<clinical_study>
  <!-- This xml conforms to an XML Schema at:
    https://clinicaltrials.gov/ct2/html/images/info/public.xsd -->
  <required_header>
    <download_date>ClinicalTrials.gov processed this data on September 20, 2023</download_date>
    <link_text>Link to the current ClinicalTrials.gov record.</link_text>
    <url>https://clinicaltrials.gov/ct2/show/NCT02259608</url>
  </required_header>
  <id_info>
    <org_study_id>NL45427.091.13</org_study_id>
    <secondary_id>2013/319</secondary_id>
    <nct_id>NCT02259608</nct_id>
  </id_info>
  <brief_title>γ-irradiated BCG to Train Innate Immunity</brief_title>
  <official_title>Training of Innate Immunity by Vaccination With γ-irradiated BCG</official_title>
  <sponsors>
    <lead_sponsor>
      <agency>Radboud University Medical Center</agency>
      <agency_class>Other</agency_class>
    </lead_sponsor>
  </sponsors>
  <source>Radboud University Medical Center</source>
  <oversight_info>
    <has_dmc>No</has_dmc>
  </oversight_info>
  <brief_summary>
    <textblock>
      Rationale: The live attenuated Bacillus Calmette-Guerin (BCG) vaccine protects against&#xD;
      extrapulmonary infection with Mycobacterium tuberculosis and leprosy. It has been shown that&#xD;
      vaccination with BCG also leads to nonspecific protective effects, e.g. reduced infant&#xD;
      mortality as a result of less severe infections, stimulation of the immune system in patients&#xD;
      with bladder cancer and higher cytokine production upon restimulation of macrophages with&#xD;
      non-related infectious pathogens in vitro. However, because the live attenuated BCG vaccine&#xD;
      cannot be used in immune compromised hosts, the investigators would like to determine whether&#xD;
      similar protective non-specific effects can be induced by γ-irradiated BCG.&#xD;
&#xD;
      Objective: To determine whether vaccination with γ-irradiated BCG results in a higher&#xD;
      cytokine response by monocytes upon restimulation in vitro with infectious pathogens,&#xD;
      compared to monocytes before the vaccination.&#xD;
&#xD;
      Study design: Explorative intervention trial. Study population: Healthy volunteers, 18 - 55&#xD;
      years old. Intervention: Healthy volunteers will be vaccinated with γ-irradiated BCG vaccine.&#xD;
&#xD;
      Main study parameters/endpoints: Blood will be drawn before and at two different time points&#xD;
      after vaccination with BCG to perform restimulation of isolated cells in vitro and compare&#xD;
      cytokine production.&#xD;
&#xD;
      Nature and extent of the burden and risks associated with participation, benefit and group&#xD;
      relatedness: There is no known direct benefit for the volunteers to participate in the trial.&#xD;
      The risks are negligible. BCG vaccination can cause pain and scarring at the site of&#xD;
      injection, just as fever and headache. Local hematoma formation can occur at the site of the&#xD;
      blood drawing. This will be minimized by the blood collection by experienced persons.&#xD;
    </textblock>
  </brief_summary>
  <detailed_description>
    <textblock>
      1. INTRODUCTION AND RATIONALE&#xD;
&#xD;
           The live attenuated Bacillus Calmette-Guerin (BCG) vaccine protects against&#xD;
           extrapulmonary infection with Mycobacterium tuberculosis and against leprosy [1, 2]. It&#xD;
           has been shown that vaccination with BCG also leads to non-specific protective effects:&#xD;
           early administration of BCG vaccination could lead to reduced child mortality, mainly as&#xD;
           a result of reduced neonatal sepsis, respiratory infection, and fever. [3-5]&#xD;
           Furthermore, BCG has also been used in patients with bladder cancer, to induce an&#xD;
           improved reaction of the immune system, which prevents tumor progression and dead. [6]&#xD;
           Recently, it was shown that BCG vaccination can induce epigenetic changes in human&#xD;
           monocytes, which result in higher pro-inflammatory cytokines response upon restimulation&#xD;
           with non-related pathogens in vitro. [7] The observed effects are proposed to be due to&#xD;
           modulation of the human innate immune system, in a process called 'trained immunity'.&#xD;
           Upon stimulation with a pathogen, the innate immune system becomes primed and is able to&#xD;
           react faster and more efficient to a secondary (and non-related) stimulus. [7, 8]&#xD;
&#xD;
           Considering these beneficial effects of BCG on innate host defense, it is reasonable to&#xD;
           hypothesize that it would be helpful to generate a 'trained immunity status' in hosts&#xD;
           defective in T- and B-cell immunity, as they are more vulnerable to infections. However,&#xD;
           because of their immune compromised status, these patients cannot be vaccinated with&#xD;
           live attenuated vaccines (e.g. BCG). [9] Vaccination with y-irradiated BCG would be&#xD;
           possible, but it is not known whether the γ-irradiated vaccine has the same protective&#xD;
           non-specific effects on innate host defense. In recent in-vitro experiments, the&#xD;
           investigators have demonstrated that y-irradiated BCG can train monocytes in vitro (not&#xD;
           published). Therefore, the aim of this study is to determine whether γ-irradiated BCG is&#xD;
           able to train the human innate immune system of healthy volunteers in-vivo, just as&#xD;
           shown before with the live attenuated BCG vaccine [7].&#xD;
&#xD;
        2. OBJECTIVES&#xD;
&#xD;
           Primary Objective: To determine whether γ-irradiated BCG can train the innate immune&#xD;
           system in vivo, by comparing cytokine production upon ex-vivo restimulation with&#xD;
           pathogens before, 2 weeks and 3 months after vaccination with y-irradiated BCG.&#xD;
&#xD;
        3. STUDY DESIGN&#xD;
&#xD;
           An explorative intervention study with a duration of three months will be performed. 15&#xD;
           Healthy volunteers will be recruited to receive a vaccination with γ-irradiated BCG.&#xD;
           After informed consent, 40ml of blood will be drawn before vaccination and at 2 weeks&#xD;
           and 3 months post-vaccination.&#xD;
&#xD;
        4. STUDY POPULATION 4.1 Population Healthy volunteers, 18 - 55 yr old. 4.2 Inclusion&#xD;
           criteria The subjects must be healthy adults, aged 18 - 55 years old. 4.3 Exclusion&#xD;
           criteria Subjects are excluded when they are from tuberculosis endemic countries, if&#xD;
           they have been in contact with tuberculosis patients or if they already have been&#xD;
           vaccinated with BCG. The subjects are excluded from the study if they suffer for any&#xD;
           disease, including intercurrent infections. Subjects are not allowed to use any&#xD;
           medication except oral anticonceptive agents.&#xD;
&#xD;
           4.4 Sample size calculation This is an explorative trial, for which sample size&#xD;
           calculation is not possible.&#xD;
&#xD;
        5. TREATMENT OF SUBJECTS 5.1 Investigational product/treatment Subjects will be vaccinated&#xD;
           with a y-irradiated BCG vaccine (BCG-Vaccin SSI [Nederlands Vaccin Instituut]), which is&#xD;
           purchased from Radboud Apotheek B.V.&#xD;
&#xD;
           γ-Irradiation of the BCG vaccine will be performed by Synergy Health Ede B.V., Ede, The&#xD;
           Netherlands (registered by the Dutch Ministry of Economic Affairs). The BCG vaccine was&#xD;
           radiated in the original packaging with an average dose of 32.2kGy in a JS6500 Tote Box&#xD;
           Irradiator. This dose is based on an in the literature accepted minimal dose of 25kGy,&#xD;
           which has been proven to be sufficient to kill M. tuberculosis [10, 11] . After&#xD;
           irradiation the vaccine will be returned unopened in the original packaging. During the&#xD;
           irradiation a dosimeter is included, so each irradiated batch will come along with a&#xD;
           certificate with the exact irradiation dose used.&#xD;
&#xD;
           5.2 Use of co-intervention Subjects are not allowed to use any medication, except for&#xD;
           oral anticonception.&#xD;
&#xD;
        6. INVESTIGATIONAL PRODUCT 6.1 Name and description of investigational product(s)&#xD;
           y-Irradiated BCG vaccine (BCG-Vaccin SSI [Nederlands Vaccin Instituut]) Danish strain&#xD;
           1331.&#xD;
&#xD;
           6.2 Summary of findings from non-clinical studies The efficacy of killing by&#xD;
           γ-irradiating of the BCG vaccine was verified by culturing the y-irradiated BCG in&#xD;
           special tuberculosis culture medium. No growth was detected after six weeks.&#xD;
&#xD;
           6.3 Summary of findings from clinical studies A related study with a comparable study&#xD;
           setup, in which 20 healthy volunteers were vaccinated with live attenuated BCG vaccine&#xD;
           has already been accomplished. [7] The investigators study will use the same study&#xD;
           protocol. The only difference is that instead of the live attenuated BCG vaccine a&#xD;
           y-irradiated BCG vaccine will be used.&#xD;
&#xD;
           6.4 Summary of known and potential risks and benefits There are no clear benefits for&#xD;
           the volunteers. Potential risks are very small, as BCG is considered a safe vaccine. BCG&#xD;
           vaccination can cause pain and scarring at the site of injection just as fever and&#xD;
           headache, and very sporadic occurring side effects as syncope, convulsions and allergic&#xD;
           reaction. Local hematoma formation can occur at the site of the blood drawing.&#xD;
&#xD;
           6.5 Description and justification of route of administration and dosage Administer at&#xD;
           the upper arm slowly, in about 10 seconds, intracutaneous 0.1ml of the suspended&#xD;
           vaccine, which accounts for 0.075mg of Mycobacterium bovis.&#xD;
&#xD;
        7. METHODS 7.1 Study parameters/endpoints Before the vaccination and at two different time&#xD;
           points after vaccination with γ-irradiated BCG (2 weeks and 3 months) blood will be&#xD;
           drawn, from which PBMCs will be isolated by density centrifugation over Ficoll-Paque.&#xD;
           Monocytes will be restimulated ex-vivo with different pathogens (M. tuberculosis&#xD;
           (1µg/ml), S. aureus, C. albicans (both 1 x 10^6 microorganisms/ml), and E. coli LPS&#xD;
           (10ng/ml). Cytokine production will be assessed after 24h (TNFa, IL 1b and IL-6), 48h&#xD;
           (IFN, IL-10) and 7 days (IL-17, IL-22) stimulation, and compared with cytokine&#xD;
           production before vaccination.&#xD;
&#xD;
      7.2 Study procedures Subjects will be vaccinated with y-irradiated BCG vaccine. Before and&#xD;
      twice after vaccination, 40ml of blood will be drawn.&#xD;
&#xD;
      7.3 Withdrawal of individual subjects Subjects can leave the study at any time for any reason&#xD;
      if they wish to do so without any consequences.&#xD;
&#xD;
      7.4 Replacement of individual subjects after withdrawal Each withdrawn subject will be&#xD;
      replaced, in order that in total 15 subjects will complete the follow-up.&#xD;
    </textblock>
  </detailed_description>
  <overall_status>Completed</overall_status>
  <start_date>November 2013</start_date>
  <completion_date type="Actual">April 2014</completion_date>
  <primary_completion_date type="Actual">March 2014</primary_completion_date>
  <phase>N/A</phase>
  <study_type>Interventional</study_type>
  <has_expanded_access>No</has_expanded_access>
  <study_design_info>
    <allocation>N/A</allocation>
    <intervention_model>Single Group Assignment</intervention_model>
    <primary_purpose>Basic Science</primary_purpose>
    <masking>None (Open Label)</masking>
  </study_design_info>
  <primary_outcome>
    <measure>Cytokine Production Measured by ELISA Compared to Baseline</measure>
    <time_frame>0 weeks and 3 months</time_frame>
    <description>Comparing tnfa production of PBMCs after 24h stimulation with candida before and 3 months after yBCG vaccination.&#xD;
Baseline is set as 1.</description>
  </primary_outcome>
  <secondary_outcome>
    <measure>Cytokine Production Measured by ELISA Compared to Baseline</measure>
    <time_frame>0 weeks and 2 weeks</time_frame>
    <description>Ex-vivo cytokine production by PBMCs upon stimulation with several pathogens</description>
  </secondary_outcome>
  <number_of_arms>1</number_of_arms>
  <enrollment type="Actual">15</enrollment>
  <condition>Innate Immune Response</condition>
  <arm_group>
    <arm_group_label>BCG vaccine SSI</arm_group_label>
    <arm_group_type>Experimental</arm_group_type>
    <description>Healthy volunteers are vaccinated with yBCG. Blood will be drawn before and at several timepoints after vaccination. Cytokine production before vaccination will be used as reference to compare later timepoints with.</description>
  </arm_group>
  <intervention>
    <intervention_type>Biological</intervention_type>
    <intervention_name>BCG vaccine SSI</intervention_name>
    <description>BCG vaccination</description>
    <arm_group_label>BCG vaccine SSI</arm_group_label>
    <other_name>J07AN01</other_name>
  </intervention>
  <eligibility>
    <criteria>
      <textblock>
        Inclusion Criteria:&#xD;
&#xD;
          -  The subjects must be healthy adults, aged 18 - 55 years old&#xD;
&#xD;
        Exclusion Criteria:&#xD;
&#xD;
          -  Subjects are excluded when they are from tuberculosis endemic countries, if they have&#xD;
             been in contact with tuberculosis patients or if they have been earlier vaccinated&#xD;
             with BCG.&#xD;
&#xD;
          -  The subjects are excluded from the study if they suffer for any disease, including&#xD;
             intercurrent infections.&#xD;
&#xD;
          -  Subjects are not allowed to use any medication except oral anticonceptive agents.&#xD;
      </textblock>
    </criteria>
    <gender>All</gender>
    <minimum_age>18 Years</minimum_age>
    <maximum_age>55 Years</maximum_age>
    <healthy_volunteers>Accepts Healthy Volunteers</healthy_volunteers>
  </eligibility>
  <verification_date>January 2016</verification_date>
  <study_first_submitted>September 26, 2014</study_first_submitted>
  <study_first_submitted_qc>October 3, 2014</study_first_submitted_qc>
  <study_first_posted type="Estimate">October 8, 2014</study_first_posted>
  <results_first_submitted>January 25, 2016</results_first_submitted>
  <results_first_submitted_qc>May 21, 2016</results_first_submitted_qc>
  <results_first_posted type="Estimate">June 29, 2016</results_first_posted>
  <last_update_submitted>May 21, 2016</last_update_submitted>
  <last_update_submitted_qc>May 21, 2016</last_update_submitted_qc>
  <last_update_posted type="Estimate">June 29, 2016</last_update_posted>
  <responsible_party>
    <responsible_party_type>Sponsor</responsible_party_type>
  </responsible_party>
  <keyword>Trained innate immunity</keyword>
  <keyword>Heterologous effects of BCG on immunity</keyword>
  <intervention_browse>
    <!-- CAUTION:  The following MeSH terms are assigned with an imperfect algorithm            -->
    <mesh_term>BCG Vaccine</mesh_term>
  </intervention_browse>

  <clinical_results>

    <participant_flow>
      <group_list>
        <group group_id="P1">
          <title>BCG Vaccine SSI</title>
          <description>Healthy volunteers are vaccinated with yBCG. Blood will be drawn before and at several timepoints after vaccination. Cytokine production before vaccination will be used as reference to compare later timepoints with.&#xD;
BCG vaccine SSI: BCG vaccination</description>
        </group>
      </group_list>
      <period_list>
        <period>
          <title>Overall Study</title>
          <milestone_list>
            <milestone>
              <title>STARTED</title>
              <participants_list>
                <participants group_id="P1" count="15"/>
              </participants_list>
            </milestone>
            <milestone>
              <title>COMPLETED</title>
              <participants_list>
                <participants group_id="P1" count="15"/>
              </participants_list>
            </milestone>
            <milestone>
              <title>NOT COMPLETED</title>
              <participants_list>
                <participants group_id="P1" count="0"/>
              </participants_list>
            </milestone>
          </milestone_list>
        </period>
      </period_list>
    </participant_flow>

    <baseline>
      <group_list>
        <group group_id="B1">
          <title>yBCG</title>
          <description>15 healthy volunteers that receive yBCG at baseline. There is no control group included in this trial.</description>
        </group>
      </group_list>
      <analyzed_list>
        <analyzed>
          <units>Participants</units>
          <scope>Overall</scope>
          <count_list>
            <count group_id="B1" value="15"/>
          </count_list>
        </analyzed>
      </analyzed_list>
      <measure_list>
        <measure>
          <title>Age</title>
          <units>Participants</units>
          <param>Count of Participants</param>
          <class_list>
            <class>
              <category_list>
                <category>
                  <title>&lt;=18 years</title>
                  <measurement_list>
                    <measurement group_id="B1" value="0"/>
                  </measurement_list>
                </category>
                <category>
                  <title>Between 18 and 65 years</title>
                  <measurement_list>
                    <measurement group_id="B1" value="15"/>
                  </measurement_list>
                </category>
                <category>
                  <title>&gt;=65 years</title>
                  <measurement_list>
                    <measurement group_id="B1" value="0"/>
                  </measurement_list>
                </category>
              </category_list>
            </class>
          </class_list>
        </measure>
        <measure>
          <title>Age</title>
          <units>years</units>
          <param>Mean</param>
          <dispersion>Full Range</dispersion>
          <class_list>
            <class>
              <category_list>
                <category>
                  <measurement_list>
                    <measurement group_id="B1" value="23" lower_limit="20" upper_limit="34"/>
                  </measurement_list>
                </category>
              </category_list>
            </class>
          </class_list>
        </measure>
        <measure>
          <title>Sex: Female, Male</title>
          <units>Participants</units>
          <param>Count of Participants</param>
          <class_list>
            <class>
              <category_list>
                <category>
                  <title>Female</title>
                  <measurement_list>
                    <measurement group_id="B1" value="9"/>
                  </measurement_list>
                </category>
                <category>
                  <title>Male</title>
                  <measurement_list>
                    <measurement group_id="B1" value="6"/>
                  </measurement_list>
                </category>
              </category_list>
            </class>
          </class_list>
        </measure>
        <measure>
          <title>Healthy</title>
          <units>participants</units>
          <param>Number</param>
          <class_list>
            <class>
              <category_list>
                <category>
                  <measurement_list>
                    <measurement group_id="B1" value="15"/>
                  </measurement_list>
                </category>
              </category_list>
            </class>
          </class_list>
        </measure>
      </measure_list>
    </baseline>

    <outcome_list>
      <outcome>
        <type>Primary</type>
        <title>Cytokine Production Measured by ELISA Compared to Baseline</title>
        <description>Comparing tnfa production of PBMCs after 24h stimulation with candida before and 3 months after yBCG vaccination.&#xD;
Baseline is set as 1.</description>
        <time_frame>0 weeks and 3 months</time_frame>
        <population>Ex-vivo cytokine production by PBMCs upon stimulation with several pathogens</population>
        <group_list>
          <group group_id="O1">
            <title>BCG Vaccine SSI</title>
            <description>Healthy volunteers are vaccinated with yBCG. Blood will be drawn before and at several timepoints after vaccination. Cytokine production before vaccination will be used as reference to compare later timepoints with.&#xD;
BCG vaccine SSI: BCG vaccination</description>
          </group>
        </group_list>
        <measure>
          <title>Cytokine Production Measured by ELISA Compared to Baseline</title>
          <description>Comparing tnfa production of PBMCs after 24h stimulation with candida before and 3 months after yBCG vaccination.&#xD;
Baseline is set as 1.</description>
          <population>Ex-vivo cytokine production by PBMCs upon stimulation with several pathogens</population>
          <units>ratio</units>
          <param>Mean</param>
          <dispersion>Full Range</dispersion>
          <analyzed_list>
            <analyzed>
              <units>Participants</units>
              <scope>Measure</scope>
              <count_list>
                <count group_id="O1" value="15"/>
              </count_list>
            </analyzed>
          </analyzed_list>
          <class_list>
            <class>
              <category_list>
                <category>
                  <measurement_list>
                    <measurement group_id="O1" value="1.2" lower_limit="0.5" upper_limit="2.2"/>
                  </measurement_list>
                </category>
              </category_list>
            </class>
          </class_list>
        </measure>
      </outcome>
      <outcome>
        <type>Secondary</type>
        <title>Cytokine Production Measured by ELISA Compared to Baseline</title>
        <description>Ex-vivo cytokine production by PBMCs upon stimulation with several pathogens</description>
        <time_frame>0 weeks and 2 weeks</time_frame>
        <group_list>
          <group group_id="O1">
            <title>BCG Vaccine SSI</title>
            <description>Healthy volunteers are vaccinated with yBCG. Blood will be drawn before and at several timepoints after vaccination. Cytokine production before vaccination will be used as reference to compare later timepoints with.&#xD;
BCG vaccine SSI: BCG vaccination</description>
          </group>
        </group_list>
        <measure>
          <title>Cytokine Production Measured by ELISA Compared to Baseline</title>
          <description>Ex-vivo cytokine production by PBMCs upon stimulation with several pathogens</description>
          <units>ratio</units>
          <param>Mean</param>
          <dispersion>Full Range</dispersion>
          <analyzed_list>
            <analyzed>
              <units>Participants</units>
              <scope>Measure</scope>
              <count_list>
                <count group_id="O1" value="15"/>
              </count_list>
            </analyzed>
          </analyzed_list>
          <class_list>
            <class>
              <category_list>
                <category>
                  <measurement_list>
                    <measurement group_id="O1" value="1.4" lower_limit="0.6" upper_limit="3.4"/>
                  </measurement_list>
                </category>
              </category_list>
            </class>
          </class_list>
        </measure>
      </outcome>
    </outcome_list>

    <reported_events>
      <group_list>
        <group group_id="E1">
          <title>BCG Vaccine SSI</title>
          <description>Healthy volunteers are vaccinated with yBCG. Blood will be drawn before and at several timepoints after vaccination. Cytokine production before vaccination will be used as reference to compare later timepoints with.&#xD;
BCG vaccine SSI: BCG vaccination</description>
        </group>
      </group_list>
      <serious_events>
        <category_list>
          <category>
            <title>Total</title>
            <event_list>
              <event>
                <sub_title>Total, serious adverse events</sub_title>
                <counts group_id="E1" subjects_affected="0" subjects_at_risk="15"/>
              </event>
            </event_list>
          </category>
        </category_list>
      </serious_events>
      <other_events>
        <frequency_threshold>5</frequency_threshold>
        <default_assessment>Non-systematic Assessment</default_assessment>
        <category_list>
          <category>
            <title>Total</title>
            <event_list>
              <event>
                <sub_title>Total, other adverse events</sub_title>
                <counts group_id="E1" subjects_affected="1" subjects_at_risk="15"/>
              </event>
            </event_list>
          </category>
          <category>
            <title>Skin and subcutaneous tissue disorders</title>
            <event_list>
              <event>
                <sub_title>Local reaction to BCG vaccin</sub_title>
                <counts group_id="E1" subjects_affected="1" subjects_at_risk="15"/>
              </event>
            </event_list>
          </category>
        </category_list>
      </other_events>
    </reported_events>

    <certain_agreements>
      <pi_employee>All Principal Investigators ARE employed by the organization sponsoring the study.</pi_employee>
      <restrictive_agreement>There is NOT 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. </restrictive_agreement>
    </certain_agreements>
    <point_of_contact>
      <name_or_title>Rob Arts</name_or_title>
      <organization>Radboud UMC</organization>
      <phone>0031 24 ext 36 67206</phone>
      <email>rob.jw.arts@radboudumc.nl</email>
    </point_of_contact>
  </clinical_results>
</clinical_study>


download date: ClinicalTrials.gov processed this data on September 20, 2023
link text: Link to the current ClinicalTrials.gov record.
url: https://clinicaltrials.gov/ct2/show/NCT02259608
org study id: NL45427.091.13
secondary id: 2013/319
nct id: NCT02259608
lead sponsor: 
has dmc: No
Rationale: The live attenuated Bacillus Calmette-Guerin (BCG) vaccine protects against
      extrapulmonary infection with Mycobacterium tuberculosis and leprosy. It has been shown that
      vaccination with BCG also leads to nonspecific protective effects, e.g. reduced infant
      mortality as a result of less severe infections, stimulation of the immune system in patients
      with bladder cancer and higher cytokine production upon restimulation of macrophages with
      non-related infectious pathogens in vitro. However, because the live attenuated BCG vaccine
      cannot be used in immune compromised hosts, the investigators would like to determine whether
      similar protective non-specific effects can be induced by γ-irradiated BCG.

      Objective: To determine whether vaccination with γ-irradiated BCG results in a higher
      cytokine response by monocytes upon restimulation in vitro with infectious pathogens,
      compared to monocytes before the vaccination.

      Study design: Explorative intervention trial. Study population: Healthy volunteers, 18 - 55
      years old. Intervention: Healthy volunteers will be vaccinated with γ-irradiated BCG vaccine.

      Main study parameters/endpoints: Blood will be drawn before and at two different time points
      after vaccination with BCG to perform restimulation of isolated cells in vitro and compare
      cytokine production.

      Nature and extent of the burden and risks associated with participation, benefit and group
      relatedness: There is no known direct benefit for the volunteers to participate in the trial.
      The risks are negligible. BCG vaccination can cause pain and scarring at the site of
      injection, just as fever and headache. Local hematoma formation can occur at the site of the
      blood drawing. This will be minimized by the blood collection by experienced persons.
1. INTRODUCTION AND RATIONALE

           The live attenuated Bacillus Calmette-Guerin (BCG) vaccine protects against
           extrapulmonary infection with Mycobacterium tuberculosis and against leprosy [1, 2]. It
           has been shown that vaccination with BCG also leads to non-specific protective effects:
           early administration of BCG vaccination could lead to reduced child mortality, mainly as
           a result of reduced neonatal sepsis, respiratory infection, and fever. [3-5]
           Furthermore, BCG has also been used in patients with bladder cancer, to induce an
           improved reaction of the immune system, which prevents tumor progression and dead. [6]
           Recently, it was shown that BCG vaccination can induce epigenetic changes in human
           monocytes, which result in higher pro-inflammatory cytokines response upon restimulation
           with non-related pathogens in vitro. [7] The observed effects are proposed to be due to
           modulation of the human innate immune system, in a process called 'trained immunity'.
           Upon stimulation with a pathogen, the innate immune system becomes primed and is able to
           react faster and more efficient to a secondary (and non-related) stimulus. [7, 8]

           Considering these beneficial effects of BCG on innate host defense, it is reasonable to
           hypothesize that it would be helpful to generate a 'trained immunity status' in hosts
           defective in T- and B-cell immunity, as they are more vulnerable to infections. However,
           because of their immune compromised status, these patients cannot be vaccinated with
           live attenuated vaccines (e.g. BCG). [9] Vaccination with y-irradiated BCG would be
           possible, but it is not known whether the γ-irradiated vaccine has the same protective
           non-specific effects on innate host defense. In recent in-vitro experiments, the
           investigators have demonstrated that y-irradiated BCG can train monocytes in vitro (not
           published). Therefore, the aim of this study is to determine whether γ-irradiated BCG is
           able to train the human innate immune system of healthy volunteers in-vivo, just as
           shown before with the live attenuated BCG vaccine [7].

        2. OBJECTIVES

           Primary Objective: To determine whether γ-irradiated BCG can train the innate immune
           system in vivo, by comparing cytokine production upon ex-vivo restimulation with
           pathogens before, 2 weeks and 3 months after vaccination with y-irradiated BCG.

        3. STUDY DESIGN

           An explorative intervention study with a duration of three months will be performed. 15
           Healthy volunteers will be recruited to receive a vaccination with γ-irradiated BCG.
           After informed consent, 40ml of blood will be drawn before vaccination and at 2 weeks
           and 3 months post-vaccination.

        4. STUDY POPULATION 4.1 Population Healthy volunteers, 18 - 55 yr old. 4.2 Inclusion
           criteria The subjects must be healthy adults, aged 18 - 55 years old. 4.3 Exclusion
           criteria Subjects are excluded when they are from tuberculosis endemic countries, if
           they have been in contact with tuberculosis patients or if they already have been
           vaccinated with BCG. The subjects are excluded from the study if they suffer for any
           disease, including intercurrent infections. Subjects are not allowed to use any
           medication except oral anticonceptive agents.

           4.4 Sample size calculation This is an explorative trial, for which sample size
           calculation is not possible.

        5. TREATMENT OF SUBJECTS 5.1 Investigational product/treatment Subjects will be vaccinated
           with a y-irradiated BCG vaccine (BCG-Vaccin SSI [Nederlands Vaccin Instituut]), which is
           purchased from Radboud Apotheek B.V.

           γ-Irradiation of the BCG vaccine will be performed by Synergy Health Ede B.V., Ede, The
           Netherlands (registered by the Dutch Ministry of Economic Affairs). The BCG vaccine was
           radiated in the original packaging with an average dose of 32.2kGy in a JS6500 Tote Box
           Irradiator. This dose is based on an in the literature accepted minimal dose of 25kGy,
           which has been proven to be sufficient to kill M. tuberculosis [10, 11] . After
           irradiation the vaccine will be returned unopened in the original packaging. During the
           irradiation a dosimeter is included, so each irradiated batch will come along with a
           certificate with the exact irradiation dose used.

           5.2 Use of co-intervention Subjects are not allowed to use any medication, except for
           oral anticonception.

        6. INVESTIGATIONAL PRODUCT 6.1 Name and description of investigational product(s)
           y-Irradiated BCG vaccine (BCG-Vaccin SSI [Nederlands Vaccin Instituut]) Danish strain
           1331.

           6.2 Summary of findings from non-clinical studies The efficacy of killing by
           γ-irradiating of the BCG vaccine was verified by culturing the y-irradiated BCG in
           special tuberculosis culture medium. No growth was detected after six weeks.

           6.3 Summary of findings from clinical studies A related study with a comparable study
           setup, in which 20 healthy volunteers were vaccinated with live attenuated BCG vaccine
           has already been accomplished. [7] The investigators study will use the same study
           protocol. The only difference is that instead of the live attenuated BCG vaccine a
           y-irradiated BCG vaccine will be used.

           6.4 Summary of known and potential risks and benefits There are no clear benefits for
           the volunteers. Potential risks are very small, as BCG is considered a safe vaccine. BCG
           vaccination can cause pain and scarring at the site of injection just as fever and
           headache, and very sporadic occurring side effects as syncope, convulsions and allergic
           reaction. Local hematoma formation can occur at the site of the blood drawing.

           6.5 Description and justification of route of administration and dosage Administer at
           the upper arm slowly, in about 10 seconds, intracutaneous 0.1ml of the suspended
           vaccine, which accounts for 0.075mg of Mycobacterium bovis.

        7. METHODS 7.1 Study parameters/endpoints Before the vaccination and at two different time
           points after vaccination with γ-irradiated BCG (2 weeks and 3 months) blood will be
           drawn, from which PBMCs will be isolated by density centrifugation over Ficoll-Paque.
           Monocytes will be restimulated ex-vivo with different pathogens (M. tuberculosis
           (1µg/ml), S. aureus, C. albicans (both 1 x 10^6 microorganisms/ml), and E. coli LPS
           (10ng/ml). Cytokine production will be assessed after 24h (TNFa, IL 1b and IL-6), 48h
           (IFN, IL-10) and 7 days (IL-17, IL-22) stimulation, and compared with cytokine
           production before vaccination.

      7.2 Study procedures Subjects will be vaccinated with y-irradiated BCG vaccine. Before and
      twice after vaccination, 40ml of blood will be drawn.

      7.3 Withdrawal of individual subjects Subjects can leave the study at any time for any reason
      if they wish to do so without any consequences.

      7.4 Replacement of individual subjects after withdrawal Each withdrawn subject will be
      replaced, in order that in total 15 subjects will complete the follow-up.
allocation: N/A
intervention model: Single Group Assignment
primary purpose: Basic Science
masking: None (Open Label)
measure: Cytokine Production Measured by ELISA Compared to Baseline
time frame: 0 weeks and 3 months
description: Comparing tnfa production of PBMCs after 24h stimulation with candida before and 3 months after yBCG vaccination.
Baseline is set as 1.
measure: Cytokine Production Measured by ELISA Compared to Baseline
time frame: 0 weeks and 2 weeks
description: Ex-vivo cytokine production by PBMCs upon stimulation with several pathogens
arm group label: BCG vaccine SSI
arm group type: Experimental
description: Healthy volunteers are vaccinated with yBCG. Blood will be drawn before and at several timepoints after vaccination. Cytokine production before vaccination will be used as reference to compare later timepoints with.
intervention type: Biological
intervention name: BCG vaccine SSI
description: BCG vaccination
arm group label: BCG vaccine SSI
other name: J07AN01
criteria: 
gender: All
minimum age: 18 Years
maximum age: 55 Years
healthy volunteers: Accepts Healthy Volunteers
responsible party type: Sponsor
mesh term: BCG Vaccine
participant flow: 
baseline: 
outcome list: 
reported events: 
certain agreements: 
point of contact: 