A Randomised, Double-blind, Placebo-controlled Phase IIb Trial to Test FLU-v Vaccine
A Randomised, Double-blind, Placebo-controlled Phase IIb Trial to Test FLU-v Vaccine
FLU-v is a vaccine that aims to protect against a wide range of flu viruses. The purpose of
 this study is to measure the immune responses induced by FLU-v vaccine. This study will look
 at how safe FLU-v is when administered and how successful it is in preventing flu or reducing
 the severity of the flu symptoms.

 The study requires 222 healthy volunteers 18-60 years old. Participation in the study will
 take a maximum of 7 months and consists of 5 visits. During visit 1, subjects will be
 examined by a doctor to make sure they are eligible to enter the study. A 15ml blood sample
 (a tablespoon) will be taken to check general health followed by a general physical exam.
 Medical history and some personal information will be collected. Subjects that have received
 the traditional flu vaccine in the past 6 months, and those females who are pregnant or
 breastfeeding will not be allowed in the study. Subjects of childbearing age must agree to
 use effective contraceptive methods.

 At visit 2, subjects will be randomly allocated to one of the four treatment groups
 summarised below:

 -  Treatment 1: FLU-v (test vaccine) at the start of the study (Day 0) and then again 21
 days later

 -  Treatment 2: FLU-v (test vaccine) with an additional substance added [known as Montanide
 ISA 51] which improves the effect of the test vaccine. Injection will be given on Day 0
 and then Placebo (no test vaccine) alone 21 days later

 -  Treatment 3: Placebo (no test vaccine) injection on Day 0 and then 21 days later

 -  Treatment 4: Placebo (no test vaccine) with an additional substance added [known as
 Montanide ISA 51] on Day 0 and then Placebo (no test vaccine) alone 21 days later
 Treatment will be injected under the skin in the upper arm on day 0 (visit 2) and 21
 days later (visit 3). Blood samples will be taken before treatment (day 0), and on days
 42 (visit 4) and 180 (visit 5) to the immune responses induced by the vaccine.

 Subjects will be asked to complete a diary card to write down any side effects that they may
 experience after vaccination. Subjects will also be asked to complete another diary card to
 document any flu-like symptoms experienced between December 2016 and March 2017, this time is
 officially considered as the flu season. During this period, if the subject experiences
 flu-like symptoms, a collection of a nose and tonsil swab will be arranged by the study site
 to confirm whether they have the flu or not.
Rationale: Current seasonal influenza vaccines mainly induce immune responses against viral
 membrane glycoproteins. These proteins, however, undergo continuous mutations by a process
 called antigenic drift. To prevent immune escape, annual vaccination with the latest
 predicted viral strains is adopted. Such vaccination strategy not only poses inconvenience
 and cost-inefficiency, but also results in poor protective effectiveness when the vaccinated
 strains are mismatched with the actual circulating strains. The latter point is especially of
 concern during a pandemic outbreak, when a large geographical area is affected and the
 general population is naïve to the newly re-assorted viral strain due to antigenic shift.

 Objective: To evaluate the safety and immunogenicity of the influenza vaccine (FLU-v, as a
 suspension or adjuvanted as emulsion) targeting conserved immunogenic regions of influenza A
 and B viruses in healthy adults, in particular to show that the TH1 cytokine response at 42
 and 180 days after the first injection is greater in the adjuvanted FLU-v and unadjuvanted
 FLU-v than in the placebo.

 Study design: A total of 222 study participants will be recruited. The study follows a
 factorial design where the two factors are treatment (FLU-v / placebo) and formulation
 (unadjuvanted / suspension, adjuvanted / emulsion). Subjects will be randomised in two strata
 (age 18 to 40, age 41 to 60) to one of the following treatment regimens:

 -  Group 1 (n=74): FLU-v (unadjuvanted) as a suspension in pH neutral HCl/NaOH (0.5mL) on
 Day 0 and Day 21

 -  Group 2 (n=74): (0.5mL) ISA-51-adjuvanted FLU-v emulsified in water for injection (WFI)
 on Day 0, saline (0.5mL) on Day 21

 -  Group 3 (n=37): saline solution (0.5mL ) on Day 0 and Day 21

 -  Group 4 (n=37): WFI and ISA-51 emulsion (0.5mL) on Day 0, saline (0.5mL) on Day 21

 Each administration will be given subcutaneously. Solicited and unsolicited adverse events
 (AEs) will be collected by AE questionnaire/diary card until day 42. Adverse events (AEs) and
 serious adverse events (SAEs) will be collected for the entire study period. The treatments
 will be administered starting in third quarter of 2016 in order to provide protection for the
 subsequent influenza season starting in December 2016. Blood samples will be taken from all
 subjects on day 0 (before FLU-v vaccination), 42 (21 days after the second dosing) and 180
 (159 days after the second dosing) for the evaluation of FLU-v-specific cellular and humoral
 immune responses. Clinical symptom scores to ascertain severity and the incidence of
 RT-PCR-confirmed influenza A and/or B infection will be recorded during the subsequent
 influenza season (December 2016 to March 2017) to decide clinical efficacy of the tested
 vaccines.

 Study population: Healthy volunteers aged 18-60 years. Intervention: FLU-v investigational
 influenza vaccine lyophilised product containing 500 micrograms of total peptides
 reconstituted in either 0.01M HCl (0.25ml) and 0.01M NaOH (0.25ml) to achieve a volume of
 0.5ml, or emulsified in WFI (water for injection, 0.25ml) and ISA-51 (0.25ml) to achieve a
 volume of 0.5ml.

 Primary study parameters/endpoints: For immunogenicity: To compare the change from baseline
 (Day 0) between treatments in cellular immune responses, specifically TH1 cytokines, in all
 groups 42 and 180 days following FLU-v vaccination. For safety: (1) To evaluate the solicited
 AEs in all subjects until 21 days after the last dosing of the study vaccine (FLU-v); (2) To
 evaluate the unsolicited AEs and SAEs in all subjects for the entire study period after the
 first dosing of FLU-v.

 Secondary study parameters/endpoints: To evaluate the humoral immune responses specific to
 FLU-v and TH2 cytokines from baseline in all groups 42 and 180 days following FLU-v
 vaccination.

 Exploratory study parameters/endpoints: For immunogenicity: To evaluate the change from
 baseline in cellular immune responses based on additional CMI assays such as ELISPOT
 (Enzyme-Linked ImmunoSpot) in all groups at 42 and 180 days following FLU-v vaccination, in a
 subset of subjects chosen at random.

 Clinical efficacy: (1) To evaluate the efficacy of FLU-v vaccine in reducing the incidence of
 RT-PCR confirmed influenza A and/or B infections in all subjects during the influenza season
 2016-2017. (2) To evaluate the efficacy of FLU-v vaccine in the reduction of symptom score
 among RT-PCR confirmed influenza A and/or B infection cases during the influenza season
 2016-2017. The relationship between efficacy and cellular and humoral response will be
 explored if possible.

 The effect of previous influenza vaccination on the immunogenicity of FLU-v will be assessed
 in a post-hoc exploratory analysis after stratification of the data based on exposure to the
 influenza vaccine in the previous 24 months or over.

 Nature and extent of the burden and risks associated with participation, benefit and group
 relatedness:

 The intended application of the IMP is as a prophylactic vaccine to prevent influenza virus
 infection by inducing an enhanced influenza-specific immune response. The FLU-v vaccine is
 designed to be delivered either as a naturally particulate suspension (i.e. no adjuvant) or
 emulsified in adjuvant. Particulate and emulsified protein preparations are preferentially
 taken up by phagocytic cells (e.g. dendritic cells) responsible for inducing primary immune
 responses.

 Treatment with FLU-v of up to four doses of 263 μg/occasion or up to two doses of 553
 μg/occasion, with or without adjuvant, was well tolerated in animals with no signs of
 systemic toxicity. In pre-clinical studies, subcutaneous administration of ISA 51 resulted in
 no systemic toxicity. At the injection sites and occasionally in adjacent tissue the injected
 material remained surrounded by a mild chronic inflammatory response. As the IMP is provided
 in one presentation (500 μg of the combined peptide) in lyophilized form for suspension in a
 sealed single-use vial, overdose is highly unlikely.

 In a single centre, randomised, double blind phase I study of the safety, tolerability and
 immunogenicity of FLU-v no safety or tolerability concerns were identified at the doses
 administered (250 μg and 500 μg FLU-v) to the subjects in this clinical study and no safety
 or tolerability concerns were identified following administration of the adjuvant to the
 subjects (1). FLU-v vaccine candidate was demonstrated to be immunogenic in humans, as
 measured by ex vivo γ-interferon production (1).

 Clinical experience with Montanide ISA 51 dates back to the 1990s and most trials were
 related to cancer and Acquired Immune Deficiency Syndrome (AIDS). Currently, cancer trials in
 melanoma, colorectal, prostate, cervical, brain cancer and leukemia are ongoing. Frequency of
 vaccination is often between 2 and 4 weeks, and the number of injections can reach up to 40.
 Route of immunization is most often subcutaneous and volume of injection can reach up to 3mL.
 Most common local reactions are local pain, tenderness, erythema and granuloma at the
 injection site. Less frequently, mild to moderate transient indurations and swelling are
 described. General reactions are mainly 'flu-like symptoms such as chills, fever and
 headaches. Lethargy and nausea are also observed. The intensity is usually mild or moderate.
 No biological changes are generally observed. As the Placebo vaccine is provided in a sealed
 single-use vial, overdose is highly unlikely (Influenza virus (FLU-v) vaccine Investigator's
 Brochure, Edition 2.0, 07 September 2015).
Inclusion Criteria:

 -  Healthy males or healthy non-pregnant females (as indicated by a negative blood
 pregnancy test done during the screening visit) between the ages of 18 and 60 years,
 inclusive;

 -  Women of childbearing potential (not surgically sterile or postmenopausal for greater
 than or equal to one year) and men must agree to practice appropriate contraception (a
 combination of barrier and hormonal methods for women and a condom for men) from
 screening and until at least 30 days (up to Study Day 51 for females) and 90 days (up
 to Study Day 111 for males), after the last vaccination.

 -  A subject is in good health, as determined by a comprehensive clinical assessment
 {vital signs (heart rate, blood pressure, oral temperature)}, blood chemistry test
 (electrolytes, renal/kidney function, liver function, C-reactive protein, complete
 blood count), medical history, general physical examination, self-reported illness}
 and the clinical judgment of the investigator;

 -  Able to understand and comply with planned study procedures;

 -  Provides signed informed consent form

 Exclusion Criteria:

 -  Has a known allergy to any of the components of the vaccine.

 -  Has a history of severe reaction following immunization.

 -  Persons with immune deficiency/disorder, whether due to genetic defect,
 immunodeficiency disease, or immunosuppressive therapy.

 -  Women who have a positive pregnancy test during the screening visit or who are
 breastfeeding.

 -  Has a history of any of the following (reported by subjects):

 -  Acute disseminated encephalomyelitis (ADEM);

 -  Neoplastic disease - current or previous;

 -  Asthma or severe allergic disease;

 -  Bleeding disorders

 -  Chronic Hepatitis B and/or C infection;

 -  Chronic liver disease;

 -  Diabetes mellitus;

 -  Guillain-Barré syndrome;

 -  HIV;

 -  Rheumatoid arthritis or other autoimmune diseases;

 -  Severe renal disease;

 -  Transplant recipients;

 -  Unstable or progressive neurological disorders.

 -  Receipt of medicines/treatments that may affect evaluation of immunogenicity such as:

 -  Oral or parenteral steroids, high-dose inhaled steroids (greater than 800
 micrograms/day of beclomethasone dipropionate or equivalent) or other
 immunosuppressive or cytotoxic drugs (azathioprine (Imuran), cyclosporine
 (Neoral, Sandimmune, SangCya); monoclonal antibodies such as basiliximab
 (Simulect), daclizumab (Zinbryta), infliximab (Remicade), rituximab (MabThera),
 alemtuzumab (Campath and Lemtrada), omalizumab (Xolair), abatacept (Orencia),
 adalimumab (Humira and Exemptia) and etanercept (Enbrel)basiliximab (Simulect),
 daclizumab (Zenapax), and muromonab (Orthoclone OKT3); corticosteroids such as
 prednisone (Deltasone, Orasone); tacrolimus (Prograf, Advagraf, Protopic);
 Glatiramer acetate (Copaxone); Mycopehnolate (Cellcept); Sirolimus (Rapamune);
 (within 6 months of vaccination in this study)

 -  Immunoglobulin or other blood products (plasma, blood cells, coagulation factors,
 haemoglobin)(within 3 months of vaccination in this study);

 -  An experimental agent (vaccine, drug, biologic, device, blood product, or
 medication) within 1 month of vaccination in this study, or expects to receive an
 experimental agent (during the study period).

 -  Influenza antiviral medication (Amantadine (Symmetrel); Rimantadine (Flumadine);
 Zanamivir (Relenza), Oseltamivir (Tamiflu) (within 4 weeks of vaccination in this
 study).

 -  Has received any influenza vaccine within 6 months of vaccination in this study.

 -  Has influenza-like illness (a sudden onset of symptoms and at least one of the four
 systemic symptoms-fever or feverishness, malaise, headache, myalgia and at least one
 of the three respiratory symptoms-cough, sore throat, shortness of breath) or acute
 respiratory infection (a sudden onset of symptoms and at least one of the four
 respiratory symptoms-cough, sore throat, shortness of breath, coryza (Rhinitis) and a
 clinician's judgement that the illness is due to an infection) within 6 months prior
 to vaccination in this study. These symptoms must have stopped the subject from
 carrying out their normal daily activities such as attending work or school for a
 period of at least 3 days.

 -  Has an acute illness, including an oral temperature greater than 38 degrees Celsius,
 within 1 week of vaccination.

 -  Has a history of alcohol or drug abuse within the last 2 years deemed unsuitable for
 inclusion by the investigator.

 -  Any abnormal haematology values and/or serum chemistries judged by the Investigator as
 clinically significant.