for binding to reverse transcriptase leading to viral DNA chain termination.

Nucleoside Analogs

• There are currently 7 FDA-approved NRTI’s and one nucleotide analog.

• The first anti-HIV drug approved was the NRTI known as AZT or Zidovudine (1987).

• AZT was discovered as a treatment of AIDS during a screening process for the identification of effective AIDS treatments4.

• Antiviral selectivity due to higher affinity for HIV RT than human DNA polymerases.

Non-Nucleoside Analogs

• Non-nucleoside analog reverse transcriptase inhibitors (NNRTI’s) inhibit viral DNA replication by binding at the allosteric non-bonding site of RT, causing a conformational change of the active site.

• NNRTI’s do not require bioactivation by kinases. • Three NNRTI’s are currently approved for clinical use

in combination therapy: nevirapine, delavirdine, and efavirenz

Non-Nucleoside Analogs

Delavirdine

Benzoxazinone

Nevirapine

Protease Inhibitors• During the reproduction cycle of HIV a specific

protease is needed to process GAG and POL polyproteins into mature HIV components.

• If protease is missing noninfectious HIV is produced.

• HIV protease inhibitors are specific to HIV protease because it differs significantly from human protease.

• The 6 PI’s currently approved for clinical use were all designed by using structure-based drug design methods.

FYI: Structural Genes

• Three main structural genes:– Group Specific Antigen (Gag)– Envelope (Env)– Polymerase (Pol)

Group Specific Antigen (Gag)

• Located in nucelocapsid of virus.• Icosahedryl capsid surrounds the internal

nucleic acids made up of p24 andp15.• p17 lies between protein core and envelope

and is embedded in the internal portion of the envelope.

• Two additional p55 products, p7 and p9, are nucleic acid binding proteins closely associated with the RNA.

HIV Protease

• The crystal structure of HIV protease was first obtained at Merck Laboratories.

• HIV protease is a 99 amino acid aspartyl protease that functions as a homodimer with one active site.

• The active sites of protease are hydrophobic.

Protease Inhibitors

• HIV PI’s target the peptide linkages in the gag and gag-pol polyproteins which must be cleaved by protease.

• All approved PI’s contain a hydroxyethylene bond instead of a normal peptide bond.

• The hydroxyethylene bond makes PI’s non-scissile substrate analogs for HIV protease

Protease Inhibitors

• ABT-378 or lopinavir was approved in 2000 for use in combination with ritonavir (a PI) (Kaletra)

• Ritonavir strongly inhibits the metabolism of ABT-378

Some Alternative Therapies • Virus adsorption inhibitors – interfere with

virus binding to cell surface by shielding the positively charged sites on the gp-120 glycoprotein– Polyanionic compounds

• Viral coreceptor antagonists – compete for binding at the CXCR4 (X4) and CCR5 (R5) coreceptors– bicyclams and ligands

Virus Adsorption Inhibitors

• Cosalane was originally developed as an anti-cancer agent by researchers at Purdue University and the U.S. National Cancer Institute8.

• Cosalane was developed from a chemical known as ATA (aurintricarboxylic acid), which has long been known to have anti-HIV activity8.

• ATA is a mixture of different polymers. Chemists took one of the low molecular weight components of ATA, and attached it to a steroid molecule in order to target the substance more effectively to the surface of viruses and of cells.

• The result was cosalane.• Cosalane binds to the HIV gp-120

protein.

Viral Coreceptor Antagonists • Bicyclams are a type of viral

coreceptor antagonist.• They are very specific and potent

X4 coreceptor antagonists.• Bicyclams belong to a

class of macrocyclic polyamines consisting of two cyclam units linked by an aliphatic bridge

• Bicyclams with an aromatic linker apparently had higher antiviral activity10.

• One such compound is AMD3100.

Combination Therapy

• Monotherapy created virus resistance to the individual drug. Some combination therapies increase the time it takes for the virus to become resistant.

• Combinations of a PI or NNRTI with one or two NRTI’s is often recommended.

• Combination therapy may reduce individual drug toxicity by lowering the dosage of each drug

Combination Therapy

• The combination of drugs chosen is based on the history of each individual patient and synergistic drug interactions.

• Some drugs compete with each other for binding sites or enzymes.– Example: zidovudine and stavudine • both nucleoside analogs compete for the same

kinase. Stavudine is not phosphorylated because zidovudine is preferred

Combination Therapy and Drug Resistance

• Some drug combinations can restore sensitivity of the virus to drugs it was previously resistant to.– Example: lamivudine and zidovudine• The HIV M184V mutation is resistant to lamivudine but

restores sensitivity to zidovudine resistant virus mutants

Drug Toxicity and Side Effects

• All available antiretroviral drugs are toxic.• Side effects of nucleoside analogs are lactic

acidosis and severe hepatomegaly with steatosis (enlarged fatty liver).

• Other side effects of anti-HIV drugs include pancreatitis, myopathy, anemia, peripheral neuropathy, nausea, and diarrhea.

Reducing Drug Toxicity

• The use of combination therapy:– Combining agents with favorable synergistic

properties allows a decrease in dose or dosing frequency

– Ritonavir alone cause gastrointestinal side effects but when used in combination with other PI’s it can be administered at a lower dose.

Conclusions

• An effective anti-HIV therapy is still needed.• Several possible targets are being studied and

tested.• The area of anti-HIV drugs has more room for

growth and the future for the discovery of new effective drugs is promising.