Study to Optimize the Quality of Samples for Cell-mediated Immunity (CMI) in ART-naïve HIV-1-infected Subjects

Study Overview
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Brief Summary
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The purpose of this study is to investigate a combined set of parameters deemed to impact the quality of CMI analyses in terms of the proportion of viable lymphocytes in antiretroviral therapy-naïve HIV-1 infected subjects.

Detailed Description
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This study will address the respective and combined impact of (i) timing between blood collection and peripheral blood mononuclear cells (PBMC) processing [time-to-process] and (ii) timing of PBMC resting before stimulation [resting -time].

Official Title
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Optimizing the Quality of Samples Used for the Evaluation of Cell-mediated Immune (CMI) Responses in Antiretroviral Therapy (ART)-naïve Human Deficiency Virus Type 1 (HIV-1)-Infected Subjects

Conditions
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AIDS, AIDS Vaccines

Intervention / Treatment
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* Procedure: Blood sample collection


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: All subjects must satisfy all the following criteria at study entry: Subjects who the Investigator believes can and will comply with the requirements of the protocol. Written informed consent obtained from the subject prior to any study procedure. A male or female between and including 18 and 55 years of age at the time of enrollment. Confirmed HIV-1 infection. ART-naïve and not eligible for ART treatment as per established guidelines. Subjects must never have received ART after HIV diagnosis, including lamivudine used for chronic hepatitis B infection. The exception to this is short-term ART for prevention of mother-to-child transmission (PMTCT) which must have been completed at least 360 days prior to enrollment. Viral load level between and including 2,000 and 100,000 copies/mL at screening. CD4+ T cell count >500 cells/mm3 at screening. If the subject is female, she must be of non-childbearing potential, i.e., have a current tubal ligation, hysterectomy, ovariectomy or be post-menopausal. Female subjects of childbearing potential may be enrolled in the study, if the subject: has practiced adequate contraception for 30 days prior to vaccination, and has a negative pregnancy test at screening, and has agreed to continue adequate contraception during the entire study period. Exclusion Criteria: The following criteria should be checked at the time of study entry. If any exclusion criterion applies, the subject must not be included in the study: Infection with HIV-2. This includes subjects with dual infection with HIV-1/HIV-2. Planned use of any hematotoxic product during the study period. Planned use of any investigational or non-registered product during the study period. Acute or chronic, clinically relevant pulmonary, cardiovascular, hepatic or renal functional abnormality, as determined by physical examination, serology and/or medical history at screening. Grade 3 or grade 4 laboratory abnormalities, as defined by Division of AIDS (DAIDS) grading table, at screening. Any condition which, in the opinion of the Investigator, could compromise the subject's adherence to the study protocol. Planned administration of a vaccine not foreseen by the study protocol during the period starting 30 days before the Sample Collection Visit (Visit 2). Vaccine can be administered as after sampling in Visit 2. Pregnant or lactating female.

Ages Eligible for Study
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Minimum Age: 18 Years
Maximum Age: 55 Years

Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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No

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Basic Science
Intervention Model: Single Group Assignment
Masking: None (Open Label)


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Experimental: HIV-1 Group<br>Antiretroviral Therapy-naïve HIV1-infected subjects, aged 18 to 55 years, from whom samples for cell-mediated immunity (CMI) were collected. No investigational vaccine was administered. | Procedure: Blood sample collection<br>* Blood samples will be collected in all subjects at two time points, at the Screening Visit (Day 0) and at the Sample Collection Visit (Day 15)<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Lymphocytes Viability Prediction (LOGIT Transformed) in CMI Samples Post-overnight Incubation Time Before Intracellular Cytokine Staining (ICS): Intercept Parameter Estimate of the Prediction Model - Condition None Resting Time Not Included | The objective was to model lymphocyte viability according to time-to-process (TP=2h, 7h or 24h) and resting time (RT=2h, 6h or 18h [none resting time not included]) conditions to select the best combination of these two parameters to maximize the post-ICS viability in CMI samples collected from ART-naïve HIV-1-infected subjects. Viability (%) = 10^P/(1 + 10^P)*100 with P for Prediction.The optimum of the viability was predicted as P (LOGIT)= intercept + a*TP + b*RT + a*b*TP*RT + a*a*TP*TP + b*b*RT*RT. Where intercept, TP, RT, TP*RT, TP *TP, RT*RT are the parameters evaluated and presented in the 3 first primary outcomes. And a and b are log-transformed parameters corresponding respectively to the TP and the RT where the prediction has to be done. This outcome is presenting the intercept i.e. expected mean value of Prediction when TP and RT = 0. The optimum of this Design of Experiment is presented in outcome 4. | At Day 15 (sample collection visit) | 
| Lymphocytes Viability Prediction (LOGIT Transformed Estimate) in CMI Samples Post-overnight Incubation Time Before ICS: Time to Process and Resting Time Parameter Estimates of the Prediction Model - Condition None Resting Time Not Included | The objective was to model lymphocyte viability according to time-to-process (TP=2h, 7h or 24h) and resting time (RT=2h, 6h or 18h [none resting time not included]) conditions to select the best combination of these two parameters to maximize the post-ICS viability in CMI samples collected from ART-naïve HIV-1-infected subjects. Viability (%) = 10 ^ P / (1 + 10 ^ P) * 100 with P for Prediction.The optimum of the viability was predicted as P (LOGIT)= intercept + a*TP + b*RT + a*b*TP*RT + a*a*TP*TP + b*b*RT*RT. Where intercept, TP, RT, TP*RT, TP *TP, RT*RT are the parameters evaluated and presented in the 3 first primary outcomes. And a and b are log-transformed parameters corresponding respectively to the TP and the RT where the prediction has to be done. This outcome is presenting TP and RT estimates expressed as log(hours). The optimum of this Design of Experiment (DOE) is presented in outcome 4. | At Day 15 (sample collection visit) | 
| Lymphocytes Viability Prediction (LOGIT Transformed Estimate) in CMI Samples Post-overnight Incubation Time Before ICS: TP*RT, TP*TP and RT*RT Parameter Estimates of the Prediction Model - Condition None Resting Time Not Included | The objective was to model lymphocyte viability according to time-to-process (TP=2h, 7h or 24h) and resting time (RT=2h, 6h or 18h [none resting time not included]) conditions to select the best combination of these two parameters to maximize the post-ICS viability in CMI samples collected from ART-naïve HIV-1-infected subjects. Viability (%) = 10 ^ P / (1 + 10 ^ P) * 100 with P for Prediction.The optimum of the viability was predicted as P (LOGIT)= intercept + a*TP + b*RT + a*b*TP*RT + a*a*TP*TP + b*b*RT*RT. Where intercept, TP, RT, TP*RT, TP *TP, RT*RT are the parameters evaluated and presented in the 3 first primary outcomes. And a and b are log-transformed parameters corresponding respectively to the TP and the RT where the prediction has to be done. This outcome is presenting TP*RT, TP*TP and RT*RT estimates expressed as log(hours^2). The optimum of this DOE is presented in outcome 4. | At Day 15 (sample collection visit) | 
| Lymphocytes Viability Prediction (LOGIT Transformed Estimate) in CMI Samples Post-overnight Incubation Time Before ICS: Optimum Mean Cell Viability Estimate by the Prediction Model - Condition None Resting Time Not Included | The objective was to model lymphocyte viability according to time-to-process (TP=2h, 7h or 24h) and resting time (RT=2h, 6h or 18h [none resting time not included]) conditions to select the best combination of these two parameters to maximize the post-ICS viability in CMI samples collected from ART-naïve HIV-1-infected subjects. Viability (%) = 10 ^ P / (1 + 10 ^ P) * 100 with P for Prediction.The optimum of the viability was predicted as P (LOGIT)= intercept + a*TP + b*RT + a*b*TP*RT + a*a*TP*TP + b*b*RT*RT. Where intercept, TP, RT, TP*RT, TP *TP, RT*RT are the parameters evaluated and presented in the 3 first primary outcomes. And a and b are log-transformed parameters corresponding respectively to the TP and the RT where the prediction has to be done. The optimum predicted mean cell viability of this Design of Experiment is presented in this outcome and expressed as percentage. | At Day 15 (sample collection visit) | 
| Lymphocytes Viability Prediction (Non-transformed Estimate) in CMI Samples Post-overnight Incubation Time Before ICS: Intercept Parameter Estimate of the Prediction Model - Condition None Resting Time Included | The objective was to model lymphocyte viability according to time-to-process (TP=2h, 7h or 24h) and resting time (RT=0h) conditions in order to select the best combination of these two parameters with the aim to maximize the post-ICS viability in CMI samples collected from ART-naïve HIV-1-infected subjects. Viability (%) = 10 ^ P / (1 + 10 ^ P) * 100 with P for Prediction. The optimum of the viability was predicted as follows. P (LOGIT)= intercept + a*TP + b*RT + a*b*TP*RT + + b*b*RT*RT. Where intercept, TP, RT, TP*RT, TP *TP, RT*RT are the parameters evaluated and presented in the primary outcomes 5, 6 and 7. And a and b are parameters (hour) corresponding respectively to the TP and the RT where the prediction has to be done. This outcome is presenting the intercept i.e. expected mean value of Prediction when TP and RT = 0. The optimum of this Design of Experiment is presented in outcome 8. | At Day 15 (sample collection visit) | 
| Lymphocytes Viability Prediction (Non-transformed Estimate) in CMI Samples Post-overnight Incubation Time Before ICS: Time to Process and Resting Time Parameter Estimates of the Prediction Model - Condition None Resting Time Included | The objective was to model lymphocyte viability according to time-to-process (TP=2h, 7h or 24h) and resting time (RT=0h) conditions in order to select the best combination of these two parameters with the aim to maximize the post-ICS viability in CMI samples collected from ART-naïve HIV-1-infected subjects. Viability (%) = 10 ^ P / (1 + 10 ^ P) * 100 with P for Prediction. The optimum of the viability was predicted as follows. P (LOGIT)= intercept + a*TP + b*RT + a*b*TP*RT + + b*b*RT*RT. Where intercept, TP, RT, TP*RT, TP *TP, RT*RT are the parameters evaluated and presented in the primary outcomes 5, 6 and 7. And a and b are parameters (hour) corresponding respectively to the TP and the RT where the prediction has to be done. This outcome is presenting TP and RT estimates expressed as log(hours). The optimum of this Design of Experiment (DOE) is presented in outcome 8. | At Day 15 (sample collection visit) | 
| Lymphocytes Viability Prediction (Non-transformed Estimate) in CMI Samples Post-overnight Incubation Time Before ICS: TP*RT and RT*RT Parameter Estimates of the Prediction Model - Condition None Resting Time Included | The objective was to model lymphocyte viability according to time-to-process (TP=2h, 7h or 24h) and resting time (RT=0h) conditions in order to select the best combination of these two parameters with the aim to maximize the post-ICS viability in CMI samples collected from ART-naïve HIV-1-infected subjects. Viability (%) = 10 ^ P / (1 + 10 ^ P) * 100 with P for Prediction. The optimum of the viability was predicted as follows. P (LOGIT)= intercept + a*TP + b*RT + a*b*TP*RT + + b*b*RT*RT. Where intercept, TP, RT, TP*RT, TP *TP, RT*RT are the parameters evaluated and presented in the primary outcomes 5, 6 and 7. And a and b are parameters (hour) corresponding respectively to the TP and the RT where the prediction has to be done. This outcome is presenting TP*RT and RT*RT estimates expressed as log(hours^2). The optimum of this DOE is presented in outcome 8. | At Day 15 (sample collection visit) | 
| Lymphocytes Viability Prediction (Non-transformed Estimate) in CMI Samples Post-overnight Incubation Time Before ICS: Optimum Mean Cell Viability Estimates by the Prediction Model -Condition None Resting Time Included. | The objective was to model lymphocyte viability according to time-to-process (TP=2h, 7h or 24h) and resting time (RT=0h) conditions in order to select the best combination of these two parameters with the aim to maximize the post-ICS viability in CMI samples collected from ART-naïve HIV-1-infected subjects. Viability (%) = 10 ^ P / (1 + 10 ^ P) * 100 with P for Prediction. The optimum of the viability was predicted as follows. P (LOGIT)= intercept + a*TP + b*RT + a*b*TP*RT + + b*b*RT*RT. Where intercept, TP, RT, TP*RT, TP *TP, RT*RT are the parameters evaluated and presented in the primary outcomes 5, 6 and 7. And a and b are parameters (hour) corresponding respectively to the TP and the RT where the prediction has to be done. The optimum predicted mean cell viability of this Design of Experiment is presented in this outcome and expressed as percentage. | At Day 15 (sample collection visit) | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Percentage of Viable Lymphocytes in the CMI Samples, Post-overnight Incubation (Classic) Before ICS and Post-6 Hour Incubation Before ICS | The percentage of viable lymphocytes was determined by Forward Scatter/Side Scatter (FSC/SSC) and LIVE/DEAD gating during flow cytometry analysis for each incubation of time-to-time process (TP) = 2h, 7h and 24 h and resting time (RT) = 18h for the comparison of resting time = 18h and classic incubation time versus resting time = 18h and post-6h incubation time. | A Day 15 (sample collection visit) | 
| Magnitude of HIV-1 RT Specific Cluster of Differentiation 40 Ligand (CD40L+) CD4+ T Cell Responses in the CMI Samples Post-overnight ICS/Post 6 Hour ICS, Expressing at Least One Cytokine | Data were collected but could not be reported as data were below level of detection. | At Day 15 (sample collection visit) | 
| Magnitude of HIV-RT Specific (Background Reduced) CD8+ T Cell Responses in the CMI Samples Post-overnight ICS/Post 6 Hour ICS, Expressing at Least One Cytokine | HIV-RT specific responses of CD8+ T cells expressing at least one cytokine, among: Interleukin-2 (IL-2), Interferon-gamma (IFN-g) and Tumor necrosis factor alpha (TNF-a),after stimulation with HIV-1 peptide pools for time-to-process (TP) (2, 7, 24 hours) and resting time (RT) (0,2,6,18 hours) post-overnight ICS and for time-to-process (7 hours) and resting time (18 hours) post 6 hours ICS. | At Day 15 (sample collection visit) | 
| Number of Subjects With Serious Adverse Events (SAEs) | Serious adverse events (SAEs) assessed include medical occurrences that result in death, are life threatening, require hospitali-zation or prolongation of hospitalization or result in disability/incapacity. | During the whole study period (From Day 0 to Day 15) | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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CMI, Antiretroviral therapy-naïve, HIV