Phase I Studies of Hypericin, the Active Compound in St. John's Wort, as an Antiretroviral Agent in HIV-Infected Adults: AIDS Clinical Trials Group Protocols 150 and 258

Hypericin is a compound found in the plant St. John's wort (Hypericum perforatum). It has received attention recently both in the medical literature (1, 2) and elsewhere (3) as a natural antidepressant. Hypericin or St. John's wort is taken commonly by HIV-infected persons (4), and studies have noted immunologic and clinical benefits (5-7). Hypericin has activity against many viruses in vitro (8), including HIV (9-13), and has shown in vivo activity in three murine retroviral systems (9). The compound appears to inactivate free virions and interfere with steps in the replicative cycle (9-13). On the basis of in vitro antiretroviral activity and community use, we evaluated hypericin given intravenously (AIDS Clinical Trials Group [ACTG] protocol 150) and orally (ACTG protocol 258) to HIV-infected patients. Methods Patients We enrolled HIV-infected adults with CD4+ lymphocyte counts less than 300 cells/mm3 for ACTG protocol 150 ( 350 cells/mm3 for ACTG protocol 258). Patients also had a hemoglobin value of 95.0 g/L or more for ACTG protocol 150 ( 80.0 g/L for ACTG protocol 258) and a leukocyte count greater than 2000 cells/mm3 for ACTG protocol 150. For both protocols, patients had a neutrophil count of 1000 cells/mm3 or more, a platelet count of 75 000 cells/mm3 or more, a serum creatinine concentration of 132.6 mol/L or less ( 1.5 mg/dL), hepatic aminotransferase and alkaline phosphatase levels 5 times the normal value, a bilirubin level of 25.65 mol/L or less ( 1.5 mg/dL), and an amylase level less than 1.5 times the normal value. Initially, ACTG protocol 150 required a screening HIV p24 antigen level of 70 pg/mL or more; this requirement was later eliminated. In addition, ACTG protocol 258 required a screening HIV p24 antigen level 35 pg/mL or more. Patients were excluded if they had significant medical illnesses or were taking monoamine oxidase inhibitors or hypertension-inducing, nephrotoxic, or hepatotoxic drugs within 14 days of study entry; antiretroviral agents or immunomodulatory agents within 1 month of study entry; opiates; or drugs known to induce photosensitivity. The institutional review boards of each individual site approved the study, and all patients gave informed consent. Study Drug Hypericin was synthesized de novo from emodin (14), which was provided by VimRx Pharmaceuticals (Wilmington, Delaware), and was filtered and lyophilized from 2.5% benzyl alcohol. Within 18 hours of administration, 40 mg of hypericin was dissolved in 36 mL of 2% benzyl alcohol and 3.6 mL of 50% dextrose solution, giving a final concentration of 1.9% benzyl alcohol, 4.5% dextrose, and 1.01 mg of hypericin per mL. Benzyl alcohol has been given previously to adults without toxic effects (15). Analogous procedures were used to reconstitute a 2-mg/mL solution for oral administration. The solution was stable at room temperature for 14 days. Study Design Four ACTG units participated in this open-label, dose-escalation study of intravenous (ACTG protocol 150) or oral (ACTG protocol 258) hypericin. Four cohorts of patients receiving intravenous hypericin were planned: 0.25, 0.5, 1.0, and 2.0 mg/kg of body weight administered twice weekly. Eight patients completed 3 weeks of therapy before entering the next cohort. The maximum tolerated dose was defined as the dose preceding the dose at which five or more patients had serious toxic reactions. When the maximum tolerated dose in the twice-weekly cohort was reached, a protocol amendment added cohorts of 0.25 and 0.375 mg/kg administered three times weekly. Four cohorts of patients receiving oral hypericin, starting with 0.5 mg/kg daily, were also planned. Treatment and Follow-up Patients were evaluated at baseline with a history, physical examination, and laboratory examinations. Patients were enrolled sequentially into cohorts. They received therapy for 8 weeks and could then continue treatment for up to 24 weeks. Patients were evaluated weekly for adverse events. The antiretroviral activity of hypericin was assessed by using baseline and biweekly CD4 lymphocyte counts and quantitative HIV peripheral blood mononuclear cell cultures and weekly p24 antigen determinations through week 8. Procedures approved by the ACTG were used (Division of AIDS, Rockville, Maryland). Plasma samples for HIV RNA (Amplicor HIV Monitor test, Roche Diagnostic Systems, Branchburg, New Jersey; lower limit, 200 HIV RNA copies/mL) were obtained at baseline and biweekly from patients enrolled in ACTG protocol 150. Pharmacologic measurements were performed (data presented elsewhere). Patients were evaluated 4 weeks after their last dose of hypericin. Toxicity The ACTG toxicity scale was used to achieve uniformity in reporting. Because of a newly observed toxic effect, a scale for phototoxicity was implemented during the study: grade 1, erythema, numbness, or pain causing mild discomfort; grade 2, erythema, numbness, pain, or temperature sensitivity that caused tolerable discomfort but required non-narcotic analgesia; grade 3, intolerable erythema or numbness (or both), temperature sensitivity, and pain despite long-term analgesic therapy; and grade 4, severe erythema with risk for desquamation or ulceration or severe intolerable pain of long duration despite analgesic therapy. Statistical Analysis A positive response in HIV p24 antigen level was defined as a decrease of 50% or more from the baseline level. Changes from baseline were evaluated by using the Wilcoxon signed-rank tests or t- test, as appropriate. Log transformations were used where necessary to more closely satisfy normality assumptions. All P values were two-tailed. The estimated toxicity and activity response rates were summarized by exact binomial CIs. Results Study Sample and Follow-up Thirty patients were enrolled into three cohorts that received intravenous hypericin and one cohort that received oral hypericin (Table). Of these patients, 16 (53%) discontinued treatment before completing 8 weeks of therapy because of grade 2 or 3 (moderate or severe) phototoxicity; 1 patient discontinued treatment voluntarily (Figure). Nine of the 13 remaining patients continued treatment after 8 weeks; only 2 patients completed 24 weeks of therapy. Table. Baseline Characteristics Figure. Disposition of the study patients. Toxicity Toxicity rates were defined as the percentage of patients who experienced a phototoxic reaction or at least one grade 3 or 4 toxic reaction that recurred at the same or lower grade, excluding baseline events. Patients were considered unevaluable for grade 3 or 4 toxicity summary if they permanently discontinued therapy with the study drug before completing 8 weeks of treatment for reasons other than serious (grade 3 or higher) toxic reactions. Almost all patients undergoing hypericin therapy experienced phototoxicity of grade 2 or higher: 8 of 11 (73%) receiving 0.25 mg/kg intravenously twice weekly, 7 of 7 (100%) receiving 0.5 mg/kg intravenously twice weekly, 8 of 9 (89%) receiving 0.25 mg/kg intravenously three times weekly, and 3 of 3 (100%) receiving 0.5 mg/kg orally each day. Grade 3 phototoxicity was observed in 11 of 23 (48% [95% CI, 27% to 69%]) evaluable patients: 2 of 8 (25%) receiving 0.25 mg/kg intravenously twice weekly, 5 of 6 (83%) receiving 0.5 mg/kg intravenously twice weekly, 1 of 6 (17%) receiving 0.25 mg/kg intravenously three times a week, and 3 of 3 (100%) receiving 0.5 mg/kg orally each day. The toxic reaction was an erythematous rash associated with painful dysesthesias that involved areas exposed to light; it was thus consistent with a cutaneous phototoxic reaction. This reaction resolved after discontinuation of hypericin therapy. No grade 4 phototoxicity occurred. Our data defined 0.25 mg/kg as the maximum tolerated dose for twice-weekly intravenous administration. No maximum tolerated dose could be determined for three-times-weekly intravenous or oral administration. Other serious clinical or laboratory adverse events occurred infrequently. One patient each had fever and diarrhea, headache, hemoglobin value less than 80.0 g/L, and glucose level greater than 13.88 mmol/L (250 mg/dL); two patients had elevation of alkaline phosphatase levels more than 5 times the normal value; and three patients had elevation of hepatic aminotransferase levels more than 5 times the normal value. Virologic and Immunologic Activity When we pooled the results across cohorts, 3 of 16 patients (19% [CI, 4% to 46%]) had a positive HIV p24 antigen response. No detectable change in HIV titer was seen from baseline to week 8 (median, 0.27 log10 titer; P=0.19, Wilcoxon signed-rank test). Among the 14 patients from ACTG protocol 150 with week 8 measurements, no change in HIV RNA log10 copies/mL from baseline to week 8 was detected (P>0.2, t-test). The median decrease in HIV RNA level was 0.07 log10 copies/mL. Among the 15 patients in the pooled cohort, a median decrease of 10 CD4 cells/mm3 (P=0.02) was seen from baseline to week 8. Discussion Hypericin therapy induced serious cutaneous phototoxicity and lacked demonstrable antiretroviral activity at the doses tested in our study of HIV-infected adults. These findings raise concerns about the safety of hypericin, which is available as an herbal preparation (1-3) and is commonly used by HIV-infected patients (4). Previous studies of hypericin and St. John's wort in both HIV-infected and non-HIV-infected patients noted a relative lack of side effects, although the doses were lower and the preparations were less pure than those used in the current study (1, 6, 7, 16, 17). Preparations containing hypericin are largely unregulated and contain varying amounts of drug (2). Persons taking herbal supplements and physicians need to be aware of the potential for phototoxicity. The photodynamic properties of hypericin have been described (18). Livestock who feed on hypericin have developed skin rashes termed hypericism (19). Animal hypericism ranges from erythema of sun-exposed skin to ex

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