Clinical Trial to Evaluate the Effect of Raltegravir Intensification (1.200 mg QD) on the Gut Microbiota of Chronically HIV-1 Infected Subject Over Time: THE RAGTIME STUDY

The gut microbiome is essential for the maturation of the neonatal immune system and the adequate development and function of adult immune responses. HIV-1 infection in children and adults exerts a rapid and severe depletion of gut-associated lymphoid tissue, which damages the intestinal barrier, allowing translocation of gut commensal bacteria into the systemic circulation. Bacterial translocation causes chronic inflammation and immune activation, which lead to immune deterioration and premature aging of HIV-1-infected subjects, including metabolic disturbances, cardiovascular diseases, cognitive disorders and HIV-associated cancers. Persistence of residual HIV-1 replication in the presence of ART has been associated to incomplete HIV-1 suppression in gut lymphatic tissues due to suboptimal tissular penetration of PI/s or NNRTIs.

In previous work in our institute, the investigators have observed that HIV-1 infection is independently associated with significant reductions in the gut microbiome richness, which is, in turn, are inversely correlated with systemic inflammation. Reduced microbial richness, for example, has been associated with intestinal inflammatory diseases and well as with metabolic syndrome, diabetes and obesity and correlated with metabolic markers.

Recovering bacterial richness might thus have a positive impact on immune activation, chronic inflammation and the overall health of HIV-infected individuals. However, achieving that goal will possibly require, alongside potential bacterial supplementations, the use of ART with high penetration into gut lymphoid tissue to limit as much as possible the continued damage exerted by residual HIV replication on the GALT. Antiretroviral drugs with higher intestinal penetration like raltegravir may be more effective at recovering the intestinal microbiome composition and function than those with lower gut penetration like darunavir or the NNRTIs. Thereby, raltegravir intensification could be associated with increases in intestinal microbial richness, implying an improvement on intestinal and overall health.

Despite the lack of evidence on that regard, previous studies from our group and others would favor that hypothesis. Residual HIV-1 replication in plasma can be deterred by ART intensification with raltegravir, which is, in part, due to the high penetration of raltegravir in intestinal tissues. Moreover, raltegravir intensification decreases peripheral CD8 T-cell activation CD45RA (-) and creates a transient CD4 T-cell redistribution, which revert after raltegravir withdrawal.

The project presented here will be the first prospective, randomized evaluation of the effect of ART on the structure and function of the gut microbiome. This study provides a unique opportunity to understand the benefits of ART with high intestinal penetration on the gut microbiome. It is thus a key study to understand the bidirectional interactions between the microbiome and the host in people living with HIV/AIDS

• Raltegravir (Drug), also known as NP: Raltegravir 1200 mg (2 tablets x 600mg) once daily plus current ART during 48 weeks from randomization
• Placebo (Drug), also known as NP: Placebo (2 tablets of placebo) once daily plus current ART during 48 weeks from randomization.
• Current ART (Drug), also known as NP: 3-drug antiretroviral treatment including PI/r/c or NNRTI

Primary outcomes

• Bacterial Gene Richness (Observed Unique Genes). Analysis of the Composition, Structure and Function of the Intestinal Microbiome. * [From baseline to 48 weeks]: -* Structure and composition of the microbiome. DNA will be extracted and purified from fecal samples and cryopreserved at -80ºC until amplification. The purified DNA will be amplified using Illumina-tagged primers to amplify the V3 and V4 16S ribosomal DNA (rDNA) regions. PCR reactions will be performed in triplicate to preserve diversity. Pooled triplicates will be sequenced ensuring adequate sampling depth. -Function of the bacteriome. The gene content will be inferred from the abundance of each bacteria in the intestinal bacteriome according to the 16S rDNA information

Secondary outcomes

• Association of the Gut Microbiome With Inflammation Markers [From baseline to week 48]: IL-6, IP-10
• Association of the Gut Microbiome With Coagulation [From baseline to 48 weeks]: D-Dimer
• Association of the Gut Microbiome With Enterocyte Damage [From baseline to 48 weeks]: Intestinal Fatty Acid Binding Protein (I-FABP)
• Association of the Gut Microbiome With Bacterial Translocation and Monocyte Activation [From baseline to 48 weeks]: LPS-binding protein (LBP), soluble CD14
• Mean Fluorescence Intensity (MFI) Measure of Maturation, Activation, Exhaustion and Immune Senescence Markers in CD4+ and CD8+ T-cells [Differences at week 48]: CCR7, CD28, CD27, HLA-DR, CD38, PD-1, CD57
• Association of the Gut Microbiome Composition and Richness With CD4 and CD8+ Counts. [Baseline]: CD4 and CD8+ counts
• Other Estimators of Richness and Diversity [From baseline to 48 weeks]: Shannon
• Association of the Gut Microbiome Composition and Richness With CD4/CD8+ Cell Ratio. [Baseline]: CD4/CD8+ cell ratio

Sex: All, ages: 18 Years to 99 Years

Inclusion Criteria:

1. Age ≥18 years old
2. Documented HIV infection
3. Stable 3-drug antiretroviral treatment including PI/r/c or NNRTI for at least 6 months.
4. Plasma HIV-1 RNA load <50 copies/mL for at least 12 months.
5. Signed Informed Consent

Exclusion Criteria:

1. PI/r monotherapy
2. INSTI therapy during the previous 6 months
3. Evidence of previous INSTI resistance
4. Creatine clearance <50 mL/min
5. Child- Pugh B or C
6. History of active uncontrolled GI disorders or diseases including:

   6.1. Major surgery of the GI tract, with the exception of cholecystectomy and appendectomy, in the previous 5 years.

   6.2. Any major bowel resection at any time.

   6.3. Any chronic digestive disease such as peptic ulcer, Crohn's disease, ulcerative colitis, coeliac disease, confirmed intolerance to lactose or indeterminate colitis.

   6.4. Persistent infectious gastroenteritis, colitis or gastritis; persistent or chronic diarrhea of unknown etiology; Clostridium difficile infection (recurrent) or Helicobacter pylori infection (untreated)

   6.5. Irritable bowel syndrome (moderate-severe)

   6.6. Chronic constipation

   6.7. Active proctitis
7. Antibiotic therapy within the previous 2 months
8. In women, pregnancy or breastfeeding*.

   * Female subjects of childbearing potential must not be pregnant, not be planning a pregnancy or breast-feeding. Sexually active women must be willing to use two approved methods of contraception (including condoms, diaphragm, spermicides, hormonal methods and/or intrauterine devices) from baseline until the end of the clinical trial. Sexually active men in heterosexual relationships must be willing to use two approved method of contraception with their partners from baseline until the end of the clinical trial.
   * condom use is considered as an additional method of contraception only and cannot be the only method of contraception used as not been considered an effective method by the Clinical Trial Facilitation Group (CTFG) guidelines.

Participant flow

Outcome results

• Bacterial Gene Richness (Observed Unique Genes). Analysis of the Composition, Structure and Function of the Intestinal Microbiome. * (Primary), Median, Number of unique genes detected: Current ART + Raltegravir 768358 (663708 to 879113); Current ART + Placebo 730732 (552699 to 869340)
• Association of the Gut Microbiome With Inflammation Markers (Secondary), Median, pg/ml: Current ART + Raltegravir 1.32 (1.14 to 1.78); Current ART + Placebo 1.53 (1.41 to 1.78)
• Association of the Gut Microbiome With Coagulation (Secondary), Median, ug/ml: Current ART + Raltegravir 1.99 (1.77 to 2.41); Current ART + Placebo 2.1 (1.91 to 2.89)
• Association of the Gut Microbiome With Enterocyte Damage (Secondary), Median, ng/ml: Current ART + Raltegravir 0.92 (0.68 to 1.33); Current ART + Placebo 0.69 (0.6 to 0.89)
• Association of the Gut Microbiome With Bacterial Translocation and Monocyte Activation (Secondary), Median, ng/ml: Current ART + Raltegravir 4238 (3395 to 4537); Current ART + Placebo 4274 (3664 to 5056)
• Mean Fluorescence Intensity (MFI) Measure of Maturation, Activation, Exhaustion and Immune Senescence Markers in CD4+ and CD8+ T-cells (Secondary), Median, MFI: Current ART + Raltegravir 8051 (1471 to 12096); Current ART + Placebo 8076 (1624 to 12144)
• Association of the Gut Microbiome Composition and Richness With CD4 and CD8+ Counts. (Secondary), Median, cells/mm^3 of blood: Current ART + Raltegravir 696 (554 to 918); Current ART + Placebo 664 (430 to 658)
• Other Estimators of Richness and Diversity (Secondary), Median, Shannon index: Current ART + Raltegravir 2.90 (2.45 to 3.16); Current ART + Placebo 2.56 (1.86 to 3.08)
• Association of the Gut Microbiome Composition and Richness With CD4/CD8+ Cell Ratio. (Secondary), Median, Ratio: Current ART + Raltegravir 1.10 (0.74 to 1.26); Current ART + Placebo 0.91 (0.82 to 1.62)

Adverse events

Time frame: From baseline to week 48

Other adverse events by term

• Study Protocol and Statistical Analysis Plan (PDF, 2017-02-10)