Aim stated in the publication

“To investigate the efficacy and safety of pritelivir in the treatment of genital infection with HSV-2 and to determine the most effective, safe dose, we conducted a proof-of-concept study to evaluate the effect of four different doses of oral pritelivir on mucocutaneous viral shedding in adults with genital HSV-2 infection.” (opens the source at this quote in a new tab)
“We tested the efficacy and safety of pritelivir in otherwise healthy persons with genital HSV-2 infection.” (opens the source at this quote in a new tab)

Aim stated in the registry record

“The aim of the study is to find out whether AIC316 is safe and efficacious for the prevention of reactivation of genital herpes” (opens the source at this quote in a new tab)

Secondary objective: monitoring for molecular signals of drug resistance

“Here we report the results of a secondary objective of this trial: the monitoring of molecular signals of drug resistance over 28 days of therapy.” (opens the source at this quote in a new tab)

Sites and conduct period in the publication

“The trial was conducted between April and December 2010 at seven sites in the United States.” (opens the source at this quote in a new tab)

Study dates and status in the registry record

Field	Registry entry
Recruitment status	“Completed”
Study start	“2010-03”
Primary completion (actual)	“2010-12”
Study completion (actual)	“2011-03”
Results first posted	“2022-02-04”
Last update posted	“2023-04-21”

Study locations in the registry record

“This study has 6 locations” (opens the source at this quote in a new tab)

Location	Facility
“Miami, Florida, United States, 33169”	“Cetero Research Miami Gardens”
“Indianapolis, Indiana, United States, 46202”	“Indiana University School of Medicine, Office for Research”
“Portland, Oregon, United States, 97210”	“Westover Heights Clinic”
“Houston, Texas, United States, 77030”	“University of Texas Health Science Centre, Center for Clincial Studies”
“Salt Lake City, Utah, United States, 84132”	“University of Utah”
“Seattle, Washington, United States, 98104”	“University of Washington, Virology Research Clinic”

Randomized, parallel, double-blind, placebo-controlled design

“The study was designed as a randomized, parallel, double-blind, placebo-controlled study by investigators at the University of Washington and by the sponsor, AiCuris, and was managed by FHI 360.” (opens the source at this quote in a new tab)

Randomization ratio, method, and stratification

“Eligible participants were randomly assigned to one of the five study groups in a 1:1:1:1:1 ratio. A statistician who was not involved in the conduct of the study generated the randomization schema using the permuted-blocks method, with stratification according to sex and study site.” (opens the source at this quote in a new tab)

Daily genital swabs, symptom diary, and clinic visits for recurrences

“Throughout the study period, participants obtained daily swabs of genital skin and mucosa and delivered them to the study site weekly for the purpose of HSV detection.” (opens the source at this quote in a new tab)
“Participants also maintained a diary of genital signs and symptoms. If a lesion developed that was consistent with genital herpes, an additional swab was obtained from the lesion. Participants who had a recurrence (i.e., papules, vesicles or pustules, ulcers, or crusts) during the study were asked to return to the clinic for an examination within 24 hours.” (opens the source at this quote in a new tab)

HSV DNA assay, positivity threshold, and threshold for resistance sequencing

“Samples of genital secretions were analyzed for HSV DNA with a real-time, quantitative, fluorescent polymerase-chain-reaction (PCR) assay (TaqMan) at the University of Washington Virology Laboratory.” (opens the source at this quote in a new tab)
“Test results were considered to be positive when 150 HSV DNA copies per milliliter or more were detected.” (opens the source at this quote in a new tab)
“Samples with an HSV DNA copy number of more than 5000 per milliliter were sequenced to detect mutations in UL5 and UL52.” (opens the source at this quote in a new tab)

Safety evaluation and adverse event grading

“Safety was evaluated by examining the rate, nature, duration, and severity of adverse events in each study group; events were graded with the use of the Division of Acquired Immunodeficiency Syndrome (DAIDS) table of adverse events” (opens the source at this quote in a new tab)
“Additional safety assessments included physical examination, laboratory measurements, and electrocardiography.” (opens the source at this quote in a new tab)

Design fields in the registry record

Field	Registry entry
Official title	“A Double-blind Randomized Placebo Controlled Dose-finding Trial to Investigate Different Doses of a New Antiviral Drug in Subjects With Genital HSV Type 2 Infection”
Other study identifier	“AIC316-01-II-01”
Phase	“Phase 2”
Allocation	“Randomized”
Interventional model	“Parallel Assignment”
Masking	“Quadruple”
Primary purpose	“Prevention”
Sponsor	“AiCuris Anti-infective Cures AG”
Collaborator	“FHI 360”

Statistical analysis responsibility, data access, and funding

“The statistical analyses were performed by authors at the University of Washington, according to a statistical plan previously developed by the academic and industry investigators. Both the academic and industry authors contributed to all drafts of the manuscript and approved the submission of the final manuscript for publication.” (opens the source at this quote in a new tab)
“Funded by AiCuris; ClinicalTrials.gov number, NCT01047540.” (opens the source at this quote in a new tab)

Resistance sequencing substudy: regions sequenced and sample thresholds

“The sequencing covered a 300–base pair region in UL5 (nucleotides 841–1140) and a 200–base pair region in UL52 (nucleotides 2601–2800). Only samples containing ≥5000 copies/mL were selected for sequencing.” (opens the source at this quote in a new tab)
“Full UL5 and UL52 genes were sequenced from participants with an HSV DNA level of ≥50 000 copies/mL. In participants with multiple samples, only the first and last samples were selected for sequencing.” (opens the source at this quote in a new tab)

Mathematical modeling study of the trial's shedding data: purpose

“The purpose of this study was to develop a mathematical model of HSV-2 pathogenesis during pritelivir therapy that captures the fundamental features of viral replication and spread, the cytolytic immune response against infected cells, and PK/PD of pritelivir. We first developed the model and tested its ability to reproduce the most relevant dynamic features of HSV-2 shedding in each of five treatment arms in a Phase 2 clinical trial.” (opens the source at this quote in a new tab)

Inclusion criteria in the publication

“We enrolled otherwise healthy adults (≥18 years of age) who were seropositive for HSV-2 and had a history of genital herpes. Eligible participants had a history of one to nine recurrences of genital herpes per year. Other inclusion criteria were a body-mass index (the weight in kilograms divided by the square of the height in meters) between 18 and 35, the use of effective contraception, and normal results on laboratory tests, including electrocardiography and urinalysis.” (opens the source at this quote in a new tab)

Exclusion criteria and prohibited concomitant treatments in the publication

“Infection with the human immunodeficiency virus or with hepatitis B or C virus, pregnancy, immunosuppression, treatment with medications that induce drug-metabolizing enzymes or transporters (e.g., carbamazepine or phenytoin) were exclusion criteria. Receipt of other anti-HSV therapy, consumption of grapefruit, and intake of quinine or products containing quinidine were not permitted during the study.” (opens the source at this quote in a new tab)

Inclusion criteria in the registry record

“Adult, Immunocompetent men and women in good health of any ethnic group” (opens the source at this quote in a new tab)
“History of recurrent episodes of genital herpes for at least 12 months” (opens the source at this quote in a new tab)
“Seropositive for Herpes Simplex Virus HSV Type 2” (opens the source at this quote in a new tab)
“Body Mass Index (BMI) between 18 and 35 kg/m2” (opens the source at this quote in a new tab)

Exclusion criteria in the registry record

“Present episode of genital herpes” (opens the source at this quote in a new tab)
“Intake of systemic drug against HSV or any topical application against HSV within 7 days before randomization for the trial” (opens the source at this quote in a new tab)
“Intake of systemic corticosteroids, other immunomodulating agents or any investigational agent within 3 months before randomization for the trial” (opens the source at this quote in a new tab)
“Positive results in any of the virology tests for human immunodeficiency virus antibody (HIV-Ab), hepatitis C antibody (HCV-Ab), hepatitis B surface antigen (HBsAg) or hepatitis B core antibody (HBc-Ab)” (opens the source at this quote in a new tab)

Age, sex, and healthy volunteer fields in the registry record

Field	Registry entry
Ages eligible for study	“18 Years and older (Adult, Older Adult )”
Sexes eligible for study	“All”
Accepts healthy volunteers	“No”

Four oral pritelivir regimens and placebo for 28 days

“The four dosing regimens for pritelivir were a loading dose of 20 mg followed by a daily dose of 5 mg, a loading dose of 100 mg followed by a daily dose of 25 mg, a loading dose of 300 mg followed by a daily dose of 75 mg, and a weekly dose of 400 mg. The regimen for the administration of placebo was the same as it was for the administration of pritelivir. The study drug was taken for a period of 28 days.” (opens the source at this quote in a new tab)

Arm descriptions in the registry results

Arm	Description
Dose regimen 1	“AIC316: Day 1 20 mg, Day 2-28 5 mg QD, Oral administration”
Dose regimen 2	“AIC316: Day 1 100 mg, Day 2-28 25 mg QD, Oral administration”
Dose regimen 3	“AIC316: Day 1 300 mg, Day 2-28 75 mg QD, Oral administration”
Dose regimen 4	“AIC316: 400 mg QWK, Oral administration”
Placebo	“Matching placebo: Oral administration”

Loading doses by arm as described in the modeling study

“To rapidly achieve representative drug concentrations, participants in the 5 mg arm (n=33) received an initial loading dose of 20 mg, participants in the 25 mg arm (n=32) received an initial loading dose of 100 mg and participants in the 75 mg arm (n=29) received an initial loading dose of 300 mg. No loading dose was administered in the 400 mg weekly arm (n=31).” (opens the source at this quote in a new tab)

Adherence to swab collection and study medication

“The median percentage of days on which swabs were obtained was 100% (range, 11 to 100%); 80% of the participants obtained more than 96% of the swabs. Mean adherence to the study medication, as calculated according to the pill count, was 99% (median, 100%; range, 72 to 100) with 94% of participants taking more than 95% of the study drug; adherence was similar in all study groups.” (opens the source at this quote in a new tab)

Primary endpoint defined in the publication

“The frequency of HSV-2 detection (the shedding rate) was defined as the number of days with a genital swab that was positive for HSV divided by the total number of days on which genital swabs were obtained. The primary end point was the reduction in the HSV shedding rate among participants receiving pritelivir relative to the shedding rate among participants receiving placebo.” (opens the source at this quote in a new tab)

Primary outcome measure in the registry record

“Efficacy of 4 Different Doses of AIC316 and Matching Placebo With Respect to the Suppression of Herpes Simplex Virus Replication (Shedding Rate)” (opens the source at this quote in a new tab)
“Subjects were assessed for shedding rate as the percentage of number of swab days with at least one swab positive for HSV DNA relative to the total number of days with analyzable swabs. Measurement was based on presence of HSV DNA measured on the material collected from swabs taken at the visits by the investigator and daily swabbing of the anogenital region by the patient during the treatment and post-treatment period.” (opens the source at this quote in a new tab)

Rationale given by the investigators for a shedding endpoint

“Our study design, in which the number of days of viral shedding was the primary outcome, provided an approach toward defining in vivo antiviral efficacy and an effective, safe dose for subsequent evaluation. Studies of the natural history of HSV-2 have shown that shedding frequency, the quantity of HSV during shedding, and the occurrence and duration of genital lesions are all correlated.” (opens the source at this quote in a new tab)
“Early studies in the development of nucleoside analogues for the treatment of genital herpes measured the reduction in the duration of symptomatic recurrences. However, lesions reflect mucosal damage as well as viral replication, and even effective antiviral agents have marginal benefits when started after the formation of lesions. Evaluations of viral shedding have shown that nearly all persons with HSV-2 shed virus.” (opens the source at this quote in a new tab)
“We selected a patient population with clinically symptomatic disease to obtain a preliminary assessment of both the effects of extended antiviral therapy on shedding and the potential clinical benefit of pritelivir.” (opens the source at this quote in a new tab)

Lesion endpoints in early-phase trials of HSV antivirals

“However, the use of genital herpes lesions as an end point in designing proof-of-concept studies and dose-ranging studies in early development of anti-HSV therapeutic agents is challenging because genital herpes lesions are highly variable in frequency and duration” (opens the source at this quote in a new tab)
“While evaluation of a clinically meaningful end point such as lesions or recurrence rate is currently required prior to approval of new HSV interventions (ie, vaccines and antivirals), the use of HSV detection at mucosal sites, or viral shedding, could be instead used as the surrogate outcome, particularly for early phase evaluations.” (opens the source at this quote in a new tab)

Surrogate outcome analysis: question and data sources

“We tested whether genital herpes simplex virus (HSV) shedding is an appropriate surrogate outcome for the clinical outcome of genital herpes lesions in studies of HSV-2 antiviral interventions.” (opens the source at this quote in a new tab)
“We analyzed prospective data from natural history studies and clinical trials of antiviral agents for HSV-2 in which HSV-2–seropositive participants provided self-collected anogenital swab specimens daily over ≥25 days for HSV DNA quantitation by polymerase chain reaction (PCR). Genital recurrences were self-reported.” (opens the source at this quote in a new tab)

Correlation of shedding rate with lesion rate in a natural history cohort

“Among 674 participants, genital HSV shedding was detected on 17% of days, and genital lesions were reported on 10% of days. Within the same session, HSV shedding rates were strongly correlated with lesion rates (ρ = 0.61, P < .0001).” (opens the source at this quote in a new tab)
“In these 90 pairs of sessions, the HSV shedding rate was consistent within individuals (ρ = 0.39, P < .0001; Figure 2A).” (opens the source at this quote in a new tab)
“Viral shedding at an earlier session was also strongly correlated with the lesion rate at a subsequent session (ρ = 0.41, P < .0001).” (opens the source at this quote in a new tab)

Proportion of the pritelivir effect on recurrences and lesions explained by shedding

“After adjustment for sex and study site, the risk ratio for HSV recurrence while receiving pritelivir versus placebo was 0.28 (95% CI, .08–.95; P = .041; Table 2). When the HSV shedding rate, the candidate surrogate outcome, was added to the model, the risk ratio for recurrence increased to 0.79 and was no longer statistically significant (95% CI, .19–3.32; P = .75; Table 2). Comparison of the effect of pritelivir on the recurrence rate between these 2 models yielded a PTE of 0.82: 82% of the effect of pritelivir on the recurrence frequency was explained by its effect on HSV shedding.” (opens the source at this quote in a new tab)
“When the shedding rate was additionally included as a predictor of the lesion rate, the risk ratio for pritelivir increased to 0.58 (95% CI, .18–1.91; P = .37; Table 2). Comparison of the treatment effect between these 2 models yielded an estimated PTE of 0.74: 74% of the effect of pritelivir on the genital lesion frequency was due to its effect on HSV shedding.” (opens the source at this quote in a new tab)

Conclusion of the surrogate outcome analysis

“HSV genital shedding measured by PCR analysis in swab specimens self-collected daily is an appropriate surrogate outcome for genital herpes lesions because it is in the causal pathway to recurrences.” (opens the source at this quote in a new tab)
“Overall, 40%–82% of the antiviral effect on recurrences was explained by its effect on HSV shedding.” (opens the source at this quote in a new tab)

Limits of shedding as a surrogate outcome

“To have research value, the HSV shedding rate need not be a perfect surrogate for lesion or recurrence rate, as new candidate treatments initially evaluated with the shedding rate as an outcome may be further evaluated for clinical outcomes, such as the recurrence rate prior to licensure. We note that there was residual association between treatment and clinical outcome for acyclovir and valacyclovir, indicating that shedding does not explain 100% of the effect of treatment on lesions or recurrence.” (opens the source at this quote in a new tab)
“Our study may be limited because recurrence and lesion rates were based on self-reports from participants. Given that lesions can be difficult to detect, their frequency may be underestimated.” (opens the source at this quote in a new tab)

Secondary endpoints

“Secondary end points included the rate of genital lesions, the reduction of HSV DNA copies, the frequency of herpes recurrence, and the rate of subclinical shedding.” (opens the source at this quote in a new tab)

Participants screened, randomized, and treated

“Among 261 persons screened, 156 underwent randomization; 1 person who underwent randomization was deemed ineligible and did not receive the study drug or participate in the study.” (opens the source at this quote in a new tab)

Enrollment in the registry record

Field	Registry entry
Enrollment (actual)	“156”

Participants started and in the full analysis set by arm, registry results

Milestone	Dose regimen 1 (5 mg daily)	Dose regimen 2 (25 mg daily)	Dose regimen 3 (75 mg daily)	Dose regimen 4 (400 mg weekly)	Placebo
Started	“34”	“32”	“29”	“31”	“30”
Full analysis set	“33”	“32”	“29”	“31”	“30”
Analyzed for the primary outcome	“33”	“32”	“29”	“31”	“30”

Swabs and participants contributing to the shedding analysis

“A total of 4180 daily genital swabs were obtained, 435 of which (10.4%) were positive for HSV. Genital HSV was detected at least once in 87 of the 155 participants (56%).” (opens the source at this quote in a new tab)

Swab specimens available to the resistance sequencing substudy

“8993 genital swab specimens were collected from 155 eligible participants.” (opens the source at this quote in a new tab)

Participant count as described in the modeling study

“We analyzed results from a randomized, double-blind phase 2 study, in which 150 participants were distributed among 5 doses (5 mg, 25 mg and 75 mg daily; 400 mg weekly; placebo)” (opens the source at this quote in a new tab)

Intention-to-treat analysis population

“The intention-to-treat analysis included all patients who took at least one dose of study medication and obtained at least one swab of genital secretions.” (opens the source at this quote in a new tab)

Statistical model for shedding and lesion rates

“Poisson regression with overdispersion was used to estimate the effect of treatment on the rates of viral shedding and lesion recurrence, with terms included for treatment and sex and a random term for investigational site.” (opens the source at this quote in a new tab)

Comparison of HSV DNA quantity and concordance of swab sources

“On days when HSV was detected, the log10 number of HSV DNA copies was compared between each of the pritelivir groups and the placebo group with the use of generalized estimating equations that accounted for the correlation between repeated measures per participant.” (opens the source at this quote in a new tab)
“The concordance between the detection of HSV from swabs obtained by a participant and detection from those obtained by an investigator was assessed with the kappa statistic and the Spearman coefficient.” (opens the source at this quote in a new tab)

Baseline analysis population in the registry results

Field	Registry entry
Baseline analysis population description	“Based on Full Analysis Set”
Overall number of baseline participants	“155”

Resistance sequencing substudy: participants and acceptance rule

“For all participants who received at least 1 dose of trial medication (placebo or pritelivir) and from whom an adequate yield of viral DNA was available, we sequenced the regions of UL5 and UL52 genes known to carry mutations mediating phenotypic resistance to HPIs in vitro” (opens the source at this quote in a new tab)
“Sequence results were accepted only if all bases in the region were sequenced at least twice by independent sequencing reactions.” (opens the source at this quote in a new tab)

Modeling study: outcome definitions and comparisons between arms

“Shedding rate was defined as number of swabs with HSV DNA detected by PCR out of the total number of swabs collected. Lesion rate was number of days with reported lesions out of total number of days with reports.” (opens the source at this quote in a new tab)
“Rate outcomes were compared by arm between persons using generalized estimating equations models with a log link, to estimate risk ratios for shedding rates, lesion rates, and episode rates by study arm.” (opens the source at this quote in a new tab)

Completion and reasons for discontinuation in the publication

“A total of 147 participants (94%) completed the study. Participants discontinued treatment prematurely for the following reasons: withdrawal of consent (4 participants), adverse events (3 participants), loss to follow-up (2 participants), pregnancy (1 participant), and noncompliance with the protocol (1 participant).” (opens the source at this quote in a new tab)

Reason for ineligibility at screening

“The predominant reason for ineligibility was an absence of serologically confirmed HSV-2 infection.” (opens the source at this quote in a new tab)

Participant flow by arm in the registry results

Milestone	Dose regimen 1 (5 mg daily)	Dose regimen 2 (25 mg daily)	Dose regimen 3 (75 mg daily)	Dose regimen 4 (400 mg weekly)	Placebo
Started	“34”	“32”	“29”	“31”	“30”
Completed	“29”	“32”	“26”	“31”	“29”
Not completed	“5”	“0”	“3”	“0”	“1”

Reasons for not completing by arm in the registry results

Reason	Dose regimen 1 (5 mg daily)	Dose regimen 2 (25 mg daily)	Dose regimen 3 (75 mg daily)	Dose regimen 4 (400 mg weekly)	Placebo
Protocol violation	“1”	“0”	“0”	“0”	“0”
Lost to follow-up	“1”	“0”	“0”	“0”	“0”
Withdrawal by subject	“1”	“0”	“3”	“0”	“1”
Pregnancy	“1”	“0”	“0”	“0”	“0”
Adverse event	“1”	“0”	“0”	“0”	“0”

Age, sex, race, duration of genital herpes, and prior suppressive therapy

“The median age of the participants was 41 years; 104 (67%) were women, and most were white (Table 1). The median duration of genital herpes before study entry was 11 years; 21% of the participants were receiving suppressive therapy before study entry.” (opens the source at this quote in a new tab)

Demographics by arm in the registry results, full analysis set

Characteristic	Dose regimen 1 (5 mg daily), n = 33	Dose regimen 2 (25 mg daily), n = 32	Dose regimen 3 (75 mg daily), n = 29	Dose regimen 4 (400 mg weekly), n = 31	Placebo, n = 30	Total, n = 155
Age, mean (standard deviation), years	“37.7 (12.56)”	“38.8 (12.00)”	“41.6 (12.47)”	“42.0 (11.79)”	“43.1 (10.81)”	“40.5 (11.97)”
Female, count (%)	“22”( “66.7%”)	“21”( “65.6%”)	“20”( “69.0%”)	“20”( “64.5%”)	“21”( “70.0%”)	“104”( “67.1%”)
Male, count (%)	“11”( “33.3%”)	“11”( “34.4%”)	“9”( “31.0%”)	“11”( “35.5%”)	“9”( “30.0%”)	“51”( “32.9%”)
American Indian or Alaska Native, count (%)	“0”( “0.0%”)	“0”( “0.0%”)	“0”( “0.0%”)	“0”( “0.0%”)	“1”( “3.3%”)	“1”( “0.6%”)
Asian, count (%)	“0”( “0.0%”)	“0”( “0.0%”)	“0”( “0.0%”)	“1”( “3.2%”)	“0”( “0.0%”)	“1”( “0.6%”)
Native Hawaiian or other Pacific Islander, count (%)	“0”( “0.0%”)	“0”( “0.0%”)	“0”( “0.0%”)	“0”( “0.0%”)	“0”( “0.0%”)	“0”( “0.0%”)
Black or African American, count (%)	“8”( “24.2%”)	“5”( “15.6%”)	“4”( “13.8%”)	“5”( “16.1%”)	“7”( “23.3%”)	“29”( “18.7%”)
White, count (%)	“25”( “75.8%”)	“27”( “84.4%”)	“20”( “69.0%”)	“23”( “74.2%”)	“21”( “70.0%”)	“116”( “74.8%”)
More than one race, count (%)	“0”( “0.0%”)	“0”( “0.0%”)	“0”( “0.0%”)	“0”( “0.0%”)	“0”( “0.0%”)	“0”( “0.0%”)
Race unknown or not reported, count (%)	“0”( “0.0%”)	“0”( “0.0%”)	“5”( “17.2%”)	“2”( “6.5%”)	“1”( “3.3%”)	“8”( “5.2%”)

Characteristics of the placebo and 75 mg daily arms combined, from a later analysis of the trial data

Characteristic	Pritelivir trial participants, n = 59
Age, median (range), years	“42 (20–67)”
Male sex, number (%)	“18 (31)”
White race, number (%)	“41 (69)”
Other race, number (%)	“18 (31)”
HSV-2 antibody only, number (%)	“35 (59)”
HSV-1 and HSV-2 antibody, number (%)	“24 (41)”
Time since acquisition of 1 year or longer, number (%)	“56 (95)”
Time since acquisition unknown, number (%)	“3 (5)”
Years since acquisition, median (range)	“15 (1–40)”
Swabbing days, median (range)	“28 (8–29)”

“We included 2 study arms: placebo and 75 mg daily.” (opens the source at this quote in a new tab)

Primary endpoint: genital HSV shedding rate by dose

“The rate of HSV shedding in the placebo group was 16.6%; among the groups receiving pritelivir daily, the rate was 18.2% with 5 mg, 9.3% with 25 mg, and 2.1% with 75 mg; in the group receiving 400 mg weekly, the rate was 5.3% (Table 2 and Figure 1A).” (opens the source at this quote in a new tab)
“The relative risk of viral shedding with pritelivir as compared with placebo was 1.11 (95% confidence interval [CI], 0.65 to 1.87; P=0.70) at a daily dose of 5 mg, 0.57 (95% CI, 0.31 to 1.03; P=0.06) at a daily dose of 25 mg, 0.13 (95% CI, 0.04 to 0.38; P<0.001) at a daily dose of 75 mg, and 0.32 (95% CI, 0.17 to 0.59; P<0.001) at a weekly dose of 400 mg (Table 2).” (opens the source at this quote in a new tab)

Primary outcome in the registry results: mean per-participant shedding rate

Measure	Dose regimen 1 (5 mg daily)	Dose regimen 2 (25 mg daily)	Dose regimen 3 (75 mg daily)	Dose regimen 4 (400 mg weekly)	Placebo
Participants analyzed	“33”	“32”	“29”	“31”	“30”
Shedding rate, mean (standard deviation), percentage of swab days	“21.2 (23.79)”	“9.2 (12.61)”	“2.0 (4.00)”	“5.2 (6.50)”	“16.5 (18.61)”

Quantity of HSV DNA on days with detection

“When HSV was detected, the median log10 number of HSV DNA copies was 5.1 with placebo, 4.5 with 5 mg of pritelivir daily, 3.6 with 25 mg daily, 2.4 with 75 mg daily, and 3.6 with 400 mg weekly (P<0.001 for the comparisons of placebo with doses of pritelivir of 25 mg or more) (Table 2 and Figure 1B).” (opens the source at this quote in a new tab)

Subclinical shedding rate by dose

“The overall rate of subclinical shedding was 9.8% with placebo, 11.9% with 5 mg of pritelivir daily, 7.4% with 25 mg daily, 1.6% with 75 mg daily, and 4.8% with 400 mg weekly (Figure 1C). The relative risks of subclinical shedding at pritelivir doses of 5 mg daily, 25 mg daily, 75 mg daily, and 400 mg weekly were 1.26 (95% CI, 0.67 to 2.35; P=0.48), 0.73 (95% CI, 0.37 to 1.47; P=0.38), 0.16 (95% CI, 0.05 to 0.49; P=0.001), and 0.47 (95% CI, 0.24 to 0.91; P=0.02), respectively.” (opens the source at this quote in a new tab)

Shedding by sex and tests of interaction

“Pritelivir reduced viral shedding in both men and women. For example, among men in the 75-mg group, HSV shedding occurred on 3.2% of days as compared with 12.5% of days among men in the placebo group; among women in the 75-mg group, HSV shedding occurred on 1.6% of days as compared with 18.3% of days in the placebo group. No significant interaction was detected between treatment with 75 mg of pritelivir daily and sex, nor was there a significant interaction between treatment and site.” (opens the source at this quote in a new tab)

Days with genital lesions by dose

“Genital lesions were noted on 9.0% of days with placebo, 12.5% with 5 mg of pritelivir, 3.5% with 25 mg, 1.2% with 75 mg, and 1.2% with 400 mg (Figure 1D). The risk of genital lesions with pritelivir as compared with placebo was 1.39 (95% CI, 0.59 to 3.30; P=0.46) with 5 mg of pritelivir daily, 0.41 (95% CI, 0.16 to 1.09; P=0.07) with 25 mg daily, 0.13 (95% CI, 0.02 to 0.70; P=0.02) with 75 mg daily, and 0.13 (95% CI, 0.03 to 0.52; P=0.004) with 400 mg weekly.” (opens the source at this quote in a new tab)

Number of recurrences by dose

“In addition, we observed 12 recurrences in participants receiving placebo, 18 in participants receiving 5 mg of pritelivir daily, 12 in those receiving 25 mg daily, 3 in those receiving 75 mg daily, and 6 in those receiving 400 mg weekly (P=0.04 for the comparison of placebo with 75 mg of pritelivir daily).” (opens the source at this quote in a new tab)

Shedding and HSV DNA quantity on days with lesions

“In the placebo group, HSV was detected on 85.3% of days on which participants had lesions, as compared with 60.4% of days in the group receiving 5 mg of pritelivir daily, 50.0% in the group receiving 25 mg daily, 44.4% in the group receiving 75 mg daily, and 50.0% in the group receiving 400 mg weekly. The median HSV DNA log10 copy number was 6.1 among participants with lesions who were receiving placebo. The log10 copy number was decreased somewhat among participants receiving lower doses of pritelivir, reaching 2.9 at a dose of 75 mg daily (P<0.001 for the comparison of 75 mg of pritelivir daily with placebo) (Table 2).” (opens the source at this quote in a new tab)

Shedding, recurrence, and lesion rates with counts for the placebo and 75 mg daily arms

“Thirty participants receiving placebo shed HSV on 16.6% of days (138 of 833), and 29 participants receiving the most effective dose of pritelivir (75 mg daily) shed HSV on 2.1% of days (16 of 766). The recurrence rate was 5.4 cases per person-year in the placebo group and 1.4 cases per person-year in the pritelivir group (Figure 3A)” (opens the source at this quote in a new tab)
“The lesion rate in the pritelivir clinical trial was 9.0% (75 of 836 days) in the placebo group and 1.2% (9 of 775 days) in the pritelivir group (Figure 4A)” (opens the source at this quote in a new tab)

Dose response as summarized by the investigators

“Our study showed that pritelivir, an HSV-specific helicase–primase inhibitor, significantly reduces the frequency of genital HSV shedding and lesions in otherwise healthy men and women with genital HSV-2 infection. The effect was dose-related, with a daily dose of 75 mg having the greatest antiviral effect. Furthermore, at this dose, pritelivir reduced the quantity of HSV in breakthrough shedding by more than 98%.” (opens the source at this quote in a new tab)
“Although the HSV shedding rate was reduced by more than 85% at the highest doses of pritelivir, as compared with placebo, some breakthrough shedding remained.” (opens the source at this quote in a new tab)

Concordance of participant-collected and investigator-collected swabs

“There was a high concordance in the frequency with which HSV was detected and the quantity of HSV DNA detected in the swabs obtained by participants and the swabs obtained by the investigators (r=0.87, P<0.001) (Fig. S1 in the Supplementary Appendix).” (opens the source at this quote in a new tab)

Viral resistance: sequencing of UL5 and UL52 reported in the publication

“Analysis of the resistance-associated regions of UL5 and UL52 revealed variations in DNA in all study groups as compared with the HSV-2 reference sequence. Four changes in UL5 were identified in five participants. Among these four changes, three were synonymous (G879A [in a recipient of 5 mg of pritelivir daily], C996T [in a placebo recipient], and G1053A [in a recipient of 400 mg of pritelivir weekly]) and one was nonsynonymous (A1001G [in a recipient of 5 mg of pritelivir daily]); the corresponding amino acid change is H334R, which is not within or downstream of the fourth functional motif of UL5 and is therefore not considered to confer resistance to pritelivir.” (opens the source at this quote in a new tab)
“In UL52, two synonymous changes were identified in two participants (G2649A [in a placebo recipient] and C2676T [in a recipient of 25 mg of pritelivir daily]). All sequences within participants were identical.” (opens the source at this quote in a new tab)
“Sequencing of samples obtained before and after treatment suggests that drug resistance did not emerge, even in participants treated with the lowest doses of pritelivir.” (opens the source at this quote in a new tab)

Resistance sequencing substudy: samples sequenced in the resistance-associated regions

“Of these, 823 swab specimens (median time of collection after treatment initiation, 22 days; range, 1–44 days), collected from 87 subjects (20 in the placebo group and 67 in one of the 4 dosage groups; Supplementary Table 4), contained an HSV-2 quantity of ≥5000 copies/mL and were subjected to resistance-region sequencing. Of the 823 samples, 688 (84%), from 80 patients, underwent successful sequencing of the UL5 gene, and 775 (94%), from 83 patients, underwent successful sequencing of the UL52 gene (Figure 2).” (opens the source at this quote in a new tab)

Resistance sequencing substudy: findings in the resistance-associated regions

“We found no evidence that pritelivir induced known resistance-mediating mutations or for amino acid variation at other loci.” (opens the source at this quote in a new tab)
“All 23 HSV-2 samples from this participant, including the sample collected at baseline (day 1), before initiating pritelivir therapy, and the sample collected on day 2, after pritelivir initiation, had the same single nucleotide change and corresponding amino acid change relative to the H334R consensus.” (opens the source at this quote in a new tab)
“As this isolate was susceptible to pritelivir, the H334R mutation was not regarded to confer resistance (Figure 3).” (opens the source at this quote in a new tab)
“In the UL52 resistance region, a synonymous change to the consensus was observed in 2 participants (1 placebo recipient and 1 participant receiving pritelivir 25 mg once daily); no nonsynonymous changes were found in any of the samples.” (opens the source at this quote in a new tab)

Resistance sequencing substudy: whole-gene sequencing of UL5 and UL52

“Forty-five participants provided the remaining 203 samples that were eligible for whole-gene sequencing.” (opens the source at this quote in a new tab)
“Of the 75 available specimens, 73 (97%) were successfully sequenced for the entire UL5 gene.” (opens the source at this quote in a new tab)
“Only 2 participants, both assigned to the 5 mg once daily treatment group, had virus that showed any between-sample variation in the UL5 gene between the first and the last positive swab specimen” (opens the source at this quote in a new tab)
“Owing to the high GC content of the UL52 gene, only 46 of 75 specimens (61%) were successfully sequenced for UL52” (opens the source at this quote in a new tab)
“However, we confirmed that these differences reflect infection with dual HSV-2 strains, rather than variation induced by the treatment.” (opens the source at this quote in a new tab)

Resistance sequencing substudy: conclusion

“This study demonstrates evidence of retained susceptibility of HSV-2 to pritelivir in immunocompetent persons following daily therapy for up to 28 days.” (opens the source at this quote in a new tab)

Modeling study: dose response of shedding in the trial data

“There was a pronounced dose response of quantitative shedding rate and quantity to pritelivir. Quantitative shedding frequency on 5 mg approximated that of placebo, while there was a sequential decrease in shedding from 5 mg daily to 25 mg daily, from 25 mg daily to 400 mg weekly, and from 400 mg weekly to 75 mg daily” (opens the source at this quote in a new tab)

Modeling study: simulated and observed shedding rates by arm

“Mean simulated shedding rate was 14.7% (range: 11.2–19.2%) versus 16.6% in the trial for placebo; 13.5% (range: 9.4–23.8%) versus 18.2% in the trial for 5 mg; 7.9% (range: 3.3 – 11.1%) versus 9.3% in the trial for 25 mg; 2.5% (range: 0.7–6.2%) versus 2.1% in the trial for 75 mg; and 4.6% (range: 1.8–10.5%) versus 5.3% in the trial for 400 mg weekly.” (opens the source at this quote in a new tab)

Modeling study: estimated in vivo EC50 and inhibition of replication by dose

“In vivo EC50 values between 50 and 70 ng/mL provided best fit” (opens the source at this quote in a new tab)
“Thus, our in vivo EC50 estimate is 2–15 fold higher than in vitro measures.” (opens the source at this quote in a new tab)
“At 75 mg, inhibition of replication was sustained but incomplete (80%). At 25 mg, inhibition was ~30%.” (opens the source at this quote in a new tab)

Modeling study: predicted reduction in shedding at 150 mg daily

“Up to 96% reduction versus the placebo shedding rate was reached at 150 mg per day, and average daily cumulative dose regardless of dosing frequency was highly predictive of simulated and observed shedding rate” (opens the source at this quote in a new tab)

Conclusion of the publication

“Pritelivir reduced the rates of genital HSV shedding and days with lesions in a dose-dependent manner in otherwise healthy men and women with genital herpes.” (opens the source at this quote in a new tab)

Overall pattern of adverse events and the serious adverse event reported in the publication

“No pattern of adverse events or laboratory abnormalities emerged during the study” (opens the source at this quote in a new tab)
“One serious adverse event occurred in a participant receiving 25 mg of pritelivir daily, who was hospitalized for pancreatitis 2 weeks after completing treatment; a review of the participant's medical history revealed prior alcohol-related pancreatitis and probable continued alcohol intake.” (opens the source at this quote in a new tab)

Nausea

“The number of days with nausea was highest in the group receiving 400 mg of pritelivir per week (33 days of nausea) and the group receiving 5 mg per day (44 days), although a single participant accounted for 75% of the days of nausea in the group receiving 400 mg per week.” (opens the source at this quote in a new tab)
“Reports of nausea did not appear to be related to the dose of pritelivir.” (opens the source at this quote in a new tab)

Discontinuations because of adverse events

“Three participants receiving pritelivir discontinued treatment because of adverse events: one participant receiving 5 mg daily discontinued treatment because of moderate headache, nausea, chest pain, and dyspnea; one receiving 25 mg daily discontinued treatment because of suspected systemic lupus erythematosus; and one receiving 400 mg per week discontinued treatment because of mild anxiety.” (opens the source at this quote in a new tab)

Safety conclusions of the investigators

“The rate of adverse events was similar in all groups.” (opens the source at this quote in a new tab)
“In our small study, pritelivir, when administered daily for 4 weeks, was not associated with any clinical, laboratory, or cardiac abnormalities that emerged during treatment.” (opens the source at this quote in a new tab)

All-cause mortality, serious adverse events, and other adverse events by arm in the registry results

Category	Dose regimen 1 (5 mg daily)	Dose regimen 2 (25 mg daily)	Dose regimen 3 (75 mg daily)	Dose regimen 4 (400 mg weekly)	Placebo
All-cause mortality	“0/33 (0.00%)”	“0/32 (0.00%)”	“0/29 (0.00%)”	“0/31 (0.00%)”	“0/30 (0.00%)”
Serious adverse events	“0/33 (0.00%)”	“0/32 (0.00%)”	“0/29 (0.00%)”	“0/31 (0.00%)”	“0/30 (0.00%)”
Other (not including serious) adverse events	“27/33 (81.82%)”	“26/32 (81.25%)”	“27/29 (93.10%)”	“23/31 (74.19%)”	“23/30 (76.67%)”

Adverse events by system organ class and term in the registry results, participants affected of participants at risk

System organ class	Term	Dose regimen 1 (5 mg daily)	Dose regimen 2 (25 mg daily)	Dose regimen 3 (75 mg daily)	Dose regimen 4 (400 mg weekly)	Placebo
Blood and lymphatic system disorders	Blood and Lymphatic System Disorders	“2/33 (6.06%)”	“1/32 (3.13%)”	“3/29 (10.34%)”	“1/31 (3.23%)”	“1/30 (3.33%)”
Gastrointestinal disorders	Gastrointestinal Disorder	“17/33 (51.52%)”	“13/32 (40.63%)”	“17/29 (58.62%)”	“15/31 (48.39%)”	“12/30 (40.00%)”
Gastrointestinal disorders	Nausea	“9/33 (27.27%)”	“5/32 (15.63%)”	“5/29 (17.24%)”	“9/31 (29.03%)”	“4/30 (13.33%)”
Gastrointestinal disorders	Diarrhoea	“3/33 (9.09%)”	“3/32 (9.38%)”	“5/29 (17.24%)”	“4/31 (12.90%)”	“3/30 (10.00%)”
Gastrointestinal disorders	Abdominal Pain Upper	“2/33 (6.06%)”	“4/32 (12.50%)”	“4/29 (13.79%)”	“2/31 (6.45%)”	“3/30 (10.00%)”
Gastrointestinal disorders	Dyspepsia	“0/33 (0.00%)”	“1/32 (3.13%)”	“2/29 (6.90%)”	“4/31 (12.90%)”	“2/30 (6.67%)”
Gastrointestinal disorders	Abdominal Pain	“0/33 (0.00%)”	“4/32 (12.50%)”	“0/29 (0.00%)”	“0/31 (0.00%)”	“0/30 (0.00%)”
General disorders	General Disorders and Administration Site Conditions	“7/33 (21.21%)”	“6/32 (18.75%)”	“8/29 (27.59%)”	“3/31 (9.68%)”	“8/30 (26.67%)”
General disorders	Fatigue	“4/33 (12.12%)”	“2/32 (6.25%)”	“5/29 (17.24%)”	“3/31 (9.68%)”	“5/30 (16.67%)”
General disorders	Pyrexia	“1/33 (3.03%)”	“1/32 (3.13%)”	“3/29 (10.34%)”	“1/31 (3.23%)”	“0/30 (0.00%)”
Infections and infestations	Infections and Infestations	“12/33 (36.36%)”	“8/32 (25.00%)”	“8/29 (27.59%)”	“8/31 (25.81%)”	“10/30 (33.33%)”
Infections and infestations	Upper Respiratory Tract Infection	“3/33 (9.09%)”	“2/32 (6.25%)”	“1/29 (3.45%)”	“3/31 (9.68%)”	“1/30 (3.33%)”
Infections and infestations	Urinary Tract Infection	“2/33 (6.06%)”	“1/32 (3.13%)”	“1/29 (3.45%)”	“0/31 (0.00%)”	“3/30 (10.00%)”
Injury, poisoning and procedural complications	Injury, Poisoning and Procedural Complications	“1/33 (3.03%)”	“1/32 (3.13%)”	“5/29 (17.24%)”	“2/31 (6.45%)”	“0/30 (0.00%)”
Investigations	Investigations	“1/33 (3.03%)”	“1/32 (3.13%)”	“4/29 (13.79%)”	“2/31 (6.45%)”	“5/30 (16.67%)”
Musculoskeletal and connective tissue disorders	Musculoskeletal and Connective Tissue Disorders	“10/33 (30.30%)”	“9/32 (28.13%)”	“7/29 (24.14%)”	“6/31 (19.35%)”	“11/30 (36.67%)”
Musculoskeletal and connective tissue disorders	Arthralgia	“1/33 (3.03%)”	“5/32 (15.63%)”	“2/29 (6.90%)”	“1/31 (3.23%)”	“2/30 (6.67%)”
Musculoskeletal and connective tissue disorders	Back Pain	“1/33 (3.03%)”	“2/32 (6.25%)”	“2/29 (6.90%)”	“1/31 (3.23%)”	“5/30 (16.67%)”
Musculoskeletal and connective tissue disorders	Pain in the Extremity	“2/33 (6.06%)”	“2/32 (6.25%)”	“2/29 (6.90%)”	“2/31 (6.45%)”	“3/30 (10.00%)”
Nervous system disorders	Nervous System Disorders	“13/33 (39.39%)”	“15/32 (46.88%)”	“14/29 (48.28%)”	“16/31 (51.61%)”	“12/30 (40.00%)”
Nervous system disorders	Headache	“12/33 (36.36%)”	“14/32 (43.75%)”	“11/29 (37.93%)”	“14/31 (45.16%)”	“12/30 (40.00%)”
Nervous system disorders	Dizziness	“3/33 (9.09%)”	“1/32 (3.13%)”	“5/29 (17.24%)”	“2/31 (6.45%)”	“1/30 (3.33%)”
Psychiatric disorders	Psychiatric Disorders	“3/33 (9.09%)”	“4/32 (12.50%)”	“5/29 (17.24%)”	“4/31 (12.90%)”	“3/30 (10.00%)”
Psychiatric disorders	Insomnia	“2/33 (6.06%)”	“3/32 (9.38%)”	“3/29 (10.34%)”	“2/31 (6.45%)”	“2/30 (6.67%)”
Reproductive system and breast disorders	Reproductive System and Breast Disorders	“7/33 (21.21%)”	“4/32 (12.50%)”	“7/29 (24.14%)”	“5/31 (16.13%)”	“8/30 (26.67%)”
Reproductive system and breast disorders	Dysmenorrhoea	“2/33 (6.06%)”	“0/32 (0.00%)”	“2/29 (6.90%)”	“2/31 (6.45%)”	“4/30 (13.33%)”
Reproductive system and breast disorders	Pruritus Genital	“1/33 (3.03%)”	“0/32 (0.00%)”	“2/29 (6.90%)”	“1/31 (3.23%)”	“3/30 (10.00%)”
Respiratory, thoracic and mediastinal disorders	Respiratory, Thoracic and Mediastinal Disorders	“4/33 (12.12%)”	“4/32 (12.50%)”	“6/29 (20.69%)”	“7/31 (22.58%)”	“6/30 (20.00%)”
Respiratory, thoracic and mediastinal disorders	Oropharyngeal Pain	“1/33 (3.03%)”	“2/32 (6.25%)”	“4/29 (13.79%)”	“4/31 (12.90%)”	“3/30 (10.00%)”
Skin and subcutaneous tissue disorders	Skin and Subcutaneous Tissue Disorders	“2/33 (6.06%)”	“6/32 (18.75%)”	“5/29 (17.24%)”	“7/31 (22.58%)”	“3/30 (10.00%)”

Method of adverse event collection in the registry results

“Indicates events were collected by systematic assessment” (opens the source at this quote in a new tab)

FDA clinical hold after the trial and the nonclinical findings behind it

“In May 2013, the clinical development of pritelivir was placed on hold by the Food and Drug Administration because of unexplained dermal and hematologic findings in a toxicology study of monkeys treated with daily doses ranging from 75 mg per kilogram of body weight to 1000 mg per kilogram (these doses were 70 to more than 900 times as high as a dose of 75 mg in humans). The reason for the findings in monkeys is currently under investigation; such findings were not observed in the current trial.” (opens the source at this quote in a new tab)

Summary

AIC316-01-II-01 enrolled otherwise healthy, immunocompetent adults with genital HSV-2 infection, a population outside the labeled indication of immunocompromised adults with refractory mucocutaneous HSV infection. The regimens tested (5, 25, or 75 mg daily, or 400 mg weekly) differ from the approved regimen of 400 mg on day 1 followed by 100 mg daily. Treatment lasted 28 days, and 155 participants were analyzed in five arms of 29 to 33 participants. The primary endpoint was a virologic measure, and the investigators state that the trial was not powered to evaluate recurrences. The publication reports one serious adverse event (pancreatitis 2 weeks after treatment), and the registry results list none.

Limitations stated by the investigators

“Although our study was not powered to evaluate the effect of pritelivir on recurrences of symptomatic genital herpes, we did observe significant reductions in the number of days of genital lesions at doses of 75 mg daily and 400 mg weekly that paralleled the trend toward fewer recurrences at these doses.” (opens the source at this quote in a new tab)
“The question of whether further increases in the daily dose of pritelivir would completely abrogate viral shedding will have to be addressed in additional studies.” (opens the source at this quote in a new tab)
“Although it is appealing to compare the efficacy of pritelivir with that of currently licensed products, the frequency of viral shedding and the magnitude of the reduction in the quantity of virus shed with antiviral medication depend heavily on the characteristics of the participants enrolled; thus, a direct comparison in a clinical trial is warranted to properly determine comparative efficacy.” (opens the source at this quote in a new tab)
“Further studies will be needed to assess the magnitude of clinical benefit in the treatment of HSV infections and to establish the usefulness of pritelivir in treating severe HSV disease and in reducing sexual transmission.” (opens the source at this quote in a new tab)

Limitations of the resistance sequencing substudy

“While these analyses involved a relatively small number of patients, at least 2 dosages in our study were less than optimal for HSV replication inhibition, as shown by persistent viral shedding during dosing” (opens the source at this quote in a new tab)
“In human studies, the longest duration of pritelivir administration so far has been 28 days. Thus, whether prolonged use will result in selection for resistance is as yet unclear. Moreover, studies in immunocompromised patients with poor cellular immunity are lacking.” (opens the source at this quote in a new tab)
“Whether resistance will be eventually observed with HPI therapy is unknown.” (opens the source at this quote in a new tab)

Limitations of the modeling study

“A few potential limitations are worth noting. Participants in the placebo arm and four treatment arms of the actual study were not matched. Some difference between arms may be due to different baseline HSV-2 shedding rates, rather than differential drug effects.” (opens the source at this quote in a new tab)
“While our model is effective for predicting outcomes of a clinical trial, it is unable to predict outcomes in treated individuals” (opens the source at this quote in a new tab)
“Finally, the model generally overestimates the proportion of short, one-day episodes, particularly in low dose simulations.” (opens the source at this quote in a new tab)

Agreement on publication recorded in the registry

“There IS an agreement between Principal Investigators and the Sponsor (or its agents) that restricts the PI's rights to discuss or publish trial results after the trial is completed” (opens the source at this quote in a new tab)