Phase II Trial to Evaluate Safety and Immunogenicity of a Dengue 1,2,3,4 (Attenuated) Vaccine

• Safety [ Time Frame: Up to day 21 after vaccination ] [ Designated as safety issue: Yes ]
  Frequency of solicited and unsolicited local and systemic adverse reactions up to Day 21 after vaccination.
• Immunogenicity [ Time Frame: Up to day 56 after vaccination ] [ Designated as safety issue: No ]
  Seroconversion rate for each of the four vaccine viruses defined by PRNT50 ≥1:10 for each vaccine virus serotype on Days 28 or 56.

• Safety [ Time Frame: up to Day 182 after the first vaccination ] [ Designated as safety issue: Yes ]
  1. Unsolicited adverse events after Day 21 and up to Day 182 after the first vaccination;
  2. Viremia for each of the four vaccine virus on Days 3, 6, 9, 12, 15 and 21 after the first vaccination of all volunteers in Step A of study and 40 volunteers with previous exposure to Dengue of Step B. For other volunteers from Step B, viremia only is measured on Day 12.
  3. Cases of suspected and confirmed dengue in five years after the first vaccination;
• Immunogenicity [ Time Frame: Annualy up to five years ] [ Designated as safety issue: No ]
  1. Response rates of monovalent, bivalent, trivalent or tetravalent, at Day 28 and 56 and annually after vaccination, as measured by PRNT50 for each one of the four vaccine viruses;
  2. The difference between the geometric means, pre and post vaccination, serum level of neutralizing antibodies measured by PRNT50 for each of the four vaccine viruses evaluated until five years after the first vaccination.

This is a phase II step-wise, randomized, multicenter, double-blind and controlled clinical trial to evaluate the safety and immunogenicity of a attenuated tetravalent lyophilized dengue vaccine manufactured by Butantan Institute. Three Clinical Sites at University of Sao Paulo - Brazil will participate in the study. A total of 300 volunteers will be recruited and divided into two steps: Step A (with no previous exposure to dengue) and Step B (with and without previous exposure to dengue). In step A the participants will be assigned to receive either the lyophilized vaccine, or the liquid vaccine(developed at NIH and produced and formulated at Butantan according to the NIH-Protocol), or the placebo. In Step B participants will be assigned to receive either the lyophilized vaccine, or the placebo. Both vaccine formulations (lyophilized and liquid) are composed of the same attenuated viruses: rDEN1∆30, rDEN2/4∆30(ME), rDEN3∆30/31, and rDEN4∆30. At the end of the study, 20 volunteers will have received the liquid formulation (NIH), 210 the lyophilized formulation (Butantan), and 70 will have received the placebo. All participants included in both steps will be followed by a period of five years after their inclusion in the study. The study hypothesis is that the investigational lyophilized dengue vaccine manufactured by Butantan Institute is safe and confers balanced immune response, after one dose of 1000PFU, to all for vaccine viruses.

This is a phase II step-wise, randomized, multicenter, double-blind and controlled clinical trial to evaluate the safety and immunogenicity of an attenuated tetravalent lyophilized dengue vaccine manufactured by Butantan Institute. A total of 300 volunteers will be recruited including men and no pregnant/breastfeeding women between 18 and 59 years of age complete, with and without previous exposure to dengue that will be randomized into Step A and Step B. Step A will include 50 volunteers with no previous exposure and they will be randomly assigned to receive one dose of either the lyophilized formulation (Butantan), or the liquid formulation (TetraVax-DV Vaccine - Admixture TV003 developed by NIH/NIAID and produced and formulated at Butantan according the NIH-Protocol), or the placebo. A second dose will be administered six months after the first vaccination as part of an exploratory assessment. Step B will include 250 participants(50 without previous exposure to dengue,and 200 with previous exposure to dengue) who will be randomly assigned to receive one dose of either the lyophilized formulation (Butantan),or the placebo. All participants included in both steps will be followed by a period of five years after their inclusion in the study. The vaccines will be administered at dose of 1000 PFU (for each of the vaccine viruses),and both vaccines and the placebo will be administered subcutaneously.

• Biological: dengue lyophilized vaccine
  Dose 1000 PFU per virus (1,2,3,4) Route:subcutaneous
  Other Name: Dengue 1,2,3,4 (attenuated) Vaccine: Lyophilized formulation
• Biological: dengue liquid vaccine
  Dose 1000 PFU per virus Route: subcutaneous
  Other Name: TetraVax-DV Vaccine - Admixture TV003
• Other: Placebo
  Route:subcutaneous
  Other Name: Placebo

• Experimental: Step A: dengue lyophilized vaccine
  Experimental: Dengue 1,2,3,4 (attenuated) Vaccine: Lyophilized formulation Active comparator:dengue liquid vaccine Placebo comparator
  Intervention: Biological: dengue lyophilized vaccine
• Active Comparator: Step A:dengue liquid vaccine
  Experimental: Dengue 1,2,3,4 (attenuated) Vaccine: Lyophilized formulation Active comparator:dengue liquid vaccine Placebo comparator
  Intervention: Biological: dengue liquid vaccine
• Placebo Comparator: Step A: Placebo
  Experimental: Dengue 1,2,3,4 (attenuated) Vaccine: Lyophilized formulation Active comparator:dengue liquid vaccine Placebo comparator
  Intervention: Other: Placebo
• Experimental: Step B: dengue lyophilized vaccine
  Experimental: Dengue 1,2,3,4 (attenuated) Vaccine: Lyophilized formulation Placebo comparator
  Intervention: Biological: dengue lyophilized vaccine
• Placebo Comparator: Step B: Placebo
  Experimental: Dengue 1,2,3,4 (attenuated) Vaccine: Lyophilized formulation Placebo comparator
  Intervention: Other: Placebo

• Blaney JE Jr, Durbin AP, Murphy BR, Whitehead SS. Development of a live attenuated dengue virus vaccine using reverse genetics. Viral Immunol. 2006 Spring;19(1):10-32.
• Durbin AP, Kirkpatrick BD, Pierce KK, Schmidt AC, Whitehead SS. Development and clinical evaluation of multiple investigational monovalent DENV vaccines to identify components for inclusion in a live attenuated tetravalent DENV vaccine. Vaccine. 2011 Sep 23;29(42):7242-50. Epub 2011 Jul 21.

Inclusion Criteria:

• Healthy adults (men and non-pregnant women), from 18 to 59 years old;
• Willingness to participate throughout the study period (approximately five years);
• Willingness to participate documented by the signature of ICF;
• Females with childbearing potential must be willing to use contraceptive methods during the first study year. All female volunteers will be considered with childbearing potential unless they have documented history of hysterectomy, tubal ligation or are postmenopausal (12 months of amenorrhea after the last menstrual period).

Exclusion Criteria:

• Pregnancy (confirmed by positive beta-hCG test) or breastfeeding;
• Evidence of active neurological, cardiac, pulmonary, hepatic or renal disease as clinical history, physical examination and/or laboratory results;
• Compromised immune system diseases including: diabetes mellitus, cancer (except basal cell carcinoma) and autoimmune diseases;
• Behavioral, cognitive or psychiatric disease that in the opinion of the principal investigator or his representative physician, affects the participant ability to understand and cooperate with all study protocol requirements;
• Values of absolute neutrophil, alanine aminotransferase (ALT) or serum creatinine count greater than or equal to Grade 1, as defined in the protocol;
• Abusive usage of alcohol or drugs in the past 12 months that has caused medical, professional or family problems, indicated by clinical history;
• History of severe allergic reactions or anaphylaxis;
• Diagnosis of asthma with a history of hospitalization in the last six months due to illness;
• Fever or suspect fever within 72 hours prior to vaccination or axillary temperature greater than 37,8°C on the day of vaccination;
• Positive result of HIV-1 serology by screening or confirmed tests;
• Screening or confirmed positive test for hepatitis C virus (HCV);
• Positive test of hepatitis B virus antigen surface (AgHBs) alone or against hepatitis B "core" antigen antibody (anti-HBc);
• Use of corticosteroids (except topical or nasal) or other immunosuppressive drugs within 42 days before study initiation/baseline. It will be considered immunosuppressive dose of corticosteroids the equivalent to a dose ≥10 mg of prednisone per day for over 14 days;
• Use of anticoagulant medication;
• Have received live virus vaccine within 28 days or killed virus vaccine in the last 14 days prior to vaccination, or have a scheduled immunization during the first 42 days after receiving the investigational product;
• History of asplenia;
• Have received blood products in the past six months, including transfusions or immunoglobulin, or scheduled administration of blood products or immunoglobulin for the first 42 days after vaccination;
• Use of any investigational product within 42 days before or after receiving this study vaccination;
• Has participated in another clinical trial six months prior to vaccination;
• Denies permission for biological material storage for future research as defined in ICF;
• Any other condition that might put in risk the safety/rights of a potential participant or hurdle his/her compliance with this protocol in investigator's opinion or his representative physician.

• Banco Nacional de Desenvolvimento Economico e Social
• Fundação de Amparo a Pesquisa do Estado de Sao Paulo
• Butantan Foundation