HIV Testing and Treatment, March 2002

Table of Contents

Prevention

Testing

Fundamental Notions of Testing

Screening Tests

Confirmatory Tests

Treatment

Approved Drugs

Not Yet Approved Drugs

Other Drug Notions

Objectives

Footnotes have been enclosed in square brackets and colored green. Students need to know the terms in blue.

Prevention

Surely, an ounce of prevention is worth a pound of cure (especially when there is no cure!). We know that HIV is transmitted by: (a) transfusion of HIV contaminated blood products, (b) sharing of intravenous drug injection apparatus with an infected user, (c) having unprotected sex with an infected person, or (d) from an infected mother to her newborn child either during pregnancy/delivery or via breast feeding.

Currently, blood products in this country are screened for HIV and other pathogens. The best estimates for risk are that one in 450,000 units of blood is tainted with HIV. That's only 0.0002% likelihood of infection (much less than dying in an automobile crash). A recent proposal (2/2001) recommends that all whole blood be subjected to leukoreduction, i.e. removal of all white blood cells. This would greatly reduce the likelihood of transmission and also reduce deleterious immune responses to the transfused blood.

Properly used condoms are estimated to be 98% effective. That means a 2% likelihood of contact with the infectious agent. Using a 0.30% probability of transmission for unprotected sex, this reflects a transmission rate of 0.006% during protected sex. This translates to odds of transmission of about 1 in 16,000.

Drug "works" can be sterilized using a simple bleach solution. Nevertheless, sharing needles is a very bad idea. Infected blood in a syringe can harbor viable virus for up to seven days.

Current thinking is being directed toward further improving the safety of sex by the use of microbicides. Several options are undergoing testing.

(1)   Dextrin sulfate seems to be an effective vaginal microbicide that does not disrupt genital epithelium  and it is not systemically absorbed.

(2)   An extract of blue-green algae, called cyanovirin-N is undergoing tests and seems promising.

(3)   A thermoreversible gel containing sodium lauryl sulfate, bearing the commercial name of Invisible Condom, seems to be effective against HIV and other sexually transmitted pathogens.

None of these products are currently FDA-approved for to prevent transmission of HIV. In vitro research published in August 2001 has shown that there are other commercially available candidates: Astroglide, Vagisil, and ViAmor

The June 2000 issue of the Journal of Virology carried an article by R.J. Shattock and colleagues from St. George's Hospital Medical School, in London. They analyzed tissue from the cervix and vagina of women who had hysterectomies. They found that HIV infects macrophages below the epithelium. Furthermore, this was enhanced by immune activation. Epithelial cells were not infected and did not seem to have a role in transferring the virus across the epithelium. Of course, any trauma that could damage the epithelium would enhance transmission. The group tested various virucides that can be applied topically. Nonoxynol-9, gramicidin, and PRO-2000 blocked HIV infection by 30%, 71%, and 97%, respectively. If these results hold up to further scrutiny, women will have an option, over which they have control, which may have a significant impact on transmission.

Research presented during the XIII^th International AIDS Conference held in Durban, South Africa (2000) indicated that the spermicide nonoxynol-9 not only does not prevent infection by HIV, but may very well enhance it. Although the infection rate was higher in this treatment group, it was not statistically significant. Other studies have reported disruption of the genital epithelium by nonoxynol-9.

Testing

If you were going to be tested for HIV, there are several steps to follow. If you are not someone who engaged in risky behaviors, e.g., unprotected sex with multiple partners, intravenous drug use sharing needles, etc., then you should take a screening test. If you test negative, then that would be that. On the other hand, if you had engaged in risk behaviors and tested negative, you would be advised to (a) come back for another test in six months to one year and (b) change your behaviors. Someone at risk who tests positive would be urged to take a second screening test. If that test were positive, then a confirmatory test would be given. In all cases and for all people, testing should be preceded and followed by counseling. It is imperative that the testee be given all the facts associated with her/his test and test results.

Testing Modalities

Anyone who is to undergo a medical procedure must give his or her informed consent. Informed consent consists of:

1. Full and complete disclosure of all relevant information to the patient.
2. The patient must be legally competent to make an informed decision.
3. The patient must understand everything that was explained.
4. The patient's decision must be purely voluntary and free of all duress.
5. The patient must make the decision whether to undergo the procedure or not.

More than half of the states allow testing without consent, but only if certain preconditions are met. Some of the conditions are:

(a)   patient or her/his designee is unable to either give or withhold consent,

(b)   the test results are essential to determine the form of treatment,

(c)    the test is needed to protect the health of health care personnel or others,

(d)   a minor patient has parental consent.

There are several forms of testing.

1. Voluntary testing: the patient agrees to a test where the results will be known to the patient and the tester, who is under no obligation to withhold the results.
2. Voluntary confidential testing: the patient agrees to a test where the results will be known to the patient and to the tester, who cannot divulge the information to anyone else.
3. Voluntary anonymous testing: the patient agrees to a test where the results will be known only to the patient. The tester knows the patient only as a code number.
4. Mandatory testing: the patient is required to be tested as a precondition for participation in some activity, e.g., giving blood. This may be anonymous, confidential, or neither. Participation in the activity is at the patient's option. The patient may choose to forgo the activity to avoid the test.
5. Compulsory testing: the patient must submit to be tested irrespective of her/his wishes, e.g., in some states people convicted of prostitution are ordered to be tested.

 

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Fundamental Notions of Testing

According to the CDC, in 1996 somewhere between 2 and 2.5 million HIV antibody tests were administered in publicly funded programs. Of these fully 60% were retests and the remaining 40% were tests given for the first time. That was the good news. The bad news is that 26% of those testing HIV+ and 33% of those testing HIV- never returned for their results. That leaves quite a few people walking the streets that don't know their HIV status, but should.

At least 70% of the HIV+ people in the developed countries know their HIV status. On the other hand, at least 90% of the HIV+ people in the developing countries do not know their HIV status. The incidence rates reflect these figures!

No form of testing is perfect. Every test can introduce both false positives (testing positive when you are really negative) and false negatives (testing negative when you are really positive). It is essentially impossible to construct a test procedure for which there are neither false positives nor false negatives in all testing situations. Instead, one designs a test either for screening or confirmatory validation. A screening test must catch as many positives as possible, even at the expense of a large number of false positives, whereas a confirmatory test should rarely give a false negative. For these reasons, tests are characterized by their sensitivity and specificity, together with their positive predictive value as defined below.

Sensitivity = probability of testing + when you are HIV+

Specificity = probability of testing - when you are HIV-

Positive predictive value = probability of being HIV+ when you test +

Negative predictive value = probability of being HIV- when you test -

Clearly, the number of interest to someone being tested is positive predictive value (PPV). It tells you how likely you are of being positive once you test positive. A high PPV assures you that the results of the test are reliable, while a low PPV indicates the need for further testing. Some comparative values of these parameters indicate their relationship to each other when we assume that the subpopulation to which the testee belongs has an HIV prevalence rate of 0.5% (that means about 1 in 200 people in the population are HIV+).

 

When sensitivity is .995, specificity is .995, and the incidence rate of HIV is 0.4% (not at all unreasonable numbers), then the results of testing 100,000 people would break down as follows. We classify whether they test positive or not and whether they are positive or not.

 

Notice that there are more false positives than true positives for this (high) caliber test. The idea behind a screening test is to be sure that as few positives as possible get through undetected. [Similar results come out of drug screening tests. The thought of being wrongfully branded a drug user by an overly sensitive test is a constant and vexing problem.]

Viewed from another perspective, suppose we have a test, which has a sensitivity of 0.999 and a specificity of 0.996 (a really high quality test). The PPV as a function of the prevalence rate for the subpopulation to which the testee belongs is given below.

 

It should be clear from this table that the riskier your behaviors, the more likely that a positive screening test which says you are positive is giving you a correct result.

 

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Screening Tests

There are several types of HIV-1 tests: enzyme-linked immunoassay (ELISA), enzyme immunoassay (EIA), various types of blot tests, immunofluorescent tests, and viral load [also called viremia] tests that measure the number of copies of viral RNA present in the blood. The enzyme-based tests, which test for HIV antibody, have high sensitivity and only moderate to high specificity. The blot tests search for many types of proteins (and there is not a uniform standard as to what constitutes a positive test) and have lower sensitivity, but high specificity. The RNA tests are highly sensitive and highly specific. There are also tests for both the p24 and p31 antigens which are characteristic of HIV. Each of these tests usually requires more than a day to complete at a laboratory, so that the testee must return to get her/his results.

Bear in mind that upon infection with any pathogen, there is a lag time (the "window period") before antibody is produced. For HIV, that could be as long as several months. The process of generating detectable antibody is referred to as seroconversion. A person who has seroconverted to HIV+ status is said to be seropositive. Only after seroconversion will any antibody test show a positive result (other than a false positive). No matter what test is used, a person who has engaged in risk behaviors who tests negative on both a screening and a confirmatory test should be retested in six months to one year.

Couples of the same serostatus are said to be seroconcordant. Couples with differing serostatuses are serodiscordant.

False positives to antibody tests may occur at disproportionately high rates in

• injection drug users;
• people who have recently been given injections of gamma globulin;
• those recently immunized against influenza or hepatitis B;
• multiparous females (that's medicalese for women who have had several children);
• people with liver disease;
• people who have had autoimmune disorders, rheumatological diseases, DNA viral infections, various cancers, leprosy, malaria, infection with Epstein-Barr virus, or alcohol induced hepatitis.

A rare case of a false negative viral load test has been recorded for an HIV+ man with a simultaneous cytomegaloviral infection.

The various EIA and ELISA tests can use different reagents: carcinogenic antigen (CEA), sex hormone binding globulin (SHBG), h-insulin-like growth factor binding protein-1 (IGFBP-1), luteinizing hormone (h-LH), thyroid stimulating hormone (hTSH), prolactin, or human chorionic gonadotrophin (hCG). Most of these cost around $15–50. Another test of this form is OraSure which does not use blood but rather oral fluids, saliva and transmucosal fluid, and it can serve as both a screening and a confirmatory test depending on the reagents used. There is also a urine test available, the Seradyn Sentinel.

To improve the return rate for screening testing and cut down on the number of people who test positive but never learn their test results, several companies have developed rapid screening tests. The only such test currently FDA-approved is the Single Use Diagnostic System for HIV-1. Many of the rapid tests return results in anywhere from 10 minutes to one hour. Preliminary data indicate that their sensitivity and specificity are as good as the regular enzyme antibody tests. Hopefully, their rapid introduction will decrease the number of HIV+ people who do not know their HIV status, thus increasing the use of safer sex precautions and resulting in a subsequent decrease of the incidence rate.

On May 10, 2000 Calypte Biomedical Corp. announced the formation of Sentinel Testing. This will involve a rapid turn-around time (3 days after receipt at their lab facilities) for testing of HIV-1 antibody, chlamydia DNA, and gonorrhea DNA using urine!

On June 8, 2000 Bio-Rad Labs announced a new HIV assay that can detect the virus earlier than other tests. The company's Genscreen Plus HIV Ag/Ab kit detects both antibody and antigen in a blood sample with a sensitivity of 99.9%. The test can detect all forms of HIV-1 group M and Group O plus all forms of HIV-2. Although approved in France, the company has yet to apply for FDA approval.

As of April 2000, there were 21 commercial HIV-1 antibody tests, 3 commercial HIV-1,2 antibody tests, 1 commercial HIV-2 antibody test, 5 HIV-1 antigen tests, and 1 HIV RNA test that have full FDA approval. A full listing is available at the FDA website.

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Confirmatory Tests

Confirmatory tests have been of either the Western Blot or immunofluorescent types.

Blot testing screens for a large number of HIV proteins. HIV is cultured in human cells, which are then ground up and placed on a gel. An electric current is applied and the lightest proteins move the farthest and the heaviest move the least. The result is a series of narrow bands corresponding to the protein weights. Bands corresponding to p17/18 [This means that p17 and p18 cannot be easily distinguished], p24/25, p31/32, gp41, p51/52/53, p55, p64/65/66, gp110/120, and gp160 can be detected. Sometimes indeterminate results are obtained at various bands. Additionally, testing methods, reagents, and test criteria are not standardized. Various companies and organizations define a positive Western Blot test differently. [The most conservative approach is to define a positive as showing determinate results for p17, p24, gp41, gp120, and gp160] Costs are typically $150+ and the time required is ordinarily from 12 to 24 hours.

The immunofluorescent tests can detect either antigen or antibody. A control preparation of antibodies labeled with a fluorescent dye is added to a sample. This test has the added advantages of taking about 90 minutes to do in a doctor's office and has no indeterminate outcomes.

Not all HIV-1 tests will respond properly to either type O HIV-1 infections or HIV-2 infections. Special tests have been designed for patients suspected of having these conditions.

The viral load [measured in virions per mL. One mL is the same as 1000 mm³] tests, which detect proviral DNA (from which the RNA numbers can be easily inferred), currently available are: Amplicor, which uses PCR [This stands for Polymerase Chain Reaction whereby the RNA is primed to DNA, which is then split and cloned. This is repeated as many times as needed to obtain the desired accuracy. Ten repeats gives 2^-10=1/1024 accuracy. It is normal to use 25 repeat cycles. Because the automated method is not 100% efficient, this usually produces only a three million-fold increase as opposed to the 2²⁵ = 33,554,432-fold theoretical increase.]; Quantiplex, which is a banded DNA testing method; and nucleic acid sequence based amplification (NASBA) of HIV-RNA. These are a bit pricier at about $160+. Only the Amplicor test is FDA-approved for general use, although many large-scale research groups use the Quantiplex test. Both the dynamic range and lowest detectable level have been improving. The range of standard viral load tests can go from 400 to 50,000 copies per mL. The lowest currently detectable viral load is 20 copies in so-called ultrasensitive screening, but accuracy at this level is unknown and variability below 400 may be as high as a factor of five! For advanced AIDS cases, tests are frequently "detuned" to provide an extended dynamic range above 50,000 copies.

On February 5, 2001, BioTech Imaging Inc. announced a new test for cells that are actively producing HIV. The gp120 Tagger Cell Expression Kit is currently undergoing testing. If successful, this test would shed an entirely different light on the production and number of virions in the body as the current PCR Amplicor test counts only viral RNA in the peripheral blood.

Viral load tests are not designed as screening tests even though they have high sensitivity. A team reported in the Annals of Internal Medicine for January 5, 1999 of three misdiagnoses of HIV by these tests (in addition to one other such case). Nevertheless, that remains a very low false positive rate.

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Treatment

Common illnesses are treated by an aggressive induction therapy designed to lower the number and virulence of pathogens in the body. Once the reproduction rate of the pathogen is below a predetermined threshold, the dosages of drugs are lowered to levels of so-called maintenance therapy designed to keep the level of the pathogen low and manageable by the immune system. If the infection resurges, then a change in medication is called for and salvage therapy begins.

If an HIV patient develops an opportunistic infection, it must be treated immediately and aggressively. In fact, when the CD4 markers fall low enough to indicate the possibility of such an infection, medication in prophylactic dosages is usually given as a preventive measure. Thus, patients are often taking medications when they have no symptoms of disease other than HIV disease.

A clinical marker of infection by HIV has been the CD4+ T cell count, which has now been displaced by the viral load. CD4 counts are not very predictive of the onset of opportunistic infections and frank AIDS [Remember, the CDC defines AIDS among HIV+ patients as having a CD4 count under 200 and/or a specific opportunistic infection such as PCP, wasting syndrome, KS, etc. when you are HIV+], but the viral load is a fairly good predictor. Therefore, people undergoing treatment are given frequent viral load tests. A viral load below 10,000 is considered to be stable. Viral loads between 10,000 and 100,000 mean there is a risk of disease progression and/or opportunistic infections. Viral loads in excess of 100,000 indicate an increased risk of progression to AIDS [Early after infection, when the viral load is maximum and the patient is most infectious, counts in the millions are not uncommon.]. Oddly enough, women tend to develop AIDS at lower viral loads than men, but equally strangely, women have a longer life expectancy once infected.

HIV uses three enzymes to reproduce: reverse transcriptase, protease, and integrase. The current crop of approved drugs attacks only the first two of these. There are currently three approved (+2 experimental) classes of anti-HIV drugs;

1. Nucleoside analogue reverse transcriptase inhibitors (NRTIs or "nukes" in the vernacular), which block the action of the enzyme reverse transcriptase and prevent the virus from making proviral DNA,
2. Non-nucleoside analogue reverse transcriptase inhibitors (NNRTIs or non-nukes), which bind directly to reverse transcriptase to prevent its action, and
3. Protease inhibitors (PIs), which block the action of the enzyme protease which is used by HIV to cut other proteins into shorter, more useful pieces for reassembly into new virions.

There is a new class of experimental drugs called pyrophosphate analogues, which also block the action of reverse transcriptase. None of the drugs in this class have yet received FDA approval. Coreceptor blockers are also being studied. Clinical trials are ongoing for two new fusion inhibitors.

Monotherapy, the use of a single drug, is strongly advised against because of the very strong (almost certain) possibility of the development of resistance to the drug by the virus. It is recommended that patients take at least a three-drug "cocktail", possibly including either one protease inhibitor or the NNRTI efavirenz (Sustiva). This is usually called highly active antiretroviral therapy or HAART. These treatments can drop the viral load by three and four logs [A log is factor of 10. Thus, two logs is a factor of 100, three logs a factor of 1000, and four logs a factor of 10,000.]. Unfortunately, HIV has managed to develop mutations that resist many of these drugs. When adverse side effects dictate a lowered dosage, the viral load can rise significantly, and drug resistance can increase. Then the drugs may have little effect on the HIV, and an alternative drug regimen is required.

There are three types of drug resistance that can develop:

·        genotype resistance, the virion mutates in the presence of the drug,

·        phenotype resistance, there is an decrease in sensitivity of the virus to the drug, and

·        cross-resistance, the virion develops a resistance to many drugs within the same class.

The administration of ZDV orally during pregnancy, intravenously during delivery, and to the baby for a short time after delivery can significantly reduce the transmission rate from a high of 16–20% down to 6–8%. An experiment on Thai women (see the March 6, 1999 issue of the Lancet) indicates that a shorter regime of ZDV is nearly as effective. Other research (8/99) done in Africa indicates that a much shorter course of nevirapine may be equally effective and significantly cheaper ($5 versus $75).

A study released on October 25, 1999 tested the trophoblastic outer layer of the placentas of several positive women. No trace of HIV RNA was found, indicating that except in unusual circumstances, HIV is not transmitted to the fetus during pregnancy. More likely is a transmission slightly before or during the birth process.

Many patients receiving HAART do not initially respond as well as they should and require changes in their treatment. In fact, 15% of those given PIs must change medications because of the side effects. An additional 15% do not respond to PIs at all. Recent numbers indicate that many patients who have been previously successfully treated using PIs are no longer responding to the drugs.

The optimal or durable response is to lower viral load below detection by ultrasensitive test, i.e., < 40 copies/mL and raise the CD4 count. Factors that decrease the chances of a durable response are: (1) low baseline CD4 count, (2) high initial viral load, (3) prior long-term use of a NRTI or PI, and (4) poor treatment adherence. On the other side of the coin, there are no reliable indicators that can assure when treatment will succeed.

On February 4, 2001 a revision of the treatment recommendations was released. Rather than the "hit early and hit hard" philosophy, federal health officials are recommending delaying treatment as long as possible. [CD4+ count below 350 and viral load above 55,000 using PCR or 30,000 using branched-DNA] The intent is to prevent the cumulative toxicity that can cause major physiological changes in the body.

 

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Approved Drugs

The table below lists the currently (6/2001) FDA-approved drugs. The abbreviations used are bid = bis in die = twice per day and tid = ter in die = three times per day.

 

 

On January 31, 2000 Bristol-Myers Squibb applied to the FDA for approval of a new formulation of Videx (ddI) that could be used for once daily dosing. Following three reports of fatalities, on January 5, 2001 the company sent a letter to practitioners warning of the possible danger of lactic acidosis for pregnant women taking either stavudine (d4T) or didanosine (ddI).

Fortovase should be stored in a refrigerator. Crixivan does not tolerate humidity well, so it must be stored in a closed container with a desiccant. Ritonavir tablets must be kept refrigerated, but the (absolutely horrid tasting) liquid can go without refrigeration for up to 30 days. The August 1998 issue of Clinical Drug Investigation reports on cases of acute renal failure and dysfunction associated with ritonavir.

A 1998 study from Australia indicates that 18–76% of the people on protease inhibitors will develop peripheral lipodystrophy, wherein they will lose subcutaneous fat from their extremities and add visceral fat to the trunk, particularly the breasts, the abdomen (called a "Crix belly" or "protease paunch"), or on the back between the shoulder blades (called a "Buffalo hump"). The Buffalo hump can be such a problem for sleeping and cardiac involvement that liposuction and/or surgery may be required. Other metabolic disorders are concomitant with antiretroviral therapy. The study also reported that 16% of the subjects had impaired glucose tolerance, which can lead to diabetes. These metabolic abnormalities have not been fully characterized, not are they restricted to the use of protease inhibitors. Cases have been reported for people on other anti-HIV drugs too.

The 7^th Conference on Retroviruses and Opportunistic Infections (held 1/30–2/1/2000) had a large number of papers dealing with this issue. A suggestion of Grunfeld was to call the syndrome FRAM, fat redistribution and metabolism. This is reasonable because there are three separate problems: peripheral lipodystrophy, truncal accumulation of visceral fat, and various metabolic disorders. FRAM has been tied to mitochondrial toxicity of the antiretroviral medications.

The following pictures are taken from the October 29, 1998 issue of the New England Journal of Medicine's section Images in Medicine.

 

What you see is someone whose lipid (fat) levels in the arms and legs has dropped precipitously, while s/he has developed fatty deposits around her/his trunk and abdominal organs. This is not the usual subcutaneous fat that lies below the skin and above the muscles, but rather visceral fat that lies directly around the vital organs. Some people have a small fat pad on the back of their neck, but the man below has a much more pronounced accumulation. Normal activities like sleeping on one's back or holding the head erect are significantly impaired by such a protrusion. Sometimes surgery is used to remedy the situation.

 

Reports beginning in September and October of 1999 tied antiretroviral therapy (nukes, non-nukes, and PIs) to increases in both cholesterol and triglyceride levels. These increases have been associated with cardiac problems, including heart attacks.

The drugs ddI and/or d4T have been given in combination with hydroxyurea, which enhances their effect, mitigates the effects of drug resistance, and allows somewhat reduced dosages with a concomitant reduction in side effects. Hydroxyurea also inhibits the production of HIV in macrophages, quiescent lymphocytes, and dendritic cells. Another drug, resveratrol, seems to have similar effects as hydroxyurea, but with fewer adverse reactions.

Efavirenz can cross the blood-brain barrier and has been shown to reduce the viral load in the cerebrospinal fluid, CSF. This raises some hope that there is a possibility that AIDS dementia can be arrested and some studies have shown this to be the case.

Abacavir can lead to such severe hypersensitivity reactions that death can result. A case study of a near fatality is available in The AIDS Reader 11(4):222-226. 2001. Once abacavir is discontinued it should never be restarted because of a possible hypersensitivity reaction which can be life threatening. One death due to rechallenge with abacavir has been reported in France. Abacavir can also cross the blood-brain barrier.

A finding by Boucher, et al. published in the December 3, 1999 issue of the journal AIDS found that suboptimal doses of an antiretroviral drug can lead to HIV mutations that can replicate better than the wild type virus. They studied a single patient who had been treated only with the protease inhibitor ritonavir. Two phases of evolution of the virus were described: in the first phase the HIV changed in only one or two nucleotides and the drug seemed to be doing what it was intended to do. In the second phase there was a five or six nucleotide difference from the wild type virus. This new form did not show increased drug resistance, but it did show an increased efficiency of protease and a concomitant increase in the replicative capacity of the virus. Thus the fitness of the virus improved as a stronger variant was selected for by these changes.

 

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Not Yet Approved Drugs

The following drugs have not yet received full FDA approval, but some of the noncoded ones are expected to be approved within a few months:

 

The November 1998 issue of Nature Medicine carried an article by Kilby, et al. describing a clinical trial of a fusion inhibitor T-20 that reduced viral loads an average of 1.96 logs (almost a factor of 91) in fourteen days. The drug is a synthetic peptide corresponding to a region of the transmembrane subunit of the HIV-1 envelope protein, thus blocking cell fusion. It must be taken either intravenously or subcutaneously. The drug is currently undergoing Phase III clinical trials. The company has also proposed another candidate, T-1249, as a possible fusion inhibitor. Positive preliminary results of the success of T-20 were announced on February 2, 2000. During May of 2001, the company (Trimeris) applied to the FDA for approval. In November of 2001, the drug went on an open-label clinical trial. Full approval and distribution is expected by 2003.

GlaxoSmithKline has an integrase inhibitor S-1360 currently in Phase II clinical trials.

Although lodenosine was previously listed as an unapproved drug, due to severe adverse reactions and one patient death, the Phase II clinical trials involving 176 people were officially put on hold on October 15, 1999. Pending reassessment, no further testing will occur.

On November 1, 1999 the FDA denied accelerated approval to adefovir dipivoxil. Kidney damage was associated with the drug during testing. The manufacturer has withdrawn it as a potential anti-HIV drug, but will continue the drug's development as a hepatitis B treatment. Although no new patients will be enrolled in the clinical trials or expanded access programs, patients in the trials will be allowed to enroll in the expanded access program so long as they benefit from treatment.

Gilead Sciences announced the expansion of access to its drug tenofovir DF on January 1, 2001. Patients would take one 300 mg pill once a day. Expanded access, as part of the Phase III clinical trials, will likely speed FDA approval.

Pfizer announced on 1/26/2001 that it was halting clinical trials of its promising drug capravine. Experiments had shown an increased incidence of vasculitis (inflamed blood vessels) in dogs. None of 650 human subjects participating in the trials had shown any symptoms of vasculitis.

Green, Stevenson, et al. authored an article published in the December 9, 1999 issue of Nature wherein they described the mechanism by which so-called matrix protein enhances the flow of HIV into the nucleus as the virus commandeers the cell's genetic machinery for the purposes of producing copies of the virus' own genetic material. Once this is complete, it abruptly reverses itself and directs the constructed viral RNA outward from the nucleus. This details two more steps in the infection process that could possibly be interrupted by appropriate drugs. On January 28, 2000, the pharmaceutical giant Merck announced the discovery of two strand transfer inhibiting drugs via chemical screening, L-731,988 and L-708,906. The drugs were shown to be effective integrase inhibitors in vitro against several different clades, M-tropic and T-tropic, clinical isolates, and strains of the virus resistant to both reverse transcriptase and protease inhibitors. Strains that were resistant to the compounds were also found. The drugs are but two in the class of diketo acids, about which more needs to be studied. This is a long way from another useful drug, but it does open new possibilities where far too few are now present.

On January 11, 2001 ScienceExpress carried an announcement of a new protein 5-Helix as a possible injectible drug for salvage therapy when other drugs fail. 5-Helix is designed to bind to gp41 and prevent viral fusion. It is also possible this mechanism can be employed to defeat other viruses like influenza or Ebola.

Other classes of drugs being researched include: integrase inhibitors, DC-SIGN inhibitors, regulating protein inhibitors, uncoating protein inhibitors, RNAase H protein inhibitors, zinc finger inhibitors, capsid protein polymerization inhibitors, assembly protein inhibitors, and Tsog protein inhibitors.

Another significant problem is the interaction of various drugs (not just anti-HIV drugs). Some work together to enhance their effects and others retard their effects. The number of possible combinations is quite large and great care is needed to avoid worsening side effects and/or weakening the antiretroviral activity. Currently software is available to check on this. There are also interaction effects between drugs and common foods, most notably grapefruit juice.

Some explanation of what the various clinical trials are is in order. During Phase I trials, human subjects who do not have the disease in question are given the drug to determine any major side effects. Phase II trials are usually double blind studies of the drug versus a control. Ethical issues arise here because in this country the control would be chosen from one of the currently used drugs. Some companies have been the recipients of severe criticism for using non-American control groups that received no drugs. In the final Phase III trials, everyone is given the drug but separate groups receive different dosages to find the optimal level. At all levels certain statistical rules must be followed. One of the most important is the randomization of subjects among groups.

 

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Other Drug Notions

Drugs in the body react and decay to the point where they are no longer effective. A measure of this decay is the half-life. A drug with a half-life of one hour will have only 50% of its maximum activity after one hour, 25% (= (1/2)²) after two hours, 12.5% (=(1/2)³) after three hours, and so on. The longer the half-life of a drug, the less often it needs to be taken. Zidovudine has a half-life of about an hour, while efavirenz has a half-life of seven to eight hours thus allowing a twice-daily dosing schedule. Some experimental drugs have a half-life exceeding 15 hours, so that once daily dosing may be in sight (see previous comments about tenofovir DF).

The ultimate goal of any dosing schedule should be the maintenance of a constant therapeutic level of the drug. In actuality, the level rises and falls. The lowest level is called the trough level. The trough must be high enough to be therapeutic, lest the virus mutate and develop resistance to the drug. The maximum level should not be so high that side effects become overwhelming.

Drug resistance is a particularly problematic for the treatment of HIV disease because of the small number of available drugs. Viral resistance occurs in 30–50% of the patients on HAART. Once a patient shows a resistance to an NNRTI, it is very likely there will be cross-resistance to all NNRTIs. The Swiss cohort study assigned people on HAART to four groups according to their viral load and CD4 count.

• 40% of the subjects showed a decreasing viral load and an increasing CD4 count (that's very good);
• 40% showed ongoing viral replication and an increasing CD4 count (fair);
• 5% showed viral suppression with a steady low CD4 count (also fair); and
• 15% were treatment failures, showing high viremia and low CD4 counts (not good).

Study of the mutations has proceeded apace, insofar as it is now known at what codons the mutations for most of the individual drugs occur. A rather expensive test for drug resistance was announced in June 1998, but it may be of questionable value for other than determining which drugs not to give. For instance, a single mutation of the RTI gene of HIV confers a high degree of resistance to 3TC and/or nevirapine, whereas multiple mutations of the protease gene are required for resistance to protease inhibitors.

The difficulty of maintaining a HAART regimen is hard to imagine. An anonymous letter to the editors of POZ magazine (accessible in part at theBody) published in July 1998 brings a few of the problems to light.

 The portrayal of the HIV positive physician's assistant, Jeanie Boulet [no longer part of the program], on the TV show ER is very unrealistic. She is on Crixivan, the protease inhibitor hardest to take, but has only ever taken one dose on the show. A more realistic picture of Jeanie's day: Wake up by 5 am. Take Crixivan, take a shower, withstand nausea for at least one hour, take other meds at 6 am. Eat breakfast. Go to work around 7 am. Once there, spend a lot of time at the water fountain or carry around a bottle of water. Lunch at 11 am (barring any emergencies), then take other meds and finish by noon so that at 2 pm she can take her next dose of Crixivan. Jeanie had better hope none of the good-looking residents asks her out to lunch, as she'd have to explain that she has to be finished eating by noon or her dosing for the day will be messed up. The same goes for any dinner dates. When not drinking water, eating meals or taking her reverse transcriptase inhibitors or Crixivan, Jeanie would be on her way to the rest room. Where would she find time to see patients?

The number of pills one needs to take can be absolutely overwhelming. 12 on the low side to a high of around 60, for those with multiple opportunistic infections. That's about a small plastic lunch bag filled with medication. Bear in mind, these pills must be timed with respect to meals, feelings of major nausea, and each other. Some must be taken with a high fat meal. By "high fat meal," they mean a minimum of 28 grams of fat. That's about half of a normal person's daily allowance of fat! Despite all these caveats, when it works, HAART can drive viremia down to undetectable levels, < 400 copies/mL. It can even force the viral load below the threshold of the ultra-sensitive tests, <40 copies/mL.

 

A paper presented at the 12^th AIDS Conference in Geneva, Switzerland on July 1, 1998 reported on the detailed analysis of T cells of patients on highly active antiretroviral therapy, HAART, whose CD4 count rose significantly. The bad news is that this increased count consisted mostly of severely damaged T cells that did not function to their full potential. Only for children (they have fully functioning thymuses) were the T cells fully functional. This somewhat dashes hopes of reviving the immune system for HIV patients by using the current crop of drugs. Even worse news was announced on the next day. There was a report of an infection with a multiply drug resistant subtype of HIV-1 in a man who was antiretroviral naïve, i.e., he had never had any antiretroviral therapy. If such an infection can be spread from person to person, it bodes ill for the future of the epidemic when people with a mutated version of the virus are having unprotected sex and spreading their resistant strain. This is as bad as the 1997 report of a variant of Yersinia pestis with a resistant plasmid, but it too was not unexpected. Since July of 1998, there have been very many reports of transmission, including vertical transmission, of these so-called recombinant forms of HIV.

The crucial question remains, is it possible to completely destroy all the HIV in the body by using HAART? Not all the data are in, but a partial answer given in the July 1, 1998 issue of JAMA by Schrager and D'Souza does not paint a rosy picture. They reported on latent cellular and anatomical reservoirs of HIV-1. The virus can enter, gain control of the cellular DNA, and remain somewhat dormant in memory or so-called resting T cells, macrophages, follicular dendritic cells in the lymph nodes, the central nervous system, and possibly the testes. Chun, et al. have determined that these latent sanctuaries are established early in the infection, even before seroconversion. They are seeded with the wild type virus and normally do not contain any mutated strains. At present it is unclear if there are any other latent reservoirs in which HIV can hide. Nor is it known for how long the virus (or provirus) can remain hidden while HAART continues. Some scientists have suggested times on the order of several years. Recent research (May 1999) suggests that for a reservoir of 100,000 virions it could take HAART as long as 60 years to wipe out the virus! Several suggestions have been advanced for "flushing" the reservoirs to accelerate the viral destruction. The announcement of a study by Fauci and his colleagues using HAART together with interleukin-2 to flush out the virus was made on October 13, 1999. Two groups were analyzed; the first received HAART and IL-2, while the other received just HAART. Both groups were taken off HAART and the results were identical. The virus rebounded for both groups. This dealt a critical blow to the concept that the virus can be eliminated from the body using the currently available drugs. Further research announced in February 2000 reinforced this pessimistic view. Work published by Levy, et al. in February 2001 showed a significant positive effect when IL-2 was used together with HAART.