Islatravir (formerly known as MK-8591; 4′-ethynyl-2-fluoro-2′-deoxyadenosine; EFdA; ISL) is a novel and potent anti-HIV-1 agent which acts as an NRTTI (nucleoside reverse transcriptase inhibitor) with anti-HIV activities. It inhibits HIV-1 (WT), HIV-1 (M184V), HIV-1 (MDR) with EC50s of 0.068 nM, 3.1 nM and 0.15 nM, respectively. Islatravir has the potential to be used as an antiviral drug for the treatment of HIV infection and AIDS. It is being developed by Merck as a subdermal drug-eluting implant to be administered.
Physicochemical Properties
| Molecular Formula | C12H12FN5O3 |
| Molecular Weight | 293.253785133362 |
| Exact Mass | 293.09 |
| Elemental Analysis | C, 49.15; H, 4.12; F, 6.48; N, 23.88; O, 16.37 |
| CAS # | 865363-93-5 |
| Related CAS # | EFdA-TP;950913-56-1; 2408129-39-3 (hydrate) |
| PubChem CID | 6483431 |
| Appearance | White to off-white solid powder |
| LogP | -0.6 |
| Hydrogen Bond Donor Count | 3 |
| Hydrogen Bond Acceptor Count | 8 |
| Rotatable Bond Count | 3 |
| Heavy Atom Count | 21 |
| Complexity | 459 |
| Defined Atom Stereocenter Count | 3 |
| SMILES | C#C[C@]1([C@H](C[C@@H](O1)N2C=NC3=C(N=C(N=C32)F)N)O)CO |
| InChi Key | IKKXOSBHLYMWAE-QRPMWFLTSA-N |
| InChi Code | InChI=1S/C12H12FN5O3/c1-2-12(4-19)6(20)3-7(21-12)18-5-15-8-9(14)16-11(13)17-10(8)18/h1,5-7,19-20H,3-4H2,(H2,14,16,17)/t6-,7+,12+/m0/s1 |
| Chemical Name | (2R,3S,5R)-5-(6-amino-2-fluoro-9H-purin-9-yl)-2-ethynyl-2-(hydroxymethyl)tetrahydrofuran-3-ol |
| Synonyms | MK-8591; 4′-ethynyl-2-fluoro-2′-deoxyadenosine; EFdA; Islatravir; 865363-93-5; 4'-Ethynyl-2-Fluoro-2'-Deoxyadenosine; MK-8591; Islatravir [USAN]; (2R,3S,5R)-5-(6-amino-2-fluoro-9H-purin-9-yl)-2-ethynyl-2-(hydroxymethyl)tetrahydrofuran-3-ol; ISLATRAVIR ANHYDROUS; ISL; MK8591; MK 8591 |
| HS Tariff Code | 2934.99.9001 |
| Storage |
Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition | Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs) |
Biological Activity
| Targets | HIV-1 (WT)(EC50=0.068 nM);HIV-1 (MDR)(EC50=0.15 nM);HIV-1 (M184V)(EC50=3.1 nM) |
| ln Vitro | EC50s for islatravir (MK-8591) (4'Ed2FA), a strong anti-HIV-1 agent, are 0.068 nM, 3.1 nM, and 0.15 nM for HIV-1 (WT), HIV-1 (M184V), and HIV-1 (MDR), respectively. It functions as a nucleoside reverse transcriptase inhibitor [1]. |
| ln Vivo | Islatravir (EFdA) treatment resulted in reduction of HIV-RNA in PB to undetectable levels in the majority of treated mice by 3 weeks post-treatment. HIV-RNA levels in cervicovaginal lavage of EFdA-treated BLT mice also declined to undetectable levels demonstrating strong penetration of EFdA into the FRT. Our results also demonstrate a strong systemic suppression of HIV replication in all tissues analyzed. In particular, we observed more than a 2-log difference in HIV-RNA levels in the GI tract and FRT of EFdA-treated BLT mice compared to untreated HIV-infected control mice. In addition, HIV-RNA was also significantly lower in the lymph nodes, liver, lung, spleen of EFdA-treated BLT mice compared to untreated HIV-infected control mice. Furthermore, EFdA treatment prevented the depletion of CD4+ T cells in the PB, mucosal tissues and lymphoid tissues[2]. |
| Enzyme Assay | An idea to use 4'-C-substituted-2'-deoxynucleoside derivatives was proposed based on a working hypothesis to solve the problems of existing acquired immune deficiency syndrome chemotherapy (highly active antiretroviral therapy). Subsequent studies have successfully proved the validity of the idea and resulted in the development of 2'-deoxy-4'-C-ethynyl-2-fluoroadenosine, a nucleoside reverse transcriptase inhibitor, which is highly potent to all human immunodeficiency viruses type 1 (HIV-1s) including multidrug-resistant HIV-1 and has a low toxicity[1]. |
| Cell Assay |
Specimen collection and processing[2] PB and CVL samples were collected longitudinally (weekly) pre- and post-HIV exposure for 6 weeks. PB was collected in EDTA and plasma separated for HIV-RNA analysis by centrifuging for 5 min at 300 g. The remaining blood cells were reconstituted with PBS to restore the original volume of the PB sample and used for flow cytometric analysis. Cervicovaginal secretions (CVS) were obtained by performing a cervicovaginal lavage (CVL, weeks 0–5) with sterile PBS (3 washes of 20 μl each, ~ 60 μl total volume). To ensure that the procedure was atraumatic, CVL were performed with 20 μl sterile filter pipette tips that were inserted no more than 1–3 mm into the vaginal cavity. Following centrifugation (300g for 5 min), cell-free supernatants were used for HIV-RNA analysis. Pellets were re-suspended in PBS and used for flow cytometric analyses. The bone marrow (BM), LN, human thymic organoid (ORG), liver, lung, spleen, GI tract (from duodenum to rectum) and FRT (vagina, cervix and uterus) were harvested at necropsy 6 weeks post-HIV exposure and mononuclear cells were isolated as previously described for HIV-RNA, HIV-DNA and flow cytometric analyses. HIV viral load and flow cytometry analysis[2] PB and CVL HIV-RNA levels were measured using one-step reverse transcriptase real-time PCR [ABI custom TaqMan Assays-by-Design (limit of detection (LOD): plasma-750 copies/ml, CVL-1400 copies/60μl). Plasma and CVL viral load levels below the limit of detection were plotted as 375 copies/ml and 700 copies/ml respectively. We used these values to calculate means for the groups. The presence of HIV-RNA and HIV-DNA in mononuclear cells isolated from tissues were determined by real-time RT-PCR (HIV-RNA, LOD-1.5 copies/105cells and HIV-DNA, LOD of 2.5 copies/105cells). As a control for the presence of DNA extracted from human cells, all samples were tested for the presence of human gamma globin DNA by real-time PCR. |
| Animal Protocol |
Virus challenge and administration of EFdA[2] Stocks of HIV-1JR-CSF were prepared via transient transfection of 293 T cells, and titred using TZM-bl cells as previously described. HIV-1JR-CSF (30,000 TCIU) was administered intravenously by tail vein injection. EFdA was reconstituted in phosphate-buffered saline (PBS) at a concentration of 1 mg/mL and administered orally to BLT mice by oral gavage at 10 mg/kg once daily for 3 weeks beginning at 3 weeks post-HIV infection. PBS (200 μL) was administered by oral gavage to (untreated) controls. |
| References |
[1]. 2'-deoxy-4'-C-ethynyl-2-fluoroadenosine, a nucleoside reverse transcriptase inhibitor, is highly potent against all human immunodeficiency viruses type 1 and has low toxicity. Chem Rec. 2006;6(3):133-43. [2]. Efficient Inhibition of HIV Replication in the Gastrointestinal and Female Reproductive Tracts of Humanized BLT Mice by EFdA. PLoS One. 2016 Jul 20;11(7):e0159517. |
| Additional Infomation |
Islatravir is an investigational drug that is being studied to treat and prevent HIV infection. Islatravir belongs to a group of HIV drugs called nucleosidereverse transcriptase translocation inhibitors (NRTTIs). NRTTIs use several different methods to block an HIV enzyme called reverse transcriptase. By blocking reverse transcriptase, NRTTIs prevent HIV from multiplying and can reduce the amount of HIV in the body. Islatravir may be effective against certain HIV strains that are resistant to other HIV drugs. Islatravir is under investigation in clinical trial NCT04233216 (Doravirine/islatravir (DOR/ISL) in Heavily Treatment-experienced (HTE) Participants for Human Immunodeficiency Virus Type 1 (HIV-1) Infection (MK-8591A-019)). Drug Indication Prevention of human immunodeficiency virus (HIV-1) infection. An idea to use 4'-C-substituted-2'-deoxynucleoside derivatives was proposed based on a working hypothesis to solve the problems of existing acquired immune deficiency syndrome chemotherapy (highly active antiretroviral therapy). Subsequent studies have successfully proved the validity of the idea and resulted in the development of 2'-deoxy-4'-C-ethynyl-2-fluoroadenosine, a nucleoside reverse transcriptase inhibitor, which is highly potent to all human immunodeficiency viruses type 1 (HIV-1s) including multidrug-resistant HIV-1 and has a low toxicity.[1] Background: The nucleoside reverse transcriptase inhibitor (NRTI) 4'-ethynyl-2-fluoro-2'-deoxyadenosine (EFdA) in preclinical development exhibits improved safety and antiviral activity profiles with minimal drug resistance compared to approved NRTIs. However, the systemic antiviral efficacy of EFdA has not been fully evaluated. In this study, we utilized bone marrow/liver/thymus (BLT) humanized mice to investigate the systemic effect of EFdA treatment on HIV replication and CD4+ T cell depletion in the peripheral blood (PB) and tissues. In particular, we performed a comprehensive analysis of the female reproductive tract (FRT) and gastrointestinal (GI) tract, major sites of transmission, viral replication, and CD4+ T cell depletion and where some current antiretroviral drugs have a sub-optimal effect. Results: EFdA treatment resulted in reduction of HIV-RNA in PB to undetectable levels in the majority of treated mice by 3 weeks post-treatment. HIV-RNA levels in cervicovaginal lavage of EFdA-treated BLT mice also declined to undetectable levels demonstrating strong penetration of EFdA into the FRT. Our results also demonstrate a strong systemic suppression of HIV replication in all tissues analyzed. In particular, we observed more than a 2-log difference in HIV-RNA levels in the GI tract and FRT of EFdA-treated BLT mice compared to untreated HIV-infected control mice. In addition, HIV-RNA was also significantly lower in the lymph nodes, liver, lung, spleen of EFdA-treated BLT mice compared to untreated HIV-infected control mice. Furthermore, EFdA treatment prevented the depletion of CD4+ T cells in the PB, mucosal tissues and lymphoid tissues. Conclusion: Our findings indicate that EFdA is highly effective in controlling viral replication and preserving CD4+ T cells in particular with high efficiency in the GI and FRT tract. Thus, EFdA represents a strong potential candidate for further development as a part of antiretroviral therapy regimens.[2] |
Solubility Data
| Solubility (In Vitro) |
DMSO : ~100 mg/mL (~341.01 mM ) H2O : ~3.57 mg/mL (~12.17 mM) |
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.08 mg/mL (7.09 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 2.08 mg/mL (7.09 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. Solubility in Formulation 3: ≥ 2.08 mg/mL (7.09 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly. Solubility in Formulation 4: ≥ 1.1 mg/mL (3.75 mM) (saturation unknown) in 5% DMSO + 40% PEG300 + 5% Tween80 + 50% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 5: ≥ 1.1 mg/mL (3.75 mM) (saturation unknown) in 5% DMSO + 95% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. Solubility in Formulation 6: 10% DMSO+40% PEG300+5% Tween-80+45% Saline: ≥ 2.08 mg/mL (7.09 mM) Solubility in Formulation 7: 1.35 mg/mL (4.60 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.4101 mL | 17.0503 mL | 34.1006 mL | |
| 5 mM | 0.6820 mL | 3.4101 mL | 6.8201 mL | |
| 10 mM | 0.3410 mL | 1.7050 mL | 3.4101 mL |