Paz Bailey G 1, Sternberg M 1, Puren AJ 2, Markowitz LE 1, Ballard R 1, Delany S 3, Hawkes S 4, Nwanyanwu O 1, Ryan C 1, and Lewis DA 5 1. NCHHSTP, CDC US; 2. Specialized Molecular Diagnostic Unit, NICD, South Africa; 3. Reproductive Health Research Unit, University of the Witwatersrand, South Africa; 4. London School of Hygiene and Tropical Medicine; 5. STI Reference Centre, NICD Addition of anti-herpes treatment to syndromic management of GUD

Rationale for study HSV2 leading cause of genital ulcer disease Data available on episodic treatment among HIV negative patients show that: –Treatment is very effective for initial genital herpes –Treatment has to be started early for recurrent herpes –Reduces duration and shedding 1-2 days No placebo controlled trials on episodic therapy among HIV+ Current WHO GUD management guidelines recommend acyclovir if HSV2>30%

Objectives of the trial Primary Efficacy of acyclovir episodic therapy on reducing the duration of an HSV2 clinical outbreak Secondary Efficacy of acyclovir on reducing HIV RNA in genital ulcers and plasma among HIV-positive with a herpetic ulcer

Methods Study design: double blind randomized placebo controlled trial of acyclovir 400mgs TID for 5 days versus placebo Sample size 600 men with GUD All patients received syndromic management At enrolment tested for HSV2, HIV, syphilis, GUD etiology and other STI in urine by PCR Questionnaire for sexual behaviour and demographics HIV+ tested for CD4, HIV RNA in plasma and ulcers, and HSV2 in ulcers Follow-up visits on days 2 or 4, 7, 14 and 28 to measure ulcer healing, HIV and HSV2 shedding in ulcers HIV shedding measured in ulcer lavages with HIV-1 ultrasensitive Amplicor monitor test version 1.5 (Roche) lower threshold 50 copies per ml

Data Analysis Intent to treat analysis Study groups –All participants –HIV-positive with HSV2 ulcer –HIV-negative with HSV2 ulcer –Initial herpes (HSV-2 seronegative and HSV-2 in the ulcer) Outcomes –Ulcer Healing Per visit analysis using modified Poisson regression to obtain relative risk (RR) Time to healing using all visits with survival analysis –HIV RNA in ulcer and plasma Detection (yes/no) using modified Poisson regression Quantitative using maximum likelihood estimation (MLE) due to the censoring –HSV2 DNA in ulcers Detection (yes/no) using modified Poisson regression Quantitative using MLE Primary endpoint: visit day 7 Secondary endpoints: other visits, all visits (survival analysis)

Study flowchart Ineligible = 924 Ulcer healed=273 Does not want HIV test=204 Unable to attend visits=180 Blisters only=163 Ulcer duration >30days=40 Ulcer area >500mm=10 Enrolled before=33 Age 60 years=9 Assessed for eligibility N=1559 Refused 20 Lost to f-up - 28 (9%) HIV+/HSV2 ulcer 18 (12.3%) Attended day (89%) HIV+/HSV2 ulcer – 126 (86%) Attended day (66%) HIV+/HSV2 ulcer – 111 (76%) Attended day (71%) HIV+/HSV2 ulcer – 98 (67%) Acyclovir -309 HIV+/HSV2 ulcer Enrolled HIV+/HSV2 ulcer 295 Eligible 635 Lost to f-up - 28 (9%) HIV+/HSV2 ulcer – 11 (7%) Attended day (89%) HIV+/HSV2 ulcer – 136 (91%) Attended day (71%) HIV+/HSV2 ulcer – 126 (85%) Attended day (71%) HIV+/HSV2 ulcer – 105 (71%) Placebo – 306 HIV+/HSV2 ulcer-149 Note: Lost to follow-up defined as missing visits on day 7, 14 and 28.

Results-Baseline Information All ParticipantsHIV+ HSV2 ulcer Characteristics Acyclovir n=309 (%) Placebo n=306 (%) Acyclovir n=146 (%) Placebo n=149 (%) Median Age (IQR)29 (25-35) 30 (25-35) 30 (27-36)32 (27-36) Condom use RP (always) Sex during last week On antiretrovirals (yes) Prior sores (yes) Median duration of ulcer (IQR)5 (3-7)6 (4-10)5 (4–8)5 (3-10) Median No. ulcers (IQR)2 (1-4)2 (1-3)2 (1-4)2 (1-3) No significant differences between groups, median duration of ulcers p=0.08

HIV and STI Prevalence- Baseline All Participants*HIV+ HSV2 ulcer* Test Acyclovir n=309 (%) Placebo n=306 (%) Acyclovir n=146 (%) Placebo n=149 (%) HIV HSV2 serology Syphilis serology (RPR) Ulcer HSV Any STI in Urine HIV-positives Median base CD4 (IQR)281 ( ) 283 ( ) 287 ( ) 249 ( ) Mean base plasma HIV VL (log 10 copies/ml) (95% CI) 4.78 ( ) 4.81 ( ) 4.85 ( ) 4.84 ( ) % detectable ulcer HIV *No significant differences between groups

Ulcer healing proportion with a healed ulcer by day 7 Group Acyclovir No. % Placebo No. (%) Adjusted* RRP All participants ( )<0.001 HIV +, HSV2+ ulcer ( )0.001 HIV -, HSV2+ ulcer ( )0.17 Initial episodes ( )0.02 CD4 <= 200 cells/ml ( )0.29 CD4> 200 cells/ml ( )0.003 *Adjusted for study site using Modified Poisson Regression, interaction of treatment and CD4 count not significant p=0.45.

Ulcer healing median duration of days Group Acyclovir Median 95% (CI) Placebo Median 95% (CI) Adjusted* Hazard Ratio (95% CI)P All participants6 (5– 6)7 (6–8) 1.3 (1.1 – 1.5)0.01 HIV +, HSV2+ ulcer6 (5 – 6)9 (7–10) 1.5 (1.2 – 2.0) HIV -, HSV2+ ulcer5 (5-6)6 (5-7)1.1 ( )0.47 Initial episodes6 (5-6)8 (6-10)1.6 ( )0.03 CD4 <= 200 cells/ml6 (5-9)9 ( ( )0.36 CD4> 200 cells/ml6 (5–7)9 (6–10)1.8 (1.3 – 2.5)0.001 *Adjusted for Site using Modified Poisson Regression **Adjusted for Site and baseline self-reported duration of ulcer using Cox Proportional Hazards model Interaction term of treatment arm and CD4 count is not significant p=0.34

KM Curve for ulcer healing among HIV+ with HSV-2+ ulcer

Detectable ulcer HIV RNA and HSV2 DNA at day 7 and 14 * Adjusted for study site and baseline HIV RNA shedding from genital ulcers Group Acyclovir n (%) Placebo n (%) Adjusted* RR (95% CI) P HIV+, HSV2 Ulcer: HIV-1 Shedding Baseline ( )0.78 Day (0.4 – 1.0)0.05 Day ( )0.01 HIV +, HSV2 Ulcer: HSV2 Shedding Baseline ( )0.63 Day (0.1 – 0.4)<0.001 Day (0.3 – 1.1)0.10

HIV and HSV-2 viral loads at day 7 Group Acyclovir Mean* (95% CI) Placebo Mean* (95% CI) Adjusted** difference (95% CI) P Ulcer HIV-1 RNA Baseline1.38 ( )1.42 ( )-0.05 (-0.5 – 0.39)0.82 Day ( )1.17 ( )-0.81 ( )0.01 Plasma HIV-1 RNA Baseline4.85 ( )4.84 ( )0.0 (-0.18 – 0.18)0.96 Day ( )4.85 ( )-0.12 ( )0.04 Ulcer HSV-2 Baseline5.50 ( )5.62 ( )-0.07 (-0.57 – 0.44)0.80 Day ( )3.28 ( )-4.38 ( )<0.001 *Based on using maximum likelihood estimation assuming a normal distribution for the log transformed ulcer HIV-1 shedding. Non-detects are treated as left censored at log10(50). *All outcomes are adjusted for site and in addition outcomes based on follow-up visits are adjusted for baseline HIV shedding

Summary Acyclovir episodic treatment improved ulcer healing among: – All participants –HIV+ with an HSV2 ulcer –Initial episodes of herpes No effect among HIV negative participants Treatment reduced HIV-1 shedding from ulcers and there was some reduction on HIV plasma viral load at day 7 Treatment reduced HSV2 lesional shedding

Recommendations Acyclovir treatment should be recommended as part of syndromic management of genital ulcer disease among HIV-positives HIV negatives? Previous evidence shows modest clinical benefit if therapy is started early The role of self-initiated anti-herpes therapy should be explored HIV-testing should be strongly enforced as part of STD management (little HIV testing is currently happening) GUD management point of entry for ARV treatment and HIV prevention

Collaborators National Institute of Communicable Diseases in South Africa US Centers for Disease Control and Prevention London School of Hygiene and Tropical Medicine Reproductive Health and HIV Research Unit Special thanks to participating clinics and study participants

Ulcer etiology at baseline 133 men with initial HSV-2