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12, NO. 2. 1992
Anti-Human Immunodeficiency Virus Therapeutics: Now and the Future
Therapeutic modalities to inhibit, inactivate, or interfere with the usual activities of the human immunodeficiency virus (HIV) are best understood in reference to its life cycle. The preceding article by Pavlakis reviews the basic pathophysiology of HIV infection, including the consequences of HIV infection and the immunopathology of the reaction between virus and host. In order to understand the mechanism of anti-HIV directed drugs, this article will first briefly review the major sequences of HIV infection and the HIV life cycle, followed by a description of anti-HIV drugs and other potential therapeutic modalities.
LIFE CYCLE OF THE HUMAN IMMUNODEFICIENCY VIRUS T lymphocytes and monocytes are the major targets for HIV. The HIV virus envelope glycoprotein, gp120, M) to the CD4 binds with high affinity (K,-lop' molecule'^' on target cells. CD4 is a glycoprotein on the surface of mature CD4 (T4) lymphocytes and serves as the primary receptor for the HIV in humans." The CD4 glycoprotein normally facilitates T-helper cell recognition of antigens presented by class I1 major histocompatibility complex (MHC) molecule^.^.^ The process by which HIV enters the cell after it has bound the CD4 receptor is not completely understood; however, once the virus is internalized and uncoated, a single strand of the double-stranded HIV RNA acts as the template for HIV enzyme reverse transcriptase. This enzyme creates a single strand DNA from the viral template. Ribonuclease H, a second HIV enzyme, partially degrades the original HIV RNA template. Reverse transcriptase than acts to create a second DNA strand from the original HIV RNA. The double-stranded HIV DNA is translocated to the host cell nucleus and the HIV DNA duplex is inserted into the host genome by integrase, a third HIV enzyme.' After incorporation into the target cell, a latent or persistent form of HIV infection may develop. The rea-
From the Department of' Medicine. University of California, San Francisco, and the AIDS Program, Sun Francisco General Hospital. Son Francisco, California Reprint requests: Dr. Kahn, Department of Medicine, San Francisco General Hospital, 995 Potrero Ave., San Francisco, CA 941 10.
sons for latency remain unknown, as are the factors that serve to activate HIV after a prolonged dormancy. Many researchers believe that activation of HIV is coupled to the NF-kappa B family of proteins.' Once HIV is active, required regulatory proteins are synthesized. Viral mRNA leaves the nucleus, migrates to the cytoplasm where regulatory protein synthesis is supcrscded by the synthesis of HIV-structural proteins via a complicated feedback mechanism. Structural protein synthesis is required for mature virion production. The structural proteins include the exterior HIV coated proteins and viral enzymes: reverse transcriptase, ribonuclease H, integrase, and an HIV aspartyl protease. HIV core proteins are cleaved from a precursor protein by the HIV-derived protease. Assembly of the infectious HIV virion is accomplished by aggregation of viral core proteins, HIV-I RNA, and enzymes while moving to the surface of the cell. The cores bud, acquire the cell lipid bilayer, and final myristylation of a core protein occurs. At least two other HIV proteins help to mediate these final stages of virion morphogenesis.' HIV has a complex life cycle with multiple enzymatic processes required for virion propagation. Its complexity is unique among the known retrovirus family and it is possible that the numerous gene products of HIV will ultimately provide a number of vulnerable sites for anti-HlV-directcd therapy. As more is known about the HIV life cycle, the potential for new active drugs or combination of drugs that act at different stages of the life cycle will provide a rational approach to HIV-directed therapy.
INHIBITORS OF THE BINDING OF HUMAN IMMUNODEFICIENCY VIRUS TO ITS CELLULAR RECEPTOR The first potentially vulnerable step of the HIV life cycle is binding of virus to the cellular receptor CD4 on T lymphocytes, monocytes, and macrophages. The mature CD4 polypeptide consists of a 370 amino acid extracellular domain with four immunoglobulin-like regions, a transmembrane region, and a hydrophilic cytoplasmic region." The complete CD4 polypeptide is insoluble in serum. A truncated, soluble version of the CD4 polypeptide, which includes most of the extraeelM a r domain, has been produced by recombinant technology. Recombinant soluble CD4 (rCD4), produced
Copyright O 1992 by Thieme Medical Publishers, Inc., 381 Park Avenue South, New York, NY 10016. All rights reserved.
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12, NUMBER 2, 1992
TABLE 1. Anti-HIV Drugs in Relation to the Virus Life Cycle Mechanism
Thercrpeutic Options
Binding of HIV to CD4 receptor
HIV uncoating RNA to DNA Degrade RNA-DNA complex Integrate HIV DNA Decrease activation Transcription of HIV HIV protein synthesis Protein cleavage Sugar addition
Stabilization and prevention of uncoating Reverse transcriptase inhibitors Ribonuclease H lntegrase Inhibit TAT and REV proteins Anti-sense HIV nucleotides Selective inhibition of HIV proteins Protease Myristylation. glycosylation
HIV release from cells Antibody response
Block buding or syncytia formation Promote effective B-, T-cells response
from genetically engineered Chinese hamster ovarian cells, is capable of binding of HIV envelope gp120 with affinity similar to that of native CD4, and blocks entry of HIV-1 into CD4-bearing cells.' Antiviral activity of soluble CD4 has been demonstrated in vitro by measuring decreases in both reverse transcriptase activity and HIV induced cell fusion. '.'l-'"n addition, a single group of investigators has reported in vivo evidence of an antiviral effect of recombinant human CD4 when tested against simian immunodeficiency virus (SIV) in rhesus monkeys." SIV is a lentivirus closely related to HIV-I and HIV-2. Soluble CD4 appears to have a half-life that is not compatible with an effective therapy.'".'7 In an effort to prolong the half-life of CD4, a hybrid molecule was created by combining the gp120 binding sites of CD4 with the constant heavy chain domains of human immunoglobulin (Ig). This molecule is termed immunoadhesion (CD4-lgG).'XImmunoadhesion retains the antiviral activity against HIV but, because of the Ig moiety, CD4IgG has a prolonged plasma half-life, higher serum concentrations with comparable doses, crosses the placenta, and exhibits the capacity to bind Fc receptors.'"-"' Prcliminary studies have not consistently demonstrated an antiviral effect; however, the optimal dose, clinical circumstances, and route of administration have not been defined." It is possible that CD4-lgG may be effective either in preventing vertical transmission of HIV, in managing HIV related immune thrombocytopenia, in the treatment of acute HIV exposure. Other conjugates to recombinant CD4, including immune toxin (Pseudomonas exotoxin), and ricin are also being evaluated clinically. Large negatively charged particles also inhibit HIV binding to the CD4 cellular receptor in vitro. Dextran sulfate, pentosan sulfate, and heparin are examples of anionic polysaccharides that impair the interaction of HIV with C D 4 . ? ' T l i n i c a l trials of these compounds have not shown antiviral or immunologic benefit; they are not active as orally absorbed medication^.'^ Anionic polysaccharides may eventually have utility if their in vitro activity can be translated into in vivo effectiveness. Small peptides such as N-carbomethoxy-caronyl-prolylpenylalanyl benzyl esters are orally active and interact with gp 120 to block HIV- I binding to CD4 receptor."
CD4, CD4-lgG, dextran sulfate, anionic polysaccharides Hypericin, pseudohypericin Zidovudine, didanosine, ddC, second site RTI None in clinical trials None in clinical trials TAT inhibitors None in clinical trials GLQ223, CD4-PE HIV protease inhibitors p17 gag inhibitors castanosperimine, deoxynojirimycin Possibly alpha-interferon Vaccines
Once HIV has bound CD4 receptors, it is internalized and uncoated to release its functional doublestranded RNA genome. Arilodone and WIN5 17 1 are uncoating inhibitors with activity against rhinoviruses and related pinornaviru~es.'~ Compounds that stabilize the HIV capsid structure may also eventually have a role in combating HIV. Hypericin and pseudohypericin are examples of compounds that may act by stabilizing HIV capsids and are active against HIV in vitro."
REVERSE TRANSCRIPTASE INHIBITORS Reverse transcriptase inhibitors have provided the greatest clinical success against HIV. The first drug that was approved by the Food and Drug Administration (FDA) to combat HIV was zidovudine (3'azido-2',3'dideoxythymidine, Retrovir; ZDV, AZT). ZDV is the prototype of a family of 2',3'-dideoxyribose moieties. The three compounds that have demonstrated in vitro and clinical utility are ZDV, 2',3'-dideoxyinosine (didanosinc, Videx; ddl), and 2'3-dideoxythymidine (ddC). The dideoxynucleoside analogs are phosphorylated in the cytoplasm of cells to ultimately yield a dideoxynucleoside 5 ' - t r i p h o ~ p h a t e . These ~~ dideoxynucleoside 5'-triphosphates compete with normal 2'deoxynucleotides for binding to reverse transcriptase and can be incorporated into the growing DNA chain to bring about termination of viral DNA synthesis. These drugs share a similar mechanism of activity but they have significantly different end-organ toxicities and pharmacologic properties. ZDV was initially studied in July 1985, and early in 1986 a multicenter randomized placebo-controlled study was initiated. The study demonstrated a survival advantage to those individuals receiving ZDV compared to those receiving p l a ~ e b o . ~Individuals ' in this original Phase 2 study were selected if their CD4+ cell count was below 200 cells/mm3 or if they had a previous episode of Pneumocystis carinii pneumonia. Subsequent studies demonstrated decreases in the frequency and severity of opportunistic infection, improvement in neurologic function, transient improvement of CD4 T-lymphocytes, and decreases in the serum concentration of HIV antigen in subjects treated with ZDV compared with placebo in pa-
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Event
HIV binding
The standard of care in the tients with HIV United States has become to treat any HIV seropositive individual irrespective of their clinical status, with 500 mglday of ZDV if their C D 4 ~ 'count is less than 500 cells/mm3. There does not appear to be any gender or racial differences to suggest a modification in this recommendation based on these characteristics." ZDV toxicity is protean, with common toxicities primarily affecting the hematologic, gastrointestinal, and neurologic organ systems.34The most common toxicity is bone marrow suppression leading to transfusion-dependent anemia. There has been some suggestion that the recombinant hormone erythropoietin may decrease the transfusion requirement in this situation. Leukopenia and specifically neutropenia are common zidovudine toxicities. This is problematic and often requires dose modification and even drug discontinuation. Gastrointestinal side effects include nausea, vomiting, bloating, cramps, and loss of appetite. These side effects can be limited to an initial 2- to 10-week period but they may also persist and require pharmacologic intervention and even drug discontinuation. Headaches, malaise, and fatigue occur with initial therapy of ZDV but are not usually clinically disabling. The search for active compounds directed against HIV led to the approval of ZDV and more recently didanosine. The process resulting in the approval of the FDA of didanosine broke new ground in the cooperation between the regulatory agency, community advocacy groups, pharmaceutical companies, the National Institutes of Health statisticians, and physician scientists. Didanosine was approved before clinical data were available to verify the utility of the surrogate markers (for example, CD4+ counts. p24 antigen). It will be important to closely follow the evolving data as clinical didanosine studies mature in order to verify that patients, not simply the surrogate markers, improve with didanosine therapy. Didanosine is a purine dideoxynucleoside that is metabolized to an active moiety, dideoxyadenoside triphosphate, with prolonged intracellular activity (half-life of 8 to 24 hours) allowing twice a day oral administration. Didanosine exhibits activity against HIV in both T cells and monocytes with a relatively low toxicity profile." The low toxicity to activity ratio and favorable safety profile led to four Phase 1 studies of didanosine in adults. Didanosine was also evaluated in two pediatric Phase 1 dose escalation clinical trials. Treatment was associated with an increase in CD4+ cells and a decrease in serum p24 antigen compared with patients' values at study entry, although clinical correlates of increased CD4+ cell counts or a decrease in p24 antigen were not identified. The major toxicities in the Phase 1 didanosine studies included abdominal pain, pancreatitis (see later), painful peripheral neuropathy, hepatic enzyme elevation, hyperuricemia, and h y p e r t r i g l y ~ e r i d e m i a . ~ ~ - ~ ~ The Acquired Immunodeficiency Syndrome (AIDS) Clinical Trials Group (ACTG) of the National Institute of Allergies and Infectious Disease has begun large-scale controlled clinical trials of didanosine in adults and children. The first study evaluates didanosine in subjects who experienced hematologic intolerance to ZDV (protocol 118). Another protocol compares ZDV
with didanosine in subjects with less than 16 weeks of prior ZDV therapy (protocol 1 16). A final protocol compares didanosine to ZDV in subjects with more than 16 weeks of ZDV therapy (protocol 117). These trials will provide clinicians with a more complete understanding of the utility of didanosine for a variety of clinical situations. They will also provide the critical information for evaluating the relationship and validity between the laboratory surrogate markers (CD4' cells, p24 antigen) and clinical HIV disease. At the same time that the ACTG centers began to enroll subjects into the protocols just cited, BristolMyers Squibb, (the manufacturers of didanosine) initiated an ambitious program to provide didanosine for those individuals who mav derive some benefit from receiving the drug but who could not participate in the ACTG trials. In all, more than 22,000 individuals in the United States participated in one of two expanded access studies of didanosine. The data collectedfrom the first 7806 individuals may have helped to define some of the clinical toxicities and predictors for the observed toxicities. In the expanded access information data base, 18% of patients developed diarrhea; 16% developed peripheral neuropathy; nausea and vomiting was observed in 8%; abdominal pain, pancreatitis, and headache were each observed in 5% of the participants. Other adverse events occurred at rates lower than 5% and involved every major organ system. The FDA reviewed available information and its Antiviral Advisory Committee recommended approval of didanosine for the treatment of adult and pediatric patients (over age 6 months) with advanced HIV infection who are intolerant of ZDV theraw .. or who have demonstrated significant clinical or immunologic deterioration during ZDV therapy, but these indications are imprecise. Intolerance usually refers to ZDV-induced hem&ologic abnormalities, gastrointestinal or neurologic organ systems. Significant clinical or immunologic deterioration during ZDV therapy may refer to a number of clinical situations. Unfortunately, there are no results from controlled studies regarding the effect of didanosine therapy on the clinical progression of HIV infection; therefore the clinician must exercise judgment regarding the appropriate situation for initiating didanosine therapy. Because there are no readily available tests to measure HIV viral burden, physicians must consider the temporal trend in CD4+ T-cell counts, occurrence of opportunistic infection, weight loss, fatigue, or other indicators of disease activity before initiating didanosine. Clinical progression in sub.jects tolerating ZDV at 500 mgiday, is not well defined. Specific guidelines that might define clinical failure and are being considered by clinical trials are listed in Table 2.
TABLE 2. Guidelines for Didanosine Therapy I. For the person who begins zidovudine therapy with 250 to 500 CD4 ' cells either a 5 0 6 decrease in CD4' cells, or a decline in CD4 ' cells to less than 200 on two occasions separated by at least 4 weeks. 2 . For individuals with 100 to 250 CD4' cells, a 50% decrease in CD4 cells or a decline in CD4' cells to less than 5 0 on two occasions separated by at least 4 weeks. +
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ANTI-HIV THERAPEUTICS-KAHN
The management of toxicities that develop in patients treated with didanosine is clear: stop the medication and treat the patient symptomatically with supportive therapy. Dosing guidelines of didanosine are listed in Table 3. Abdominal pain and pancreatitis must be fully evaluated and alternative explanations of abdominal pain and pancreatitis should be ruled out. These include alcohol intoxication or withdrawal, Kaposi's sarcoma, coadministration of other agents (such as intravenous pentamidine), cytomegalovirus (CMV), AIDS cholangitis, and mycobacterium avium-intracellulare. If didanosine is strongly implicated, then rechallenge with the drug should not be considered. Nine percent of patients who received 12.5 mglkglday or less of didanosine developed pancreatitis. Pancreatitis can be fatal; in the expanded access study 27 of 7806 (0.35%) developed fatal pancreatitis. The risk factors for pancreatitis in subjects treated with didanosine are a previous episode of pancreatitis, concurrent intravenous pentamidine treatment, excessive alcohol consumption, high triglycerides, or intravenous ganciclovir treatment. Peripheral neuropathy developing in subjects treated with didanosine tends to begin as sensory changes, especially numbness, tingling, or pain, in the feet or hands and ascends up the extremities. Subjects who complain of numbness or pain or who have decreased deep tendon reflexes should have their medication stopped. When the neurologic disturbance has abated (usually in 4 to 12 weeks), the medication can be resumed at a 25 to 50% dose reduction. Other toxicities can be managed as indicated by the clinical symptomatology. In general, subjects tolerating didanosine should have, standard serum chemistries and hematologic profiles evaluated every 4 to 8 weeks or as clinically indicated. Over time and in part as a function of disease activity, or immunologic competence, HIV develops in vitro TABLE 3.
Recommended Dosing Guide of Didanosine*
Patient W e i ~ h t
> 75 kg 50-74 kg 35-49 kg Body surface area ( m 2 ) t 1.1-1.4 0.8-1 .O 0.5-0.7 5 0.4
Didrrriosinr~ Ttrhlets
Two 150 mg tablets twice daily Two 100 mg tablets twice daily One I00 and one 25 mg tablet twice daily Two 50 mg tablets twice daily One 50 and one 25 mg tablet twice daily Two 25 mg tablets twice daily One 25 mg twice daily
"Didanosine is available as 25. 50, 100, or 150 mg tablets packaged in child-resistant bottles. The buffering in a single tablet is not adequate to increaae the gastric pH. To prevent gastric acid degradation, adult patients and children older than 1 year of age must take two tablets of didanosine: children under I year (but older than 6 months) receive a one tablet dose. Initial studies in a small number of adult subjects administered didanosine chewable buffered tablets indicates that it is 20 to 25% more bioavailable from the tablet compared to the sachet formulation of didanosine. The administration of the chewable didanosine tablet following a meal results in a 50% decrease in the serum concentration of didanosine. Therefore it is recommended that didanosine be taken on an empty stomach. ?Doses of didanosine in children older than 6 months is baaed on body surface area.
12. NUMBER 2, 1992
resistance to ZDV and didanosine. The selective drug pressure that contributes to the development of this resistance has been documented in vitro and in vivo. However, there is no clear correlation between in vitro HIV resistance to ZDV and clinical deterioration. Moreover, although in vitro HIV resistance occurs with continued ZDV or didanosine treatment, there is no defined length of therapy that will result in the development of resistance or clinical or immunologic deteriorati~n.'~The durability of ZDV resistance in HIV isolates is also unknown. ACTG protocol 117 will help provide information regarding clinical and immunologic benefits for individuals treated with didanosine who have received variable duration ZDV therapy. ddC is the most ~otent-antiretroviralin vitro. Preliminary studies suggest that it may not be as effective a monotherapy modality as ZDV. The exact role of ddC as monotherapy or in combination will be determined by some of the large studies under way. ddC has not been approved by the FDA, but it is widely available through an expanded access program and unofficially through unapproved buyers club mechanisms. The toxicity profile of ddC is similar to ddI; however, pancreatitis appears to have a decreased incidence, although peripheral neuropathy occurs with somewhat increased frequency. Recent studies suggest that alternating ddC with ZDV may decrease the toxicity profile of ddC. The combination of ZDV and didanosine or ZDV and ddC is presently being evaluated in several small studies and one large ACTG study (protocol 155). As yet, the utility of combining nucleoside analogues with similar mechanisms of action is unknown. The risks and toxicities are also unknown and might limit combination therapy. Although combination therapy offers some promise, this approach cannot be recommended4' until Its benefits and-Asks are known. It is understood, however, that decisions between patients and their physicians regarding individualized treatment options are often difficult and based on information that may be incomplete. Second-site reverse transcriptase (RT) inhibitors also inhibit HIV-I RT but with a different mechanism from that of the 2,3'dideoxynucleotides. The prototypic compounds are based on the benzodiazepine structure and are referred to as tetrahydroimidazobenzodiazepinone derivative^.^' Other compounds with specific HIV-I RT inhibition include L-697, 661, and nivarapene. The mechanism of their activity is not completely known but probably involves inhibition of the p66 heterodimer of the RT complex. These compounds are protein-avid, with prolonged half-life, and good cerebrospinal fluid penetration. In many respects second-site RT inhibitors-have characteristics that-are desirable for antiviral therapeutics. Clinical evaluation of these compounds has begun in subjects with advanced HIV disease. The early clinical trials suggest that the unique specificity and activity of these compounds may limit their clinical utility. Resistance to the test compounds appears to occur rapidly when they are used as monotherapy. Resistance to the test compounds does not confer resistance to other dideoxynucleotides. Perhaps, these compounds will be most effective in combination with other therapies or in subjects without advanced disease.
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ANTI-HIV THERAPEUTICS-KAHN TABLE 4.
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Current Guidelines for HIV Reverse Transcriptase Inhibitors in Adults*
Drug
Indicution
Dosuge
Schedule
ZDV Didanosine
4 0 0 CD4 ' See table 2
100-200 300 mg
TID BID
ddC
Unknown
0.75 mg
BID
Toxicit?; Anemia Neuropathy, rash, GI pancreatitis Neuropathy, rash, GI pancreatitis
Monitoring
Cotnnzetzts
CBC, LFTs CBC, amylase. neurologic examination
Initial HIV therapy ZDV intolerance and immunologic deterioration
CBC, amylase, neurologic examination
Available through expanded access
POST-REVERSE TRANSCRIPTION, INHIBITORS OF HUMAN IMMUNODEFICIENCY VIRUS ACTIVATION AND OF PROTEIN INHIBITORS The action of ribotz~~cleuse H and integrase present two other obvious targets to inhibit HIV. Drug development targeted to these enzymes is an area of active research. There are fewer drugs that actively inhibit HIV once it has integrated into the host genome. After HIV has intcgrated itself, efforts to inhibit the virus have focused on the products of HIV TAT and REV gene^.^^,^' This strategy aims to maintain HIV in a latent stage of infection. TAT inhibitors have reached the stage of clinical trials, but inhibitors of REV are in laboratory development. Oligodeoxynucleotides are complementary nucleotides to the HIV genome that prevent transcription of the viral DNA to viral mRNA. Oligondeoxynucleotides may also bind to viral mRNA, forming an RNAIDNA hybrid, preventing translation of the viral mRNA into protein. Oligodeoxynucleotides have been shown to inhibit HIV replication in ~ i v o . ~Therapeutic ~.~' utility of antisense oligonucleoties face serious developmental challenges. The small (15 to 20 nucleotides) must traverse the plasma, contact and then penetrate into cells, resist premature degradation by nucleases, and ultimately reach their target sequences at the nucleus. Despite these formidable problems, antisense-based therapy is undergoing active development and will probably lead to clinical trials. GLQ223 is a highly purified formulation of trichosanthin, a basic (pl 9.6) 27 kd protein consisting of 247 amino acids that is purified from the root tubers of Trichosanthes kirilowii, M a x i m o ~ i c z . ~ In ~ - ~China ' single doses of trichosanthin-containing products have been used as a midtrimester abortifacient and multiple doses have been used for the treatment of trophoblastic tum o r ~ . ~ In ~ ' .vitro ~ ' studies suggest that trichosanthin selectively inhibits the replication of HIV in lymphoid and mononuclear cells.52On the basis of structural and functional properties, trichosanthin is considered a member of the family of single-chain ribosome inactivating prot e i n ~ . ~ ~The . ~ ' -selective ~~ inhibition of HIV-infected cells is probably a post-translational event. Data from the initial Phase I evaluation indicated that bolus infusion of GLQ223 provided serum concentrations of study drug that predicted effects; however, the duration of GLQ223
concentration was inadequate to observe anti-HIV eff e c t ~ Computer .~~ modeling of pharmacokinetic data from the single infusion study suggested that a brief bolus followed by a prolonged infusion would achieve serum concentrations that would reproduce in vitro drug concentrations that exerted anti-HIV effects. In order to evaluate the pharmacokinetic profile of GLQ223 during prolonged administration, a second phase 1 study was begun and completed at San Francisco General Hospital. GLQ223 was administered as a loading dose followed by a 3-hour infusion. Each subject received a test dose of 50 ng of GLQ223 prior to study drug administration. Subjects received weekly infusions of GLQ223 for 4 consecutive weeks. This was followed by a 21-day follow-up period during which no study drug was administered. All subjects were monitored for tolerance and toxicity. GLQ223 administration was not associated with notable toxicity except for a flulike syndrome characterized by muscle aches, throat pain, low-grade fevers, and chills associated with an increase in the CPK (MM fraction) isoenzyme level. This syndrome was easily controlled. Serum concentrations of GLQ223 were achieved in the two highest doses that were comparable to concentrations shown to have anti-HIV activity. Immunologic parameters suggest that GLQ223 has dose-dependent activity in HIV-infected individuals. A Phase 213 study comparing clinical and immunologic events in subjects with 200 to 500 CD4 ' cells and at least 9 months of ZDV therapy has been initiated. The subjects are randomized to one of three treatment arms, ZDV alone, GLQ223 alone, and ZDV plus GLQ223. GLQ223 is administered as a 3-hour intravenous infusion. Sixteen infusions are planned and all subjects will be followed for 48 weeks. Rational drug design has had the greatest role in developing compounds that inhibit the viral enzyme HIV protease. HIV protease recognizes as substrate the peptide bond between tyrosine (phenylalanine) and proline .57-59 The crystal structure of protease enzyme has been obtained. Inhibitors to protease have been designed based on the three-dimensional structure of the enzyme. Potent inhibitors have been found from large chemical catalogues and improvements have been designed in a few promising inhibitor candidates. As a group, these compounds inhibit HIV-1, HIV-2, and SIV. They have a poor oral bioavailability profile but potent in vitro antiHIV effects. Phase 1 clinical trials of protease inhibitors have begun in Europe. Protease inhibitors have been de-
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"BID: twice daily; CBC: complete blood count; GI: gastrointestinal; HIV: human immunodeficiency virus: LFT: liver function tests: TID: three times daily: ZDV: zidovudine.
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veloped for clinical medicine. There is clinical precedent for the development of this type of compound, for example, angiotension converting enzyme inhibitors. The attachment of myristic acid to the NH2 terminal of the gag and gag-pol precursor protein and glycosylation of the viral envelope proteins are critical factors for the maturation of viral proteins and the viability of new virions. Products that interfere with the normal sugar attachment to viral proteins may therefore represent a class of compounds that selectively inhibit HIV activity. Castanospermine and deoxynorjirimycin, (which inhibit glycosylation) reactions catalyzed by alpha-glucosideases I and 11, are only weakly active against HIV, probably because they are not HIV s p e ~ i f i c . ~Compounds ~,~' that inhibit the myristylation of p17 gag protein interfere with the assembly of HIV and mature infectious virion^,^^.^' and thus may provide a useful therapeutic option. A final area of anti-HIV research is the development of HIV vaccines to stimulate the immune system of HIV-infected patients. United States Army data have already been presented that indicate that monthly vaccination with a prototype HIV gp160 vaccine can induce new specific HIV antibodies and new T-cell functional response^.^ Those individuals with new antibody and cellular responses have developed stabilization of their CD4 lymphocyte counts compared with individuals who failed to develop these new responses. Other studies of vaccine constructs in seropositive individuals have begun, including a Phase 112 study at San Francisco General Hospital. The exact mechanism of action for postinfection vaccination-inducing immunologic stabilization is not known, however this strategy appears to have limited toxicity. Well-controlled clinical trials will define the role of vaccination for infected individuals.
SUMMARY Knowledge of the HIV life cycle helps to plan rational modalities directed against the virus. The complex nature of the life cycle of HIV presents multiple unique targets. HIV binding to the cellular receptor CD4, viral enzymatic targets, HIV activation, viral protein synthesis, and protein packaging are examples of the types of targets available to inhibit the life cycle of HIV. Patients, community groups, regulatory agencies, national research groups, and the private pharmaceutical companies have joined forces to work to a common goal, effective HIV therapies. RT inhibitors have provided the best therapeutic options, but other drugs are being developed at a rapid pace. New drugs are entering clinical investigation and will be more widely available at early signs of clinical and laboratory effectiveness.
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Fischl MA. et al: Prolonged zidovudine therapy in patients with AIDS and advanced AIDS-related complex. AZT collaborative Working Group. JAMA 262:2405-2410, 1989. Volberding, et al: Zidovudine in asymptomatic human immunodeficiency virus infection: A controlled trial in persons with fewer than 500 CD4-positive cells per cubic millimeter. N Engl J Med 322:941-949. 1990. Fischl MA, et al: Thc safety and efficacy of zidovudine (AZT) in the treatment of subjects with mildly symptomatic human immunodeficiency virus type I (HIV) infection A doubleblind, placebo-controlled trial. Ann Intern Med 112:727-737, 1990. Yarchoan R, et al: Long-term administration of 3'-azido2',3'dideocythymidine to patients with AIDS-related neurologic disease. Ann Neurol 23(Suppl):S82-87, 1988. Chaisson RE. ct al: Effect of zidovudine on serum human immunodeficiency virus core antigen levels. Results from a placebo-controlled trial. Arch Intern Med 148:215 1-2 153, 1988. Lagakos S, et al: Effects of zidovudine therapy in minority and other subpopulations with early HIV infection. JAMA 266:2709-27 12, 199 1. Richman DD, et al: The toxicity of azidothymidine (AZT) in the treatment of patients with AIDS and AIDS-related complex: A double-blind, placebo-controlled trial. N Engl J Med 317:192-197, 1987. Yarchoan R, et al: In vivo activity against HIV and favorable toxicity profile of 2',3'-dideoxyinosine. Science 245:412415, 1989. Lambert JS, et al: 2',3'-Dideoxyinosine (ddI) in patients with the acquired immunodeficiency syndrome or AIDS-related complex. A phase I trial. N Engl J Med 322:1333-1339, 1990. Cooley TP, et al: Once-daily administration of 2',3'-dideoxyinosine (ddI) in patients with the acquired immunodeficiency syndrome or AIDS-related complex; results of a Phase I trial. N Engl J Med 322: 1340- 1345, 1990. Yarchoan R, et al: Long-term toxicitylactivity profile of 2',3'dideoxyinosine in AIDS or AIDS-related complex. Lancet 336:526-529, 1990. St. Clair M H , et al: Resistance to dd1 and sensitivity to AZT induced by a mutation in HIV-I reverse transcriptase. Science 253:1557-1559, 1991. Johnson VA, et al: Two-drug combinations of zidovudine, didanosine and recombinant interferon-alpha A inhibit replication of zidovudine-resistant human irnmunodeficicncy virus type I synergistically in vitro. J Infect Dis 164:646-655, 1991. Pauwcls R , et al: Potent and selective inhibition of HIV-I replication in vitro by a novel series of TlBO derivatives. Nature 343:470-474, 1990. Kao S-Y. et al: Anti-termination of transcription within the long-terminal repeat of HIV-I by tat gene product. Nature 330:489493, 1987. Baltimore D: lntracellular immunization. Nature 335:395396, 1988. Sarin P, et al: Inhibition of acquired immunodeficiency syndrome by oligodeoxynucleoside methyphosphonates. Proc Nall Acad Sci USA 85:7448-7451, 1988. Matsukura M, et al: Regulation of viral expression of human immunodeficiency virus in vitro by an antisense phophorothioate oligodeocynucleotide against rev (arrltrs) in chroni-
cally infected cells. Proc Natl Acad Sci USA 86:42444248, 1989. Yeung HW, et al: Purification of three isolectins from root tubers of Trichosanthes kirilowii (Tiahuafen). Int J Pept Protein Res 27:325-333, 1986. Wang Y, et al: Scientific evaluation of Tian Hua Fen (THF)history, chemistry and application. Pure Appl Chem 58:789798, 1986. Caaellas P, et al: Trichokirin. a ribosome-inactivating protein from the seeds of Trichosanthes kirilowii Maximowicz. Purification. partial characterization and use for preparation of immunotoxins. Eur J Biochem 176:58 1-588, 1988. Maraganore JM, et al: Purification and characterization of trichosantm. Homology to the ricin A chain and implications as to mechanism of abortifiaceint activity. J Biol Chem 262:11628-ll633, 1987. Cheng KF. Midtrimester abortion induced by Radix l'richosanthis: Morphologic observations in placenta and fetus. Obstet Gynecol 59:494-498, 1982. Chan Y, Tong MK, Lau MJ. Shanghai J Clin Med 1982;3:303 1 (in Chinese). McGrath MS, et al: GLQ223: An inhibitor of human immunodeficiency virus replication in acutely and chronically infected cells of lymphocyte and mononuclear phagocyte lineage. Proc Natl Acad Sci USA 86:2844-2848. 1989. Yeung H. et al: Trichosanthin, alpha-momorcharin and betamomorcharin: Identity of abortifacient and ribosome-inactivating proteins. Int J Pept Protein Res 31:265-268. 1988. Kubota S. et al: Confirmational similarities of ricin A-chain and tr~chosanthin.Intl J Pept Protein Res 30:646-651, 1987. Zhang XJ. Wang JH: Homology of trichosanthin and ricin A chain. Nature 321:47-48, 1986. Kahn JO, et al: The safety and pharmacokinetics of GLQ223 in subjects with acquired immunodeficiency syndrome (AIDS) and AIDS-related complex: A Phase 1 study. AIDS. 4: 11971204, 1990. Graves MC. et al: An 1 I-kDA form of human immunodeficiency virus protease expressed in Escherichia coli is sufficient Tor enzymatic activity. Proc Natl Acad Sci USA 85:24492453, 1988. Kotler M, et al: Synthetic peptides as substrates and inhibitors of a retroviral protease. Proc Natl Acad Sci USA 85:41854189, 1988. Roberts NA, et al: Rational design of peptide-based HIV protcinase inhibitors. Science 248: 148-1 5 1 , 1990. Walker BD, et al: Inhibition of human immunodeficiency virus syncytium formation and virus replication by castanospermine. Proc Natl Acad Sci USA 84:8120-8124, 1987. Karpas A, et al: Aminosugar derivatives as potential anti-human immunodeficiency virus agents. Proc Natl Acad Sci USA 85:9229-9233, 1988. Bryant ML, et al: Replication of human immunodeficiency virus 1 and Moloney murine leukemia virus is inhibited by different heteroatom-containing analogs of myristic acid. Proc Natl Sci USA. 86:8655-8659, 1989. Trono D, et al: HIV-I Gag mutants can dominantly interfere with the replication of the wild-type virus. 59:l 13-120, 1989. Redfield RR, et al: A phase I evaluation of the safety and immunogenicity of vaccination with recombinant gp 160 in patients with early human immunodeficiency virus infection. N Engl J Med 324: 1677-1684, 1991.
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ANTI-HIV THERAPEUTICS-KAHN
12, NO. 2. 1992
Anti-Human Immunodeficiency Virus Therapeutics: Now and the Future
Therapeutic modalities to inhibit, inactivate, or interfere with the usual activities of the human immunodeficiency virus (HIV) are best understood in reference to its life cycle. The preceding article by Pavlakis reviews the basic pathophysiology of HIV infection, including the consequences of HIV infection and the immunopathology of the reaction between virus and host. In order to understand the mechanism of anti-HIV directed drugs, this article will first briefly review the major sequences of HIV infection and the HIV life cycle, followed by a description of anti-HIV drugs and other potential therapeutic modalities.
LIFE CYCLE OF THE HUMAN IMMUNODEFICIENCY VIRUS T lymphocytes and monocytes are the major targets for HIV. The HIV virus envelope glycoprotein, gp120, M) to the CD4 binds with high affinity (K,-lop' molecule'^' on target cells. CD4 is a glycoprotein on the surface of mature CD4 (T4) lymphocytes and serves as the primary receptor for the HIV in humans." The CD4 glycoprotein normally facilitates T-helper cell recognition of antigens presented by class I1 major histocompatibility complex (MHC) molecule^.^.^ The process by which HIV enters the cell after it has bound the CD4 receptor is not completely understood; however, once the virus is internalized and uncoated, a single strand of the double-stranded HIV RNA acts as the template for HIV enzyme reverse transcriptase. This enzyme creates a single strand DNA from the viral template. Ribonuclease H, a second HIV enzyme, partially degrades the original HIV RNA template. Reverse transcriptase than acts to create a second DNA strand from the original HIV RNA. The double-stranded HIV DNA is translocated to the host cell nucleus and the HIV DNA duplex is inserted into the host genome by integrase, a third HIV enzyme.' After incorporation into the target cell, a latent or persistent form of HIV infection may develop. The rea-
From the Department of' Medicine. University of California, San Francisco, and the AIDS Program, Sun Francisco General Hospital. Son Francisco, California Reprint requests: Dr. Kahn, Department of Medicine, San Francisco General Hospital, 995 Potrero Ave., San Francisco, CA 941 10.
sons for latency remain unknown, as are the factors that serve to activate HIV after a prolonged dormancy. Many researchers believe that activation of HIV is coupled to the NF-kappa B family of proteins.' Once HIV is active, required regulatory proteins are synthesized. Viral mRNA leaves the nucleus, migrates to the cytoplasm where regulatory protein synthesis is supcrscded by the synthesis of HIV-structural proteins via a complicated feedback mechanism. Structural protein synthesis is required for mature virion production. The structural proteins include the exterior HIV coated proteins and viral enzymes: reverse transcriptase, ribonuclease H, integrase, and an HIV aspartyl protease. HIV core proteins are cleaved from a precursor protein by the HIV-derived protease. Assembly of the infectious HIV virion is accomplished by aggregation of viral core proteins, HIV-I RNA, and enzymes while moving to the surface of the cell. The cores bud, acquire the cell lipid bilayer, and final myristylation of a core protein occurs. At least two other HIV proteins help to mediate these final stages of virion morphogenesis.' HIV has a complex life cycle with multiple enzymatic processes required for virion propagation. Its complexity is unique among the known retrovirus family and it is possible that the numerous gene products of HIV will ultimately provide a number of vulnerable sites for anti-HlV-directcd therapy. As more is known about the HIV life cycle, the potential for new active drugs or combination of drugs that act at different stages of the life cycle will provide a rational approach to HIV-directed therapy.
INHIBITORS OF THE BINDING OF HUMAN IMMUNODEFICIENCY VIRUS TO ITS CELLULAR RECEPTOR The first potentially vulnerable step of the HIV life cycle is binding of virus to the cellular receptor CD4 on T lymphocytes, monocytes, and macrophages. The mature CD4 polypeptide consists of a 370 amino acid extracellular domain with four immunoglobulin-like regions, a transmembrane region, and a hydrophilic cytoplasmic region." The complete CD4 polypeptide is insoluble in serum. A truncated, soluble version of the CD4 polypeptide, which includes most of the extraeelM a r domain, has been produced by recombinant technology. Recombinant soluble CD4 (rCD4), produced
Copyright O 1992 by Thieme Medical Publishers, Inc., 381 Park Avenue South, New York, NY 10016. All rights reserved.
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12, NUMBER 2, 1992
TABLE 1. Anti-HIV Drugs in Relation to the Virus Life Cycle Mechanism
Thercrpeutic Options
Binding of HIV to CD4 receptor
HIV uncoating RNA to DNA Degrade RNA-DNA complex Integrate HIV DNA Decrease activation Transcription of HIV HIV protein synthesis Protein cleavage Sugar addition
Stabilization and prevention of uncoating Reverse transcriptase inhibitors Ribonuclease H lntegrase Inhibit TAT and REV proteins Anti-sense HIV nucleotides Selective inhibition of HIV proteins Protease Myristylation. glycosylation
HIV release from cells Antibody response
Block buding or syncytia formation Promote effective B-, T-cells response
from genetically engineered Chinese hamster ovarian cells, is capable of binding of HIV envelope gp120 with affinity similar to that of native CD4, and blocks entry of HIV-1 into CD4-bearing cells.' Antiviral activity of soluble CD4 has been demonstrated in vitro by measuring decreases in both reverse transcriptase activity and HIV induced cell fusion. '.'l-'"n addition, a single group of investigators has reported in vivo evidence of an antiviral effect of recombinant human CD4 when tested against simian immunodeficiency virus (SIV) in rhesus monkeys." SIV is a lentivirus closely related to HIV-I and HIV-2. Soluble CD4 appears to have a half-life that is not compatible with an effective therapy.'".'7 In an effort to prolong the half-life of CD4, a hybrid molecule was created by combining the gp120 binding sites of CD4 with the constant heavy chain domains of human immunoglobulin (Ig). This molecule is termed immunoadhesion (CD4-lgG).'XImmunoadhesion retains the antiviral activity against HIV but, because of the Ig moiety, CD4IgG has a prolonged plasma half-life, higher serum concentrations with comparable doses, crosses the placenta, and exhibits the capacity to bind Fc receptors.'"-"' Prcliminary studies have not consistently demonstrated an antiviral effect; however, the optimal dose, clinical circumstances, and route of administration have not been defined." It is possible that CD4-lgG may be effective either in preventing vertical transmission of HIV, in managing HIV related immune thrombocytopenia, in the treatment of acute HIV exposure. Other conjugates to recombinant CD4, including immune toxin (Pseudomonas exotoxin), and ricin are also being evaluated clinically. Large negatively charged particles also inhibit HIV binding to the CD4 cellular receptor in vitro. Dextran sulfate, pentosan sulfate, and heparin are examples of anionic polysaccharides that impair the interaction of HIV with C D 4 . ? ' T l i n i c a l trials of these compounds have not shown antiviral or immunologic benefit; they are not active as orally absorbed medication^.'^ Anionic polysaccharides may eventually have utility if their in vitro activity can be translated into in vivo effectiveness. Small peptides such as N-carbomethoxy-caronyl-prolylpenylalanyl benzyl esters are orally active and interact with gp 120 to block HIV- I binding to CD4 receptor."
CD4, CD4-lgG, dextran sulfate, anionic polysaccharides Hypericin, pseudohypericin Zidovudine, didanosine, ddC, second site RTI None in clinical trials None in clinical trials TAT inhibitors None in clinical trials GLQ223, CD4-PE HIV protease inhibitors p17 gag inhibitors castanosperimine, deoxynojirimycin Possibly alpha-interferon Vaccines
Once HIV has bound CD4 receptors, it is internalized and uncoated to release its functional doublestranded RNA genome. Arilodone and WIN5 17 1 are uncoating inhibitors with activity against rhinoviruses and related pinornaviru~es.'~ Compounds that stabilize the HIV capsid structure may also eventually have a role in combating HIV. Hypericin and pseudohypericin are examples of compounds that may act by stabilizing HIV capsids and are active against HIV in vitro."
REVERSE TRANSCRIPTASE INHIBITORS Reverse transcriptase inhibitors have provided the greatest clinical success against HIV. The first drug that was approved by the Food and Drug Administration (FDA) to combat HIV was zidovudine (3'azido-2',3'dideoxythymidine, Retrovir; ZDV, AZT). ZDV is the prototype of a family of 2',3'-dideoxyribose moieties. The three compounds that have demonstrated in vitro and clinical utility are ZDV, 2',3'-dideoxyinosine (didanosinc, Videx; ddl), and 2'3-dideoxythymidine (ddC). The dideoxynucleoside analogs are phosphorylated in the cytoplasm of cells to ultimately yield a dideoxynucleoside 5 ' - t r i p h o ~ p h a t e . These ~~ dideoxynucleoside 5'-triphosphates compete with normal 2'deoxynucleotides for binding to reverse transcriptase and can be incorporated into the growing DNA chain to bring about termination of viral DNA synthesis. These drugs share a similar mechanism of activity but they have significantly different end-organ toxicities and pharmacologic properties. ZDV was initially studied in July 1985, and early in 1986 a multicenter randomized placebo-controlled study was initiated. The study demonstrated a survival advantage to those individuals receiving ZDV compared to those receiving p l a ~ e b o . ~Individuals ' in this original Phase 2 study were selected if their CD4+ cell count was below 200 cells/mm3 or if they had a previous episode of Pneumocystis carinii pneumonia. Subsequent studies demonstrated decreases in the frequency and severity of opportunistic infection, improvement in neurologic function, transient improvement of CD4 T-lymphocytes, and decreases in the serum concentration of HIV antigen in subjects treated with ZDV compared with placebo in pa-
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Event
HIV binding
The standard of care in the tients with HIV United States has become to treat any HIV seropositive individual irrespective of their clinical status, with 500 mglday of ZDV if their C D 4 ~ 'count is less than 500 cells/mm3. There does not appear to be any gender or racial differences to suggest a modification in this recommendation based on these characteristics." ZDV toxicity is protean, with common toxicities primarily affecting the hematologic, gastrointestinal, and neurologic organ systems.34The most common toxicity is bone marrow suppression leading to transfusion-dependent anemia. There has been some suggestion that the recombinant hormone erythropoietin may decrease the transfusion requirement in this situation. Leukopenia and specifically neutropenia are common zidovudine toxicities. This is problematic and often requires dose modification and even drug discontinuation. Gastrointestinal side effects include nausea, vomiting, bloating, cramps, and loss of appetite. These side effects can be limited to an initial 2- to 10-week period but they may also persist and require pharmacologic intervention and even drug discontinuation. Headaches, malaise, and fatigue occur with initial therapy of ZDV but are not usually clinically disabling. The search for active compounds directed against HIV led to the approval of ZDV and more recently didanosine. The process resulting in the approval of the FDA of didanosine broke new ground in the cooperation between the regulatory agency, community advocacy groups, pharmaceutical companies, the National Institutes of Health statisticians, and physician scientists. Didanosine was approved before clinical data were available to verify the utility of the surrogate markers (for example, CD4+ counts. p24 antigen). It will be important to closely follow the evolving data as clinical didanosine studies mature in order to verify that patients, not simply the surrogate markers, improve with didanosine therapy. Didanosine is a purine dideoxynucleoside that is metabolized to an active moiety, dideoxyadenoside triphosphate, with prolonged intracellular activity (half-life of 8 to 24 hours) allowing twice a day oral administration. Didanosine exhibits activity against HIV in both T cells and monocytes with a relatively low toxicity profile." The low toxicity to activity ratio and favorable safety profile led to four Phase 1 studies of didanosine in adults. Didanosine was also evaluated in two pediatric Phase 1 dose escalation clinical trials. Treatment was associated with an increase in CD4+ cells and a decrease in serum p24 antigen compared with patients' values at study entry, although clinical correlates of increased CD4+ cell counts or a decrease in p24 antigen were not identified. The major toxicities in the Phase 1 didanosine studies included abdominal pain, pancreatitis (see later), painful peripheral neuropathy, hepatic enzyme elevation, hyperuricemia, and h y p e r t r i g l y ~ e r i d e m i a . ~ ~ - ~ ~ The Acquired Immunodeficiency Syndrome (AIDS) Clinical Trials Group (ACTG) of the National Institute of Allergies and Infectious Disease has begun large-scale controlled clinical trials of didanosine in adults and children. The first study evaluates didanosine in subjects who experienced hematologic intolerance to ZDV (protocol 118). Another protocol compares ZDV
with didanosine in subjects with less than 16 weeks of prior ZDV therapy (protocol 1 16). A final protocol compares didanosine to ZDV in subjects with more than 16 weeks of ZDV therapy (protocol 117). These trials will provide clinicians with a more complete understanding of the utility of didanosine for a variety of clinical situations. They will also provide the critical information for evaluating the relationship and validity between the laboratory surrogate markers (CD4' cells, p24 antigen) and clinical HIV disease. At the same time that the ACTG centers began to enroll subjects into the protocols just cited, BristolMyers Squibb, (the manufacturers of didanosine) initiated an ambitious program to provide didanosine for those individuals who mav derive some benefit from receiving the drug but who could not participate in the ACTG trials. In all, more than 22,000 individuals in the United States participated in one of two expanded access studies of didanosine. The data collectedfrom the first 7806 individuals may have helped to define some of the clinical toxicities and predictors for the observed toxicities. In the expanded access information data base, 18% of patients developed diarrhea; 16% developed peripheral neuropathy; nausea and vomiting was observed in 8%; abdominal pain, pancreatitis, and headache were each observed in 5% of the participants. Other adverse events occurred at rates lower than 5% and involved every major organ system. The FDA reviewed available information and its Antiviral Advisory Committee recommended approval of didanosine for the treatment of adult and pediatric patients (over age 6 months) with advanced HIV infection who are intolerant of ZDV theraw .. or who have demonstrated significant clinical or immunologic deterioration during ZDV therapy, but these indications are imprecise. Intolerance usually refers to ZDV-induced hem&ologic abnormalities, gastrointestinal or neurologic organ systems. Significant clinical or immunologic deterioration during ZDV therapy may refer to a number of clinical situations. Unfortunately, there are no results from controlled studies regarding the effect of didanosine therapy on the clinical progression of HIV infection; therefore the clinician must exercise judgment regarding the appropriate situation for initiating didanosine therapy. Because there are no readily available tests to measure HIV viral burden, physicians must consider the temporal trend in CD4+ T-cell counts, occurrence of opportunistic infection, weight loss, fatigue, or other indicators of disease activity before initiating didanosine. Clinical progression in sub.jects tolerating ZDV at 500 mgiday, is not well defined. Specific guidelines that might define clinical failure and are being considered by clinical trials are listed in Table 2.
TABLE 2. Guidelines for Didanosine Therapy I. For the person who begins zidovudine therapy with 250 to 500 CD4 ' cells either a 5 0 6 decrease in CD4' cells, or a decline in CD4 ' cells to less than 200 on two occasions separated by at least 4 weeks. 2 . For individuals with 100 to 250 CD4' cells, a 50% decrease in CD4 cells or a decline in CD4' cells to less than 5 0 on two occasions separated by at least 4 weeks. +
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ANTI-HIV THERAPEUTICS-KAHN
The management of toxicities that develop in patients treated with didanosine is clear: stop the medication and treat the patient symptomatically with supportive therapy. Dosing guidelines of didanosine are listed in Table 3. Abdominal pain and pancreatitis must be fully evaluated and alternative explanations of abdominal pain and pancreatitis should be ruled out. These include alcohol intoxication or withdrawal, Kaposi's sarcoma, coadministration of other agents (such as intravenous pentamidine), cytomegalovirus (CMV), AIDS cholangitis, and mycobacterium avium-intracellulare. If didanosine is strongly implicated, then rechallenge with the drug should not be considered. Nine percent of patients who received 12.5 mglkglday or less of didanosine developed pancreatitis. Pancreatitis can be fatal; in the expanded access study 27 of 7806 (0.35%) developed fatal pancreatitis. The risk factors for pancreatitis in subjects treated with didanosine are a previous episode of pancreatitis, concurrent intravenous pentamidine treatment, excessive alcohol consumption, high triglycerides, or intravenous ganciclovir treatment. Peripheral neuropathy developing in subjects treated with didanosine tends to begin as sensory changes, especially numbness, tingling, or pain, in the feet or hands and ascends up the extremities. Subjects who complain of numbness or pain or who have decreased deep tendon reflexes should have their medication stopped. When the neurologic disturbance has abated (usually in 4 to 12 weeks), the medication can be resumed at a 25 to 50% dose reduction. Other toxicities can be managed as indicated by the clinical symptomatology. In general, subjects tolerating didanosine should have, standard serum chemistries and hematologic profiles evaluated every 4 to 8 weeks or as clinically indicated. Over time and in part as a function of disease activity, or immunologic competence, HIV develops in vitro TABLE 3.
Recommended Dosing Guide of Didanosine*
Patient W e i ~ h t
> 75 kg 50-74 kg 35-49 kg Body surface area ( m 2 ) t 1.1-1.4 0.8-1 .O 0.5-0.7 5 0.4
Didrrriosinr~ Ttrhlets
Two 150 mg tablets twice daily Two 100 mg tablets twice daily One I00 and one 25 mg tablet twice daily Two 50 mg tablets twice daily One 50 and one 25 mg tablet twice daily Two 25 mg tablets twice daily One 25 mg twice daily
"Didanosine is available as 25. 50, 100, or 150 mg tablets packaged in child-resistant bottles. The buffering in a single tablet is not adequate to increaae the gastric pH. To prevent gastric acid degradation, adult patients and children older than 1 year of age must take two tablets of didanosine: children under I year (but older than 6 months) receive a one tablet dose. Initial studies in a small number of adult subjects administered didanosine chewable buffered tablets indicates that it is 20 to 25% more bioavailable from the tablet compared to the sachet formulation of didanosine. The administration of the chewable didanosine tablet following a meal results in a 50% decrease in the serum concentration of didanosine. Therefore it is recommended that didanosine be taken on an empty stomach. ?Doses of didanosine in children older than 6 months is baaed on body surface area.
12. NUMBER 2, 1992
resistance to ZDV and didanosine. The selective drug pressure that contributes to the development of this resistance has been documented in vitro and in vivo. However, there is no clear correlation between in vitro HIV resistance to ZDV and clinical deterioration. Moreover, although in vitro HIV resistance occurs with continued ZDV or didanosine treatment, there is no defined length of therapy that will result in the development of resistance or clinical or immunologic deteriorati~n.'~The durability of ZDV resistance in HIV isolates is also unknown. ACTG protocol 117 will help provide information regarding clinical and immunologic benefits for individuals treated with didanosine who have received variable duration ZDV therapy. ddC is the most ~otent-antiretroviralin vitro. Preliminary studies suggest that it may not be as effective a monotherapy modality as ZDV. The exact role of ddC as monotherapy or in combination will be determined by some of the large studies under way. ddC has not been approved by the FDA, but it is widely available through an expanded access program and unofficially through unapproved buyers club mechanisms. The toxicity profile of ddC is similar to ddI; however, pancreatitis appears to have a decreased incidence, although peripheral neuropathy occurs with somewhat increased frequency. Recent studies suggest that alternating ddC with ZDV may decrease the toxicity profile of ddC. The combination of ZDV and didanosine or ZDV and ddC is presently being evaluated in several small studies and one large ACTG study (protocol 155). As yet, the utility of combining nucleoside analogues with similar mechanisms of action is unknown. The risks and toxicities are also unknown and might limit combination therapy. Although combination therapy offers some promise, this approach cannot be recommended4' until Its benefits and-Asks are known. It is understood, however, that decisions between patients and their physicians regarding individualized treatment options are often difficult and based on information that may be incomplete. Second-site reverse transcriptase (RT) inhibitors also inhibit HIV-I RT but with a different mechanism from that of the 2,3'dideoxynucleotides. The prototypic compounds are based on the benzodiazepine structure and are referred to as tetrahydroimidazobenzodiazepinone derivative^.^' Other compounds with specific HIV-I RT inhibition include L-697, 661, and nivarapene. The mechanism of their activity is not completely known but probably involves inhibition of the p66 heterodimer of the RT complex. These compounds are protein-avid, with prolonged half-life, and good cerebrospinal fluid penetration. In many respects second-site RT inhibitors-have characteristics that-are desirable for antiviral therapeutics. Clinical evaluation of these compounds has begun in subjects with advanced HIV disease. The early clinical trials suggest that the unique specificity and activity of these compounds may limit their clinical utility. Resistance to the test compounds appears to occur rapidly when they are used as monotherapy. Resistance to the test compounds does not confer resistance to other dideoxynucleotides. Perhaps, these compounds will be most effective in combination with other therapies or in subjects without advanced disease.
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ANTI-HIV THERAPEUTICS-KAHN TABLE 4.
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Current Guidelines for HIV Reverse Transcriptase Inhibitors in Adults*
Drug
Indicution
Dosuge
Schedule
ZDV Didanosine
4 0 0 CD4 ' See table 2
100-200 300 mg
TID BID
ddC
Unknown
0.75 mg
BID
Toxicit?; Anemia Neuropathy, rash, GI pancreatitis Neuropathy, rash, GI pancreatitis
Monitoring
Cotnnzetzts
CBC, LFTs CBC, amylase. neurologic examination
Initial HIV therapy ZDV intolerance and immunologic deterioration
CBC, amylase, neurologic examination
Available through expanded access
POST-REVERSE TRANSCRIPTION, INHIBITORS OF HUMAN IMMUNODEFICIENCY VIRUS ACTIVATION AND OF PROTEIN INHIBITORS The action of ribotz~~cleuse H and integrase present two other obvious targets to inhibit HIV. Drug development targeted to these enzymes is an area of active research. There are fewer drugs that actively inhibit HIV once it has integrated into the host genome. After HIV has intcgrated itself, efforts to inhibit the virus have focused on the products of HIV TAT and REV gene^.^^,^' This strategy aims to maintain HIV in a latent stage of infection. TAT inhibitors have reached the stage of clinical trials, but inhibitors of REV are in laboratory development. Oligodeoxynucleotides are complementary nucleotides to the HIV genome that prevent transcription of the viral DNA to viral mRNA. Oligondeoxynucleotides may also bind to viral mRNA, forming an RNAIDNA hybrid, preventing translation of the viral mRNA into protein. Oligodeoxynucleotides have been shown to inhibit HIV replication in ~ i v o . ~Therapeutic ~.~' utility of antisense oligonucleoties face serious developmental challenges. The small (15 to 20 nucleotides) must traverse the plasma, contact and then penetrate into cells, resist premature degradation by nucleases, and ultimately reach their target sequences at the nucleus. Despite these formidable problems, antisense-based therapy is undergoing active development and will probably lead to clinical trials. GLQ223 is a highly purified formulation of trichosanthin, a basic (pl 9.6) 27 kd protein consisting of 247 amino acids that is purified from the root tubers of Trichosanthes kirilowii, M a x i m o ~ i c z . ~ In ~ - ~China ' single doses of trichosanthin-containing products have been used as a midtrimester abortifacient and multiple doses have been used for the treatment of trophoblastic tum o r ~ . ~ In ~ ' .vitro ~ ' studies suggest that trichosanthin selectively inhibits the replication of HIV in lymphoid and mononuclear cells.52On the basis of structural and functional properties, trichosanthin is considered a member of the family of single-chain ribosome inactivating prot e i n ~ . ~ ~The . ~ ' -selective ~~ inhibition of HIV-infected cells is probably a post-translational event. Data from the initial Phase I evaluation indicated that bolus infusion of GLQ223 provided serum concentrations of study drug that predicted effects; however, the duration of GLQ223
concentration was inadequate to observe anti-HIV eff e c t ~ Computer .~~ modeling of pharmacokinetic data from the single infusion study suggested that a brief bolus followed by a prolonged infusion would achieve serum concentrations that would reproduce in vitro drug concentrations that exerted anti-HIV effects. In order to evaluate the pharmacokinetic profile of GLQ223 during prolonged administration, a second phase 1 study was begun and completed at San Francisco General Hospital. GLQ223 was administered as a loading dose followed by a 3-hour infusion. Each subject received a test dose of 50 ng of GLQ223 prior to study drug administration. Subjects received weekly infusions of GLQ223 for 4 consecutive weeks. This was followed by a 21-day follow-up period during which no study drug was administered. All subjects were monitored for tolerance and toxicity. GLQ223 administration was not associated with notable toxicity except for a flulike syndrome characterized by muscle aches, throat pain, low-grade fevers, and chills associated with an increase in the CPK (MM fraction) isoenzyme level. This syndrome was easily controlled. Serum concentrations of GLQ223 were achieved in the two highest doses that were comparable to concentrations shown to have anti-HIV activity. Immunologic parameters suggest that GLQ223 has dose-dependent activity in HIV-infected individuals. A Phase 213 study comparing clinical and immunologic events in subjects with 200 to 500 CD4 ' cells and at least 9 months of ZDV therapy has been initiated. The subjects are randomized to one of three treatment arms, ZDV alone, GLQ223 alone, and ZDV plus GLQ223. GLQ223 is administered as a 3-hour intravenous infusion. Sixteen infusions are planned and all subjects will be followed for 48 weeks. Rational drug design has had the greatest role in developing compounds that inhibit the viral enzyme HIV protease. HIV protease recognizes as substrate the peptide bond between tyrosine (phenylalanine) and proline .57-59 The crystal structure of protease enzyme has been obtained. Inhibitors to protease have been designed based on the three-dimensional structure of the enzyme. Potent inhibitors have been found from large chemical catalogues and improvements have been designed in a few promising inhibitor candidates. As a group, these compounds inhibit HIV-1, HIV-2, and SIV. They have a poor oral bioavailability profile but potent in vitro antiHIV effects. Phase 1 clinical trials of protease inhibitors have begun in Europe. Protease inhibitors have been de-
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"BID: twice daily; CBC: complete blood count; GI: gastrointestinal; HIV: human immunodeficiency virus: LFT: liver function tests: TID: three times daily: ZDV: zidovudine.
SEMINARS IN LIVER DISEASE-VOLUME
veloped for clinical medicine. There is clinical precedent for the development of this type of compound, for example, angiotension converting enzyme inhibitors. The attachment of myristic acid to the NH2 terminal of the gag and gag-pol precursor protein and glycosylation of the viral envelope proteins are critical factors for the maturation of viral proteins and the viability of new virions. Products that interfere with the normal sugar attachment to viral proteins may therefore represent a class of compounds that selectively inhibit HIV activity. Castanospermine and deoxynorjirimycin, (which inhibit glycosylation) reactions catalyzed by alpha-glucosideases I and 11, are only weakly active against HIV, probably because they are not HIV s p e ~ i f i c . ~Compounds ~,~' that inhibit the myristylation of p17 gag protein interfere with the assembly of HIV and mature infectious virion^,^^.^' and thus may provide a useful therapeutic option. A final area of anti-HIV research is the development of HIV vaccines to stimulate the immune system of HIV-infected patients. United States Army data have already been presented that indicate that monthly vaccination with a prototype HIV gp160 vaccine can induce new specific HIV antibodies and new T-cell functional response^.^ Those individuals with new antibody and cellular responses have developed stabilization of their CD4 lymphocyte counts compared with individuals who failed to develop these new responses. Other studies of vaccine constructs in seropositive individuals have begun, including a Phase 112 study at San Francisco General Hospital. The exact mechanism of action for postinfection vaccination-inducing immunologic stabilization is not known, however this strategy appears to have limited toxicity. Well-controlled clinical trials will define the role of vaccination for infected individuals.
SUMMARY Knowledge of the HIV life cycle helps to plan rational modalities directed against the virus. The complex nature of the life cycle of HIV presents multiple unique targets. HIV binding to the cellular receptor CD4, viral enzymatic targets, HIV activation, viral protein synthesis, and protein packaging are examples of the types of targets available to inhibit the life cycle of HIV. Patients, community groups, regulatory agencies, national research groups, and the private pharmaceutical companies have joined forces to work to a common goal, effective HIV therapies. RT inhibitors have provided the best therapeutic options, but other drugs are being developed at a rapid pace. New drugs are entering clinical investigation and will be more widely available at early signs of clinical and laboratory effectiveness.
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ANTI-HIV THERAPEUTICS-KAHN
