The goal of the project is to determine the difference in immune activation and HIV reservoir size between People living with HIV (PWH) on tenofovir-containing antiretroviral therapy (ART) versus PWH on nucleoside reverse transcriptase inhibitor (NRTI)-sparing ART. Tenofovir (TFV), a phosphonated nucleoside reverse transcriptase inhibitor (NRTI), is being used for oral pre-exposure prophylaxis (PrEP).

The investigators will test this hypothesis: tenofovir, and perhaps NRTIs in general, stimulate a type I/III interferon also in PWH who take these drugs. Because chronic interferon stimulation may promote the survival and proliferation of cells with integrated provirus, the investigators also hypothesize that these drugs antagonize decay of the HIV latent reservoir in PWH on ART. Consequently, the researchers hypothesize that PWH who have switched from NRTI-containing ART to NRTI-sparing ART exhibit lower type I/III interferon pathway activation and lower latent HIV reservoir size.

The investigators also hypothesize that independently of treatment, the extent of type I/III interferon activation correlates with latent HIV reservoir size.

Thus, the proposed study seeks to answer these two questions. Can the gastrointestinal epithelium be impacted by ART, and contribute to chronic immune activation and expansion of the HIV-1 reservoir? If so, what therapeutic approaches can the investigators implement to reduce the HIV-1 proviral load? The data will reveal pathways that can be targeted therapeutically to treat chronic immune activation in PWH. The findings of this study will immediately translate to optimize the standard of care in PWH.

The study will be open-label, cross-sectional, two-cohort study (20 participants per cohort will be recruited).

Cohort 1: Tenofovir-containing ART (tenofovir disoproxil fumarate [TDF] OR tenofovir alafenamide [TAF] PLUS any other ART drugs) prescribed for daily use by participants' primary care providers.

Cohort 2: NRTI-sparing ART (specifically: rilpivirine PLUS dolutegravir OR rilpivirine PLUS cabotegravir) prescribed for daily use by participants' primary care providers.

The investigators plan to test type I/III IFN (interferon) pathway activation using Crystal digital PCR (Crystal-dPCR) to quantify mRNA copy numbers of ISG15 (ISG15 ubiquitin-like modifier), MX1 (MX dynamin like GTPase 1) and IFI6 (interferon alpha inducible protein 6). RNA will be isolated from the rectal and duodenal biopsies stored in RNALater using the RNeasy Fibrous Tissue Mini Kit (Qiagen) and from the PBMC using the RNeasy Plus Mini Kit (Qiagen). Crystal-dPCR will be performed on a 6-color Naica Crystal digital PCR instrument (Stilla Technologies).

The latent HIV reservoir will be measured in DNA derived from peripheral blood mononuclear cells (PBMCs) using a novel intact/defective proviral HIV DNA Crystal-dPCR assay developed jointly by the Hladik and Jerome groups in Seattle.

Inclusion Criteria:

1. Confirmed HIV infection, by two different positive antibody tests and/or detectable plasma HIV RNA on two different dates
2. ≥18 and ≤65 years of age
3. Stable use of ART medication for ≥ 1 year
4. No switch of ART regimen within the past 180 days
5. CD4 > 350/mm3 within the past 180 days
6. HIV RNA <40 copies / mL on ≥ 2 occasions during continuous ART of ≥ 1 years, with no blip of >1000 HIV RNA copies / mL
7. Karnofsky score ≥80
8. Willingness and ability to provide informed consent for study participation
9. Willingness to undergo all required study procedures

Exclusion Criteria:

1. Active malignancy including myelodysplastic syndrome, or myeloproliferative disease within 24 weeks prior to study entry

2. Prior organ or bone marrow transplantation

3. Diagnosed autoimmune disease

4. Medical need for ongoing treatment with an immunosuppressive drug

5. Diagnosis of AIDS (defined as any AIDS-defining opportunistic infection or cancer, or a history of blood CD4+ T cell count < 200/μL)

6. Active opportunistic infection

7. Vomiting or diarrhea which prohibits consistent use of ART

8. Pregnant or breastfeeding

9. Excessive ingestion of ethanol determined by an AUDIT score of >8

10. Substance abuse

11. History of medical non-compliance

12. The following laboratory values (< 30 days before enrollment):

    • Hemoglobin < 8.5 mg/dL
    • Platelet count < 100,000/μL
    • Coagulation (PT/PTT) tests above the normal reference
    • Creatinine clearance < 60 mL/min

13. Using disallowed medications:

    • Systemic corticosteroids
    • Other immunosuppressive medications (e.g., cyclosporine, sirolimus, tacrolimus, pimecrolimus, tofacitinib)

14. BMI > 40

15. Pulmonary dysfunction.

16. Use of narcotics.

People living with HIV (PWH) on stable use of antiretroviral therapy (ART). They will be between 18 and 65 years of age. Children less than 18 years of age are excluded. Women of childbearing potential should not be pregnant to participate.

We may share information encompassing global transcriptome analysis (e.g. RNA-Seq data) derived from biopsy or peripheral blood mononuclear cells (PBMC) mRNA, DNA sequencing of the bacterial 16S RNA gene from cytobrush material (to determine microbial composition in duodenum or rectum) and HIV proviral sequencing (to characterize the HIV reservoir). HIV status and the size/structure of the HIV reservoir will be data that will be shared. Direct identifiers (e.g., names, telephone numbers, emails, social security numbers, medical record numbers, etc.) will not be shared. Data will be submitted to the National Institute of Health (NIH) Genomic Data repository, with unrestricted access (i.e. data will be made accessible to anyone via a public website). The data will be coded or anonymized prior to submission to NIH such that the identities of subjects cannot be readily ascertained or otherwise associated with the data by the repository staff or secondary data users.

Data will be shared at the time of submission of the scientific paper(s).

The information from this study will be stored in a public unrestricted data bank that anyone can use. The National Institutes of Health (NIH) has developed data banks that collect genomic study data. The NIH will store the de-identified information in these data banks for other researchers to use in future studies on any topic. This will include de-identified information about the participants' HIV status, mRNA sequences or genomic allelic variants (e.g. single nucleotide polymorphisms and copy number variants). The researchers could be from government, academic, or commercial institutions. We have a Extramural Institutional Certification dated 01/07/2022, Genomic Program Administrator: Chris Marcus from the National Institute of Allergy and Infectious Diseases (NIAID), NIH, HHS.