STUDY TITLE: A randomized, double blind, placebo-controlled
study to evaluate the immunomodulatory role of Bacillus coagulans SNZ 1969 in adults.
PROTOCOL
NUMBER: SBPL/Immuno/NK/2023
VERSION NO. 01, DATED 11/09/2023
PROTOCOL....................................................................................................................... 1
1 STUDY TITLE............................................................................................................. 4
2 INTRODUCTION & BACKGROUND............................................................................... 5
2.1 IMMUNITY IN ADULTS.................................................................................... 5
3 STUDY AIM AND OBJECTIVES...................................................................................... 7
3.1 AIM............................................................................................................... 7
4 STUDY DESIGN.......................................................................................................... 8
5 STUDY SITE AND DURATION....................................................................................... 9
6 STUDY POPULATION- SAMPLE SIZE........................................................................... 10
7 TREATMENT ARMS.................................................................................................. 11
8 STUDY OUTCOME MEASURES.................................................................................. 12
9 ELIGIBILITY CRITERIA................................................................................................ 13
10 METHODS AND ASSESSMENTS............................................................................. 14
10.1 Randomization and Blinding:........................................................................ 14
10.2 Study Visits:................................................................................................. 14
11 ETHICAL CONSIDERATIONS.................................................................................. 15
11.1 Informed Consent........................................................................................ 15
11.2 IRB review................................................................................................... 15
11.3 Confidentiality of Data and
Participant Records:............................................ 15
12 STATISTICAL ANALYSIS......................................................................................... 16
13 PARTICIPANT WITHDRAWAL AND
REPLACEMENT.................................................. 17
14 LOST TO FOLLOW-UP........................................................................................... 18
15 PREMATURE TERMINATION OR SUSPENSION
OF TRIAL.......................................... 19
16 ADVERSE EVENT MANAGEMENT.......................................................................... 20
16.1 Definition of Adverse Event
(AE)................................................................... 20
16.2 Definition of Adverse Drug
Reaction (ADR).................................................... 20
16.3 Definition of Unexpected
Adverse Drug Reaction........................................... 20
16.4 Definition of Serious Adverse
Events............................................................. 20
16.5 Recording of AE........................................................................................... 21
16.6 Recording of SAE.......................................................................................... 22
16.7 Assessment of Intensity of
AE/SAE................................................................ 23
16.8 Assessment of Causality of AE
&SAE -WHO-UMC
Causality Assessment System (Annexure 2) 23
16.9 Assessment of Outcome of AE
&SAE............................................................. 24
16.10 Notification of SAE................................................................................... 24
17 PUBLICATION...................................................................................................... 25
18 FUNDING............................................................................................................ 26
Annexure 1. Schedule of events & study flow
chart.......................................................... 27
Annexure 2. WHO-UMC system for standardized case
causality assessment...................... 28
20. REFERENCES............................................................................................................. 29
1
STUDY
TITLE
A randomized, double blind, placebo-controlled study
to evaluate the immunomodulatory role of Bacillus coagulans SNZ 1969 in adults.
1. Able to detect abnormal
cells and directly kill them without any specific antigens. (NK cells can
detect abnormal cells such as cancer cells and remove them immediately using
various receptors without specific antigens.)
2. Controls the immune
system. (NK cells regulate immune and inflammatory responses by inducing the
activity of acquired immune cells (Dendritic cell, T-cell, B-cell)
3. Effectively inhibits the
proliferation, recurrence, and metastasis of cancer cells. (NK cells
effectively remove the Cancer Stem Cells (CSC) and Circulating Tumor Cell
(CTC).)
NK Cell
Activity Test is a blood test that quantifies the innate immune system’s
ability to function. The significance of NK Cell Activity
Test - NK cells, why not NUMBERS, but ACTIVITY? Among NK cells, not all NK
cells can fight abnormal cells, such as cancer cells. There are many NK cells
that lack the ability to attack those abnormal cells. In other words, having
high number of NK cells does not necessarily mean that the antitumor immunity
is high. It is the highly active NK cells that matters to maintain our immune
system stronger. The Advantage of NK Cell Activity Test is that it measures the
activity of NK cells using a small amount of blood and get results in a short
time. It measures NK cell activity using whole blood, which includes both
innate immune cytotoxicity and immunomodulation of the adaptive immune
response. Natural killer (NK) cells are identified by the absence of CD3 and
the expression of CD16 and/or CD56. NK cells are divided according to the
expression of CD16 and CD56 into cytotoxic NK cells (CD3-CD16+CD56dim) that
represents approximately 90 percent of NK cells, and cytokine-secreting or
regulatory NK cells (CD3-CD16-CD56bright) that represents approximately 10
percent of NK cells. NK cells act against virally-infected cells and tumor
cells and may be increased or decreased in various immunologic abnormalities.
NK cells also have a role in the adaptive immune response through cytokine
production. The cytotoxic NK cells (CD3-CD16+CD56dim) and cytokine-secreting or
regulatory NK cells (CD3-CD16-CD56bright) are reported as a percentage of the
total NK cells.
Regular
monitoring of the immune system to maintain good health in elderly subjects is
recommended at least once in a year.
Prospective
clinical studies conducted found probiotics especially Bacillus coagulans to be
effectively modulating the immune system and thereby improving the immunity in
elderly subjects. (1,2)
2.2 IMMUNOMODULATORY ROLE OF PROBIOTICS IN ELDERLY
Probiotics are defined by the World Health
Organization as “live microorganisms which when administered in adequate
amounts confer a health benefit on the host.†Nutraceutical dietary regime
which may offer promise as a restorative agent for immune function is the use
of probiotic supplements, such as lactic acid bacteria (LAB). Some strains of
LAB have been shown to be potent modulators of NK cell function in rodent
models (3,4) and can effectively reduce tumor growth or cytomegalovirus
infection following oral or systemic delivery (3,4,5,6). In human studies, LAB
have been shown to increase NK cell activity in colorectal cancer patients and
to limit the recurrence of superficial tumors in bladder cancer patients (7,8).
Probiotics that can enhance key aspects of immune function have recently been
highlighted for their potential benefits to human health (9,10). Since many
probiotic LAB are derived from fermented foodstuffs already in common use,
there is the possibility that defined strains of LAB may be used as biological
agents to benefit health, including geriatric health to boost the immunity and
confer resistance to combat infections in general.
The present study is being undertaken to determine
whether dietary consumption of LAB – Bacillus coagulans SNZ 1969 probiotic
strains could significantly affect NK cell parameters in elderly subjects.
To evaluate the immunomodulatory
role of oral probiotic Bacillus coagulans SNZ 1969
in adults.
PRIMARY OBJECTIVES
To evaluate the NK Cell cytolytic
activity (NK cell activity assay) and Proportion of volunteers showing change
in NK cell activity as a marker of Immunity in adult subjects – at Baseline and
at the end of the treatment of 12 weeks with Bacillus coagulans SNZ 1969.
To evaluate the absolute number and
percentage of NK cells as measured by CD16/CD56 positive cells - at Baseline
and at the end of the treatment of 12 weeks with Bacillus coagulans SNZ 1969.
To evaluate Immunoglobulins
(Serum Ig M/Serum Ig G/Salivary Ig A) - at Baseline and at the end of the
treatment of 12 weeks with Bacillus coagulans SNZ 1969.
SECONDARY OBJECTIVES
- To record and assess the occurrence
(Incidence + Duration) of gastrointestinal infections and respiratory
infections throughout the study duration of 12 weeks.
- To record and assess the total number of days
of illness OR the number of emergency medical visits
- To observe the serious infections
necessitating antibiotic use.
- To record CRP as a marker of inflammation - at
Baseline and at the end of the treatment of 12 weeks with Bacillus coagulans
SNZ 1969.
A
randomized, double blind, placebo controlled study to evaluate the immunomodulatory
role of oral probiotic Bacillus coagulans SNZ 1969 in adults.
All the participants will be
randomized in a 1:1 ratio into two groups of 25 each:
•
Group A: n =25
Probiotic Bacillus coagulans SNZ 1969 NLT
2 Billion CFU in Capsule form administered once daily.
•
Group B: n =25
Identical Placebo capsule containing Maltodextrin
administered once daily
All the participants will be
treated with the above supplements for a period of 12 weeks.
Study site: Department of General
Medicine / Geriatrics
The total study duration is
expected to be approximately 6 months with screening period, treatment period
of 12 weeks of supplementation.
A sample size of 50 completed subjects was
calculated with 25 subjects in each group (n=25) with a power of 80% at the 5%
level of statistical significance. Considering 20% dropout rate, the
total no of subjects to be randomized and enrolled will be 60 subjects with 30
subjects in each group.
A Total
of 50 elderly subjects in the age group of 60 – 65 years in good health with no chronic
diseases like HTN/DM/CAD/RA/any chronic illness, but susceptible to
cold/cough/fever during seasonal changes throughout the year.
Such elderly subjects will be selected
from study sites.
|
Group
|
Study medication
|
Medication Frequency
|
Sample size
|
|
A
|
Probiotic Bacillus coagulans SNZ 1969
2 Billion CFU / Capsule
|
Once daily
|
25
|
|
B
|
Identical Placebo capsule containing
Maltodextrin
|
Once daily
|
25
|
Each participant will be identified using a center number, a treatment
number (provided by the treatment code found in the intervention drug label)
and the participant’s initials.
Primary Outcome:
·
To evaluate the NK Cell cytolytic activity (NK cell activity
assay) and Proportion of volunteers showing change in NK cell activity as a
marker of Immunity in adult subjects – at Baseline and at the end of the
treatment of 12 weeks with Bacillus coagulans SNZ 1969.
·
To evaluate the absolute number and
percentage of
NK cells as measured by CD16 CD56 positive cells - at Baseline and at the end
of the treatment of 12 weeks with Bacillus coagulans SNZ 1969.
·
Improvement of Immunoglobulin levels (Serum Ig M/Serum Ig G/Salivary Ig A) at the end of 12 weeks
supplementation.
Secondary Outcomes:
•
Reduction in occurrence (Incidence + Duration) of gastrointestinal
infections and respiratory infections.
•
Reduction in total number of days of illness OR
the number of emergency medical visits.
•
Reduction in serious infections necessitating
antibiotic use.
•
Reduction in CRP levels (marker of inflammation)
All the Participants will be
evaluated as per the scheduled visits until 12 weeks of supplementation.
INCLUSION CRITERIA:
All the following criteria must be
met at screening prior to randomization and at enrolment: Candidates for inclusion in the study are
clinically stable elderly subjects of either gender in the age group of 60 to
65 years in general good health but susceptible to cold or cough or fever
during seasonal changes throughout the year.
EXCLUSION CRITERIA:
1.
Any Chronic diseases like Hypertension, Diabetes, Coronary Artery
Disease, Rheumatoid Arthritis OR any chronic illness.
2.
Active dependency on controlled substances and alcohol,
3.
Not willing to sign the Informed Consent Form
4.
Administration of an investigational drug either currently or
within 30 days of screening.
5.
Active addictive drug or alcohol use or dependence that, in the opinion
of the investigator, would interfere with adherence to study requirements or
assessment.
6.
Any illness or condition that in the opinion of the investigator
may affect the safety of the participant or the evaluation of any study endpoint.
7.
Significant pre-existing cardiac or pulmonary or hepatic or neurological
co-morbidities
8.
Patients with social conditions or medical debilitating disease/disorder,
which, in the judgment of the investigator, would interfere with or serve as a
contraindication to adherence to the study protocol or ability to give informed
consent or affect overall prognosis of the patient.
Randomization will be performed using a computer-generated
randomization numbers. Investigators, participants and research associates will
be blinded to treatment. Double
blinding will be accomplished by independent blinding of the dosing kits.
Visit 1 – Screening / Baseline /
Randomization
Visit 2 – 12 Weeks after initiation of
supplementation
Each subjects parent will be provided with
detailed participant information sheet (PIS), in the language he understands. Before
participation in the study, Assent from either parent will be obtained and
their sign on written informed consent form (ICF), in the language he/she
understands, in front of the investigator. Both the PIS and ICF will be read
out and explained to the subjects parents in case he/she is an illiterate.
The study will be performed consistent with
the ICH/GCP guidelines and to the ethical principles that have their origin in
the Declaration of Helsinki. Ethical approval of the study will be obtained
from the IRB/Institutional Ethics Committee of respective study center before
the study is initiated.
The identity of the subjects and data
generated in the study will be handled with strict confidence. Data will be
available only to the investigators/auditors involved in the study and to the EC
members and regulatory authorities.
This confidentiality is extended to cover
testing of biological samples addition to the clinical information relating to
the participating subjects.
Data
will be compiled and analysed as
per the study objectives for both safety and efficacy. All
analyses will be performed in the per-protocol (PP) set, i.e. treated participants
that would have no major protocol violations, will meet the minimum protocol
requirements, and who will able to be evaluated for the primary endpoint. Differences
between the groups will be evaluated using appropriate statistical tests.
All values will
expressed as means ± standard deviation (SD). A ‘p’ value <0.05 will
be considered as statistically significant. The baseline descriptors will be
summarized as mean and standard deviation for continuous variables and as
frequencies and percentages for categorical variables.
A study participant will be withdrawn/ discontinued at
any time from the study if any of the following situations occur:
1.
Study participant / parent wants to withdraws voluntarily
from the study
2.
Lost to follow-up
3.
At the discretion of the
Investigator (e.g., rapid clinical deterioration)
4.
Study termination
5.
Any treated study participant who
has an adverse reaction to study medication that threatens his/her well-being.
The study participant should be monitored for resolution of the adverse event.
6.
Significant deterioration in the
study participant’s clinical status
7.
Failure of study participant for
whatever reason to comply with requirements of the protocol
8.
The study participant dies during
the study period
When a study participant is removed from the
study, the Investigator will clearly document the reason in the medical record
and complete the appropriate case report form page describing the reason for
discontinuation. In addition, every effort will be made to complete the
appropriate assessments.
If a trial participant fails to appear for a
follow-up examination, extensive effort (i.e., documented phone calls) should
be undertaken to locate or recall him/her or at least to determine his/her
health status. These efforts should be documented in the trial participants CRF
and another pertinent source document.
Any study participant who is not available
for the final follow up should be classified as “lost to follow-up†and the
classification noted on the CRF together with the reason, if known.
This study may be temporarily suspended or
prematurely terminated if there is sufficient reasonable cause. Written
notification, documenting the reason for study suspension or termination, will
be provided by the suspending or terminating party. If the study is prematurely
terminated or suspended, the PI will promptly inform the IEC and will provide
the reason(s) for the termination or suspension.
Circumstances that may warrant termination or
suspension include, but are not limited to:
·
Determination of unexpected
significant, and unacceptable risk to participants
·
Insufficient compliance to
protocol requirements
·
Data that are not sufficiently
complete and/or evaluable
·
Determination of futility
(pointlessness or uselessness)
Any
untoward medical occurrence in a participant or clinical investigation subject, administered a IMP and which does not necessarily have to have a causal
relationship with this treatment. All
conditions, which are pre-existing prior to study drug administration, must be
recorded on the participant CRF.
An
adverse event (AE) can therefore be any unfavorable and unintended sign (including an abnormal laboratory
finding, for example), symptom, or disease temporally associated with the
use of a medicinal product, whether or not considered related to the
medicinal product.
All noxious and unintended responses to a medicinal
product related to any dose should be considered adverse drug reactions.
The
phrase "responses to an investigational medicinal product" means that
a causal relationship between an IMP
and an adverse event is at least a reasonable possibility, i.e., the
relationship cannot be ruled out.
An adverse reaction,
the nature or severity of which is not consistent with the applicable product
information (e.g., Investigator’s Brochure for an unapproved investigational
medicinal product).
Over-dose
of IMP
without resulting in abnormal findings, worsening of the study disease if
expected, and hospitalization on admission due to study disease will be
not considered as an adverse event.
The
term "severe" is often used to describe the intensity (severity) of a specific event (as in mild,
moderate, or severe myocardial infarction); the event itself, however, may be of
relatively minor medical significance (such as severe headache). This is not
the same as "serious," which is based on participant/event outcome or
action criteria usually associated with events that pose a threat to a participant’s
life or functioning. Seriousness (not severity) serves as a guide for defining
regulatory reporting obligations.
A
serious adverse event is any untoward medical occurrence that at any dose:
- Results
in death
- Is
life
threatening*
- Leads
to or prolongs hospitalization
- Leads
to a persistent or significant disability or incapacity
- Leads
to a congenital
abnormality or birth defect
- Anything
else leading
to any of the above or requiring intervention to prevent them
- Anything
determined by the investigator to be serious
NOTE: The term
"life-threatening" in the definition of "serious" refers to
an event in which the participant was at risk of death at the time of the
event; it does not refer to an event which hypothetically might have caused
death if it were more severe.
Medical and scientific judgment should be
exercised in deciding whether expedited reporting is
appropriate in other situations, such as important medical events that may not be immediately
life-threatening or result in death or hospitalization but may jeopardize the participant or may require
intervention to prevent one of the other outcomes listed in
the definition above. These should also usually be considered
serious.
Examples
of such events are intensive treatment in an emergency room or at home for allergic bronchospasm;
blood dyscrasias or convulsions that do not result in hospitalization; or
development of drug dependency or drug abuse.
Non-serious (for e.g. AE) and serious adverse
events should be evaluated as two distinct events given their different medical
nature. If an event meets the criteria to be determined “seriousâ€, the
Investigator to the extent should be able to determine all contributing factors
applicable to each serious adverse event and examine it.
Baseline
assessment should be performed immediately prior to the study medication
administration and should be transcribed on to the CRF. If the signs and
symptoms continue and worsen during the follow-up period, they will then be
considered as adverse events and transcribed in the Adverse Event section of
the CRF.
No
AEs will be recorded prior to Day 0. AEs, if it occurs should be
evaluated by the Investigator or his/her delegate and documented in terms of a
medical diagnosis corresponding to a specific WHO preferred term.
The
occurrence of all AE should be documented in the CRF with the following type of
information where appropriate:
1.
Nature of adverse event
(symptoms/ description/ diagnosis)
2.
When the adverse event first
occurred (date and time)
3.
Date of resolution (date and
time)
4.
Intensity of the adverse event
(Mild, moderate, severe)
5.
Relationship to IMP (Definite, probable, possible,
Not related)
6.
Action taken (None, treatment
continued; additional treatment prescribed; and treatment stopped)
7.
Outcome (Recovered with
treatment, Recovered without treatment, Event continuing without additional
treatment, Event continuing and controlled with additional treatment, Event
continuing and not controlled with additional treatment, Died, Unknown)
Adverse
events already documented in the CRF, i.e. at a previous visit and designated,
as ‘ongoing’ should be reviewed as necessary. If these have resolved, the
documentation in the CRF should be completed.
The
occurrence of all SAE should be documented in the CRF with the following type
of information where appropriate:
1.
Study participant identifiers
2.
Demographics
3.
Details of event
§ Date
of report (The alert form should be faxed within 24 hours, followed by the detailed report of the Study
participant within 72 hours of the onset of the SAE)
§ Date
SAE first known
§ When
the event first occurred (date and time)
§ When
the event ended (date and time)
§ Whether
SAE ended
§ Study
drug administered by
§ Name
of responsible PI
§ Type
of report (initial, follow-up)
4.
Type of SAE:
Life-threatening/fatal, congenital anomaly, required/ prolonged hospitalization,
permanently disabling, required intervention to prevent permanent impairment/
damage, or any other)
5.
Nature: Diagnosis, description,
and confirmatory tests
6.
How long the adverse event
persisted. Whether the event was once or intermittent (ideally each occurrence
of an AE will be reported. However, certain adverse events may occur
frequently, such as vomiting or diarrhea, and it is more sensible to record
these as a single event with an intermittent periodicity if the intervals are
less than 24 hours).
7.
Relevant medical history
8.
Preexisting medical history
9.
Family history (similar adverse
events seen in the family members with the same drug)
10. Causality
11. Relationship
to IMP
12. Action
taken
13. Outcome
(Recovered with treatment, Recovered without treatment, Event ongoing without
additional treatment, Event continuing and controlled with additional
treatment, Event continuing and not controlled with additional treatment, Hospitalization,
Died)
14. Concomitant
medications
All Study participants
experiencing adverse events, whether considered associated with the use of the
IMP or not, must be monitored until symptoms
subside and any clinically relevant changes of laboratory values have returned
to baseline, or until there is a satisfactory explanation for the changes
observed, or until death, in which case a full pathologist’s report should be
supplied, if possible. All findings must be reported in the study participant’s
file (source document) and in the AE and SAE forms (available as loose sheets).
The
Investigator should assess the intensity of adverse event and grade accordingly
as described below:
- None
- Mild: An adverse event, which is easily
tolerated by the Study participant, causing minimal discomfort but not
interfering with everyday activities
- Moderate: An adverse event, which is
sufficiently discomforting and interfering with everyday activities
- Severe: An adverse event, which prevents
normal everyday activities and requires medical treatment.
16.8 Assessment of Causality of AE &SAE -WHO-UMC
Causality Assessment System (Annexure 2)
Every effort should be made by
the Investigator to explain each adverse event and assess its causal
relationship, if any, to IMP administration.
Emergency
Room Visits:
Events that result in emergency room visits that do not result in admission to
the hospital are not routinely considered to be serious events; however, these
events should be evaluated for one of the other serious outcomes [e.g.,
life-threatening, other serious (medically important) events].
The
degree of certainty with which an adverse event can be attributed to treatment
administration (or alternative causes, e.g. natural history of the underlying
diseases, concomitant therapy, etc.) will be determined by how well the event
can be understood in terms of one or more of the following:
- Reaction of similar nature having
previously been observed with this type of treatment
- The event having often been reported in
literature for similar types of treatments
- The event being temporally associated
with study drug administration or reproduced on re-administration.
The
relationship of an adverse event to IMP should be classified as defined
in the WHO-UMC system for standardized case causality assessment
(Annexure 2).
The outcome should be assessed
as:
- Recovered with treatment
- Recovered without treatment
- Event continuing without additional
treatment
- Event continuing and controlled with
additional treatment
- Event continuing and not controlled with
additional treatment
- Died
- Unknown (not for SAE)
Any
Unexpected Adverse Events or Serious Adverse Events including death due to any
cause, which occurs to any Study participant entered into treatment in this
study or within 30 days following cessation of treatment, whether or not
related to the IMP
must be reported within 24 hours by email/ telephone to the overseeing Ethics
Committee.
All
SAEs will be reported promptly to regulatory authority, Sponsor (or its
representative) and the Ethics Committee as per local regulations. SAEs
follow-up reports will be reported as per applicable local regulations. SAE
report, after due analysis, will be submitted by Sponsor/designee and study
Investigator(s), as per the applicable local regulations.