Clinical trial NCT03568812
Conditions: HIV Infections Immunological Abnormality
Keywords: HIV immunological non responder gut inflammation
Sponsor: Indonesia University
Start date: 2018-05-07; Completion date: 2018-12-27
Enrollment: 80 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
This study will address HIV patients who are on antiretroviral treatment (ART) and experiencing immunological failure. The cause of immunological failure in HIV patients could be due to several factors such as age, gender, Cluster of Differentiation (CD4)+ count before started treatment and some inflammation in the gut. The C-C Chemokine Receptor 5 (CCR5) receptor on T lymphocyte CD4+ are abundantly found in the gut and attacked by HIV virus during acute infection causing irreversible damage.

The disruption of gut integrity and chronic inflammation further causing translocation of bacteria in gut lumen to the blood. Thus resulting persistent low CD4+ or immunological failure.

This evaluation plan is designed to establish the role of investigation product (probiotics) to improve gut inflammation in HIV patients.

Detailed description:
HIV patients could develop immunological failure despite being treated with antiretroviral therapy (ART). Thus resulting the increase of morbidity and mortality among HIV patients, especially in Indonesia. The immunological failure is associated with decreased gut integrity and immunity, increased gut inflammation, and gut bacterial translocation to the blood. Memory CD4+ T cell damage in gut associated lymphoid tissue (GALT) by HIV-1 virus leads to decreased production of Th17 in gut mucosal layer. Furthermore, low level of Th17 facilitates bacterial translocation in HIV patients.

Probiotic supplementation (Lactobacillus) could decrease the gut inflammation in some diseases including HIV infection; thus helps increasing the gut integrity and immunity by repairing the gut mucosal tight junction, increasing mucin production, and regulating gut Th17. As a result, the gut inflammation and bacterial translocation decreases. Nevertheless, the benefit in HIV patients with immunological non responder status has not been established.

The investigators plan to perform double blind randomized clinical trial of probiotics (containing Lactobacillus plantarum, Streptococcus thermophiles, Bifidobacterium bifidum) in HIV patients with immunological non responder status. The Subject who fulfill inclusion criteria, willing to participate and sign informed consent will be randomized into two groups: the group receiving probiotics (n=40) and the group of placebo (n=40). Intervention will be held for 12 weeks. This study will evaluate the gut mucosal integrity and immunity by Th17 measurement; bacterial translocation by 16S ribosomal RNA (16S RNA) measurement; and gut inflammation by fecal calprotectin measurement before and after the intervention. Moreover, the investigators will measure the CD4+ level to assess immune status recovery before and after the intervention. Side effects, HIV symptom index, Food Frequency Questionnaire will also be assessed every 4 weeks.

Statistical analysis will uses: paired and independent t test (if normally distributed) or using Wilcoxon and Mann Whitney test (if not normally distributed). HIV symptom index and Food frequency will be reported descriptively.

Arms: arm 1: Rillus®, Chewing tablet containing viable cell 1.0 x 10\^9 colony forming unit (Lactobacillus plantarum 8.55 mg, Streptococcus thermophilus 8.55 mg, Bifidobacterium bifidum 2.55 mg, fructooligosaccharide 480 mg), isomalt, xylitol, milk flavour, vanilla flavour Dosage: 1 chewing tablet Frequency: once daily every night Duration: 12 weeks arm 2: Placebo: Chewing tablet with identical flavour, colour, smell, and size as investigational drug Dosage: 1 chewing tablet Frequency: once daily every night Duration: 12 weeks
Interventions: intervention 1: Rillus® intervention 2: Placebo
Intervention descriptions: intervention 1: Eligible subjects will be divided into 2 groups randomly (Rillus® or placebo). The colour, smell, flavour, and size of the chewing tablet are the same between active and placebo drugs. Subjects will consume the tablet once daily at night for 12 weeks. intervention 2: Eligible subjects will be divided into 2 groups randomly (Rillus® or placebo). The colour, smell, flavour, and size of the chewing tablet are the same between active and placebo drugs. Subjects will consume the tablet once daily at night for 12 weeks.

Protocol documents:
ICF_001.pdf: 
 
Informed Consent Form 
 
The Role of Probiotics L. Plantarum, S. thermophiles, B. bifidum on Gut 
Inflammation, Bacterial Translocation, and CD4+ Cell Count in 
HIV Patients with Immunological Non-Responder (PIONIR) 
 
 
 
INVESTIGATORS: 
Principal 
: Erni Juwita Nelwan, MD, Ph.D 
Others 
: Rina Agustina, MD, M.Sc., Ph.D 
  Prof. Marcellus Simadibrata, MD, Ph.D 
 
 
FACULTY OF MEDICINE 
Universitas Indonesia 
Approved October 30th, 2017 
 


RSCM
  
RSUPN Dr. Cipto Mangunkusumo 
Jl. Diponegoro No. 71 Jakarta 10430 
 
 
 
 
 
Telp: (021) 3918301   Fax: (021) 3148991 
 
             
 
Subject’s Initial _____ 
 
0265/rev02/LIT/2015 
MRN 
 
: 
Name 
 
: 
Sex 
 
: 
Date of Birth 
: 
(Please fill in the blank or put the ID sticker here) 
 
INFORMED CONSENT FORM 
 
Principal Investigator  
:  Erni J. Nelwan, MD, Ph.D 
Informant 
: 
Subject 
Name 
Date of Birth (Age) 
Sex 
Address 
Phone Number (Cellphone) 
 
: 
: 
: 
: 
: 
 
TYPE OF INFORMATION 
CONTENT OF INFORMATION  
MARK 
1 
Title of The Study 
 
The Role of Probiotics  Lactobacillus 
plantarum, 
Streptococcus 
thermophiles, 
Bifidobacterium 
bifidum 
on 
Gut 
Inflammation, Bacterial Translocation, and 
CD4+ Cell Count in HIV Patients with 
Immunological Non-Responder:  Double 
Blind Randomized Controlled Trial 
 
2 
Objectives of The Study 
 
To determine the effects of probiotics (good 
bacteria) 
Lactobacillus 
plantarum,  
Streptococcus 
thermophilus 
dan 
Bifidobacterium bifidum  in HIV patients with  
immune response failure 
 
3 
Methods of The Study 
 
You will be asked about HIV disease, 
physically examined, fill HIV symptoms 
index form, and have your blood tested for 
immune cells (CD4+ and Th17) level, viral 
load level, and bacteria translocation marker 
(16SrRNA) level. You will also have your 
stool tested for gut inflammation marker 
(fecal calprotectin) level and gut cells 
(colonocytes) level. 
We will take 12 cc of blood for test 
sample. 
You will get candies which you need to 
chew, 1 (one) tablet each day for 12 
(twelve) weeks. ARV medication should be 
continued as usual, but should not be taken 
with the candies in the same time in a day. 
You will be examined physically and have 
 


RSCM
  
RSUPN Dr. Cipto Mangunkusumo 
Jl. Diponegoro No. 71 Jakarta 10430 
 
 
 
 
 
Telp: (021) 3918301   Fax: (021) 3148991 
 
             
 
Subject’s Initial _____ 
 
0265/rev02/LIT/2015 
MRN 
 
: 
Name 
 
: 
Sex 
 
: 
Date of Birth 
: 
(Please fill in the blank or put the ID sticker here) 
 
 
 
 
 
 
 
 
the same laboratory tests as in the 
beginning of the study. 
This is a voluntary study. You may not 
participate in this study and it will not affect 
your treatment in Cipto Mangunkusumo 
National Central General Hospital. 
4 
Total of Subject 
80 people 
 
5 
Time of Study 
1 (one) year with 12 weeks intervention 
 
6 
Significances of Study 
(Including to Study 
Subjects) 
This study will bear the cost of laboratory 
study including CD4+, viral load, Th17, 
16SrRNA levels by blood test and fecal 
calprotectin and colonocytes levels by fecal 
test. These tests will be conducted 2 (two) 
times. 
This study will help doctors in understanding 
HIV patients with immune response failure 
thus increasing the quality of practice. 
 
7 
Risks and Adverse Events 
of The Study 
The risk is pain at area of blood sampling. 
The 
adverse 
events 
that 
may 
occur 
including nausea, vomiting, diarrhea which 
may 
cause 
lack 
of 
body 
fluid, 
and 
lactobacilemia which may cause fever and 
severe stomachache. 
 
8 
Study Subject 
Inconvenience 
You need to consume the candies with the 
flavor you do not like every day for 12 
(twelve) weeks. 
 
9 
Adverse Event 
Compensation 
If there is any lactobacilemia symptom, 
sepsis protocol will be executed and the 
probiotics treatment will be stopped.  
If there is lack of body fluid, a procedure of 
fluid therapy will be executed and probiotics 
treatment will be stopped. 
 
10 
Alternative Managements 
(if occurred) 
You may not participate in this study and 
take the ARV treatment as usual. 
 
 
 
 
 


RSCM
  
RSUPN Dr. Cipto Mangunkusumo 
Jl. Diponegoro No. 71 Jakarta 10430 
 
 
 
 
 
Telp: (021) 3918301   Fax: (021) 3148991 
 
             
 
Subject’s Initial _____ 
 
0265/rev02/LIT/2015 
MRN 
 
: 
Name 
 
: 
Sex 
 
: 
Date of Birth 
: 
(Please fill in the blank or put the ID sticker here) 
I have received information in page 1, 2, and 3 about the study conducted by Erni J. Nelwan, 
MD, Ph.D titled The Role of Probiotics Lactobacillus plantarum, Streptococcus thermophiles, 
Bifidobacterium bifidum on Gut Inflammation, Bacterial Translocation, and CD4+ Cell Count in 
HIV Patients with Immunological Non-Responder:  Double Blind Randomized Controlled Trial. I 
clearly understand the information. 
 
By signing this form, I agree to participate in the study voluntarily without any threat from 
anyone. I have right to cancel this agreement if I feel being harmed in any way. 
 
 
_________________________ 
Subject’s Signature or Thumbprint 
 
_________________________ 
Date 
_________________________ 
 
Subject’s Name 
 
 
_________________________ 
Subject’s Guardian/Witness Signature 
 
Date 
_________________________ 
 
Subject’s Guardian/Witness Name 
 
 
Note: Subject’s Guardian/Witness Signature is needed if the subject is illiterate, having a decreased 
consciousness, having mental disorder, or under 18 years old. 
 
 
 
 
 
TYPE OF INFORMATION 
CONTENT OF INFORMATION  
MARK 
11 
Data Confidentiality 
 
All your information will be kept and will not 
be given to any unauthorized person related 
to your treatment. 
 
12 
Cost to Subjects 
 No cost will be borne by subjects 
 
13 
Incentive for Subjects 
 
No incentive will be given to subjects 
 
14 
Name, Address, and 
Phone Number of Principal 
Investigator 
Erni J. Nelwan, MD, Ph.D 
Divisi 
Penyakit 
Tropik 
dan 
Infeksi, 
Departemen Ilmu Penyakit Dalam, FKUI-
RSCM 
Cellphone no: 021-3914190; 0811961229 
 


RSCM
  
RSUPN Dr. Cipto Mangunkusumo 
Jl. Diponegoro No. 71 Jakarta 10430 
 
 
 
 
 
Telp: (021) 3918301   Fax: (021) 3148991 
 
             
 
Subject’s Initial _____ 
 
0265/rev02/LIT/2015 
MRN 
 
: 
Name 
 
: 
Sex 
 
: 
Date of Birth 
: 
(Please fill in the blank or put the ID sticker here) 
I have given the information to subject appropriately and truthfully about the study objectives, 
significances, procedure, and potential risk and uncomforting situation which may happen 
(detailed information provided). I also have answered the questions related to the study in the 
best possible way. 
 
_________________________ 
 
 
 
 
______________________ 
Investigator’s Signature 
 
 
 
 
 
Date 
 
_________________________ 
 
 
 
 
 
Investigator’s Name  
 
 
 
 
 
 

Prot_SAP_002.pdf: 
Research Protocol 
 
The Role of Probiotics L. plantarum, S. thermophiles, B. bifidum on Gut 
inflammation, Bacterial Translocation, and CD4+ Cell Count in HIV Patients 
with Immunological Non-Responder 
(PIONIR) 
 
 
 
 
INVESTIGATORS: 
Principal       : Erni J. Nelwan, MD, Ph.D 
Others 
: Rina Agustina, M.Sc, Ph.D 
 
 
  Prof. Marcellus Simadibrata, MD, Ph.D 
 
 
  
 
 
 
 
FACULTY OF MEDICINE 
UNIVERSITY OF INDONESIA 
Approved May 1st, 2018 


 
I. Background 
Human Immunodeficiency Virus (HIV) infected patients treated with anti retroviral drugs may 
experiencing immunological failure known as immunological non responders (INR). In this 
particular group of patient, the level of CD4 still below normal limit even though the HIV virus 
is suppressed (undetected). Immunological non responders (INR) were associated with high 
morbidity.1 Hypothetically, INR patients have decreased gut integrity and immunity due to  
increased gut inflammation and further causing microbial translocation.2 Probiotics may 
improved gut inflammation condition, however the direct benefits of probiotics supplementation 
for INR HIV patients have not been studied.3 Growing number of INR HIV patients will be 
associated with higher economical burden and decreased quality of life.  
 
Microbial translocation was thought as the underlying mechanism of persistently immune 
activation which may lead to immunological failure condition in virally suppressed HIV 
patients.2 Decreased number of T- helper 17 (Th17) in HIV patients may facilitated microbial 
translocation. HIV Virus destroyed the CD4 T memory cells which mainly found in gut 
associated lymphoid tissue (GALT), thus resulting in lower production of Th17 in gut mucosal 
tissue.2,4 One of the microbial translocation marker is 16S ribosomal DNA (16S rDNA) bacteria 
blood level.5 Gut inflammation may be represented by fecal calprotectin level.6 
 
Probiotics supplementation in HIV patients was proposed may increase gut integrity and 
immunity through several mechanisms: improving tight junction gut mucosal, increasing mucin 
production, regulating Th17; therefore decreasing gut inflammation and microbial translocation. 
Improved gut inflammation and decreased microbial translocation may increased the immune 
recovery in INR HIV patients 7-9  
 
This clinical trial is intended to evaluate the immune recovery effect of Probiotics 
supplementation in INR HIV patients. 
 
 


 
II. 
Objectives  
A. General Objectives 
To determine the effect of Probiotic in immune response for immunological non 
responder HIV patients 
B. Specific Objectives 
a. Primary Objectives 
i. To determine the average CD4 value in immunological non 
responder HIV patients treated with Probiotic compared to 
placebo 
ii. To determine the average T helper 17 value in immunological non 
responder HIV patients treated with Probiotic compared to 
placebo 
iii. To determine the average 16S rDNA value in immunological non 
responder HIV patients treated with Probiotic compared to 
placebo 
iv. To determine the fecal calprotectin value in immunological non 
responder HIV patients treated with Probiotic compared to 
placebo 
b. Secondary Objectives  
To determine the effect of Probiotic supplementation towards quality of 
life improvement in immunological immunological non responder HIV 
patients, using HIV symptom index.  
 
III. 
Methods 
A. Study Design 
This study is double-blind randomized control trial design 
B. Time and Place 
This study will be conducted in Cipto Mangunkusumo National Central General 
Hospital, especially in Human Infectious Disease polyclinic. Enrollment will 
start on May 2018, and recruited until minimum sample size was fulfilled 


C. Study Population 
1. Targeted Population: HIV patients with Immunological Non-responder 
condition in Indonesia 
2. Sample: HIV patients with Immunological Non-responder condition in 
Indonesia 
3. Inclusion Criteria:  
a. HIV positive patients aged 18-55 years old 
b. had already treated with first line Highly Active Anti Retroviral 
Therapy for at least 6 months 
c. had CD4 level of 200-410 cell/L  
d. undetectable Viral Load (<34 copies/ml blood) 
e. agreed to participate in the study and signed the informed consent 
 
4. Exclusion Criteria: 
a. Pregnant patients 
b. Lactating patients 
c. Known Lactobacillus allergy 
d. Body Mass Index (BMI) <16 kg/m2 
e. Under Tuberculosis treatment or other acute illness, acute diarrhea,  
f. routinely taking selenium containing vitamin in last 1 month,  
g. routinely consuming probiotics containing product in last 1 month 
5. Estimation Sample Size: 80 samples. Which will be divided into two groups 
equally. 
6. Study Procedure 
Eligible patients will be approached to be enrolled in the clinical trial. 
Informed consent will be  obtained. Patients will be randomized and 
divided into two groups, intervention and placebo group. Intervention 
group will receive 
Probiotics 
(Chewable tablet) containing 
Lactobacillus plantarum, Stretococcus thermophiles, Bifidobacterium 
bifidum once daily. While, Placebo group will receive placebo which 
had identical shape, flavor, and smell as the interventional drug. 


Intervention will be held for 12 weeks. CD4, Th17, 16S Ribosomal 
RNA, and fecal calprotectin level will be checked at the 0 week and 
repeated after 12 weeks of intervention. Self administered HIV 
Symptoms Index questionnaire will be filled by the respondent on 
0,4,8,12 weeks of intervention. Food frequency questionnaire will be 
filled by the respondents with the help of nutritionist on 0,4,8,12 
weeks of intervention. 
 
7. Side effects 
All side effects that may arise and related to probiotics intake will be assessed 
and reported. Appropriate medical advice will be provided. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


 
 
 
 
 
D. Research Flow 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Eligibility of subjects 
Informed Consent 
Randomized 
Probiotic 
group 
Placebo 
group 
Intervention for 12 weeks 
Outcome 
Inclusion and 
Exclusion 
criteria 


 
 
 
 
 
 
E.  Ethics 
This research has been approved by Ethics Committee of the Faculty of Medicine 
Universitas Indonesia (Approval number: 981/UN2.F1/ETIK/2017) 
 
IV. 
Statistical Analysis Plan 
 
All Data will be analyzed using SPSS 24.0 (SPSS) for Windows. Laboratory parameters 
will be compared between two different times and also compared between two groups by using 
paired and independent t test (if normally distributed) or using Wilcoxon and Mann Whitney (if 
not normally distributed). Statistically significance was determined by p value <0.05.  Intention 
to Treat analysis will be used. HIV symptom index and food frequency will be reported 
descriptively. 
 
V. References 
1. Wilson EMP, Sereti I. Immune restoration after antiretroviral therapy: the pitfalls of hasty or 
incomplete repairs. Immunological reviews. 2013;254(1):343-54.  
2. Gaardbo JC, Hartling HJ, Gerstoft J, Nielsen SD. Incomplete Immune Recovery in HIV 
Infection: Mechanisms, Relevance for Clinical Care, and Possible Solutions. Clinical and 
Developmental Immunology. 2012;2012. 3. d'Ettorre G, Ceccarelli G, Giustini N, Serafino S, 
Calantone N, De Girolamo G, et al. Probiotics Reduce Inflammation in Antiretroviral Treated, 
HIV-Infected Individuals: Results of the "Probio-HIV" Clinical Trial. PloS one. 
2015;10(9):e0137200.  
4. Brenchley JM, Douek DC. The mucosal barrier and immune activation in HIV pathogenesis. 
Current opinion in HIV and AIDS. 2008;3(3):356-61.   
5. Harris KA, Hartley JC. Development of broad-range 16S rDNA PCR for use in the routine 
diagnostic clinical microbiology service. Journal of medical microbiology. 2003;52(Pt 8):685-


91.  
6. Xiang JY, Ouyang Q, Li GD, Xiao NP. Clinical value of fecal calprotectin in determining 
disease activity of ulcerative colitis. World journal of gastroenterology. 2008;14(1):53-7.  
7. Karczewski J, Troost FJ, Konings I, Dekker J, Kleerebezem M, Brummer RJ, et al. Regulation 
of human epithelial tight junction proteins by Lactobacillus plantarum in vivo and protective 
effects on the epithelial barrier. American journal of physiology Gastrointestinal and liver 
physiology. 2010;298(6):G851- 9.  
8. Anderson RC, Cookson AL, McNabb WC, Kelly WJ, Roy NC. Lactobacillus plantarum DSM 
2648 is a potential probiotic that enhances intestinal barrier function. FEMS microbiology 
letters. 2010;309(2):184- 92.  
9. Lopez P, Gonzalez-Rodriguez I, Gueimonde M, Margolles A, Suarez A. Immune response to 
Bifidobacterium bifidum strains support Treg/Th17 plasticity. PloS one. 2011;6(9):e24776.