HIV-1

BMS-663068 is a prodrug of the small-molecule inhibitor Temsavir/BMS-626529, which inhibits human immunodeficiency virus type 1 (HIV-1) infection by binding to gp120 and interfering with the attachment of virus to CD4+ T-cells.The activity of BMS-626529 is virus dependent, due to heterogeneity within gp120. In order to better understand the anti-HIV-1 spectrum of BMS-626529 against HIV-1, in vitro activities against a wide variety of laboratory strains and clinical isolates were determined. BMS-626529 had half-maximal effective concentration (EC(50)) values of <10 nM against the vast majority of viral isolates; however, susceptibility varied by >6 log(10), with half-maximal effective concentration values in the low pM range against the most susceptible viruses. The in vitro antiviral activity of BMS-626529 was generally not associated with either tropism or subtype, with few exceptions. Measurement of the binding affinity of BMS-626529 for purified gp120 suggests that a contributory factor to its inhibitory potency may be a relatively long dissociative half-life. Finally, in two-drug combination studies, BMS-626529 demonstrated additive or synergistic interactions with antiretroviral drugs of different mechanistic classes. These results suggest that BMS-626529 should be active against the majority of HIV-1 viruses and support the continued clinical development of the compound.[1]

The maximum median decrease in plasma HIV-1 RNA load from baseline ranged from 1.21 to 1.73 log(10) copies/mL. Plasma concentrations of BMS-626529 were not associated with an antiviral response, while low baseline inhibitory concentrations and the minimum and average steady-state BMS-626529 plasma concentrations, when adjusted by the baseline protein binding-adjusted 90% inhibitory concentration (inhibitory quotient), were linked with antiviral response. BMS-663068 was generally well tolerated. Conclusions: Administration of BMS-663068 for 8 days with or without ritonavir resulted in substantial declines in plasma HIV-1 RNA levels and was generally well tolerated. Longer-term clinical trials of BMS-663068 as part of combination antiretroviral therapy are warranted. Clinical Trials Registration.NCT01009814.[J Infect Dis. 2012 Oct 1;206(7):1002-11]

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BMS-663068, also known as fostemsavir, demonstrates strong antiviral activity against subjects harboring virus infection and IC50 values less than 100 nM[1].

Affinity and off-rates of attachment inhibitors from gp120.[1]
Micro BioSpin 6 columns were used to measure the binding of [3H]BMS-488043 or [3H]BMS-626529 to gp120. Binding solutions (30 μl) containing 25 mM Tris-HCl (pH 7.5), 125 mM NaCl, 50 nM gp120JRFL, and serial dilutions of [3H]BMS-488043 or [3H]BMS-626529 were allowed to equilibrate and then adsorbed to a MicroBioSpin 6 column. The column was centrifuged (∼14,000 rpm) for 5 min, the eluent was collected, and radioactivity was determined with a scintillation counter. To measure dissociative kinetics, 150 nM [3H]BMS-626529 or 90 nM [3H]BMS-488043 was incubated with 60 nM gp120 at ambient temperature for 1 h to achieve equilibrium binding, and then a large molar excess (14-fold) of soluble CD4 protein was added to drive dissociation. Aliquots were taken at the indicated time intervals, adsorbed to a spin column, and centrifuged, and the radioactivity in the eluent was quantitated. Comparison of the tritium signal from parallel samples with and without the soluble CD4 challenge allowed for the determination of the percent compound bound.[1]

Cytotoxicity assays. [1]
Cytotoxicity assays were performed in the presence of serially diluted BMS-626529 for up to 6 days, and cell viability was quantitated using an XTT (2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxanilide) assay. To determine CC50 values (concentration of drug required to kill 50% of cells), laboratory-adapted cells were initially plated at a density of 0.1 × 106 cells/ml. In the absence of compounds, the cell densities typically reached 1.0 × 106 to 1.2 × 106/ml after 6 days.

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Drug susceptibility assays using laboratory virus strains. [1]
MT-2 (for CXCR4 or dual-tropic viruses) or PM1 (for CCR5-tropic viruses) cells were infected with virus at a multiplicity of infection of 0.005 and incubated in the presence of serial dilutions of drug at 37°C for 4 to 6 days. Virus yields were quantified by determination of reverse transcriptase (RT) activity for CXCR4 viruses or by a p24 enzyme-linked immunosorbent assay for CCR5 viruses.

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Drug susceptibility assays using clinical isolates. [1]
Pellets of PBMCs were infected with clinical isolates at a multiplicity of infection of 0.005 and incubated in a 0.5-ml volume at 37°C for 3 h prior to resuspension in medium and addition to plates containing serial dilutions of drug. The final cell density was 1 × 106 cells/ml. Plates were incubated at 37°C, and virus yields were monitored from day 5 postinfection by using a p24 ELISA kit according to the manufacturer's instructions. The incubation was terminated when the control infection yielded a level of p24 in the supernatant within a dynamic range (0.6 < A490 < 2.0).

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Drug susceptibility assays using envelopes derived from clinical isolates. [1]
Plasma samples obtained during Bristol-Myers Squibb-sponsored trials were tested by Monogram Biosciences, together with additional samples from the Monogram collection. Drug susceptibilities of the envelopes were determined using the PhenoSense Entry assay. Envelope sequences (gp160) were amplified by reverse transcriptase PCR (RT-PCR) and ligated into the pCXAS expression vector. Envelope expression vectors were prepared as large pools of sequences (>200) in order to ensure an accurate representation of the diversity of viral quasispecies present in each sample. Recombinant HIV-1 pseudovirus stocks were prepared by cotransfecting HEK293 cells with the envelope expression vectors and a replication-defective HIV-1 genomic vector containing luciferase within the deleted envelope region. Recombinant pseudovirus particles were used to infect U87 cell lines expressing CD4/CCR5/CXCR4. Drug susceptibility was measured by comparison of luciferase activities in the presence and absence of BMS-626529. Drug susceptibility data were provided as half-maximal inhibitory concentration (IC50) values by Monogram Biosciences and are reported as such.

Fifty HIV-1-infected subjects were randomized to 1 of 5 regimen groups (600 mg BMS-663068 plus 100 mg ritonavir every 12 hours [Q12H], 1200 mg BMS-663068 plus 100 mg ritonavir every bedtime, 1200 mg BMS-663068 plus 100 mg ritonavir Q12H, 1200 mg BMS-663068 Q12H plus 100 mg ritonavir every morning, or 1200 mg BMS-663068 Q12H) for 8 days in this open-label, multiple-dose, parallel study. The study assessed the pharmacodynamics, pharmacokinetics, and safety of BMS-663068.[J Infect Dis. 2012 Oct 1;206(7):1002-11.]

Absorption, Distribution and Excretion
The absorption of temsavir is significantly limited by suboptimal dissolution and solubility following oral administration. Fostemsavir, a phosphonooxymethyl prodrug of temsavir, has improved aqueous solubility and stability under acidic conditions as compared to its parent drug - following oral administration of fostemsavir, the absolute bioavailability is approximately 26.9%. The Cmax and AUCtau following oral administration of fostemsavir 600mg twice daily was 1770 ng/mL and 12,900 ng.h/L, respectively, with a Tmax of approximately 2 hours. Co-administration of fostemsavir with a standard meal increases its AUC by approximately 10%, while co-administration with a high-fat meal increases its AUC by approximately 81%.
Temsavir is highly metabolized, after which it is excreted in the urine and feces as inactive metabolites. Approximately 51% of a given dose is excreted in the urine, with <2% comprising unchanged parent drug, and 33% is excreted in the feces, of which 1.1% is unchanged parent drug.
The steady-state volume of distribution of temsavir following intravenous administration is approximately 29.5 L.
The mean clearance and apparent clearance of temsavir, the active metabolite of fostemsavir, are 17.9 L/h and 66.4 L/h, respectively.

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Metabolism / Metabolites
Fostemsavir is rapidly hydrolyzed to temsavir, its active metabolite, by alkaline phosphatase(s) present at the brush border membrane of the intestinal lumen. Temsavir undergoes further biotransformation to two predominant inactive metabolites: BMS-646915, a product of hydrolysis by esterases, and BMS-930644, an N-dealkylated metabolite generated via oxidation by CYP3A4. Approximately 36.1% of an administered oral dose is metabolized by esterases, 21.2% is metabolized by CYP3A4, and <1% is conjugated by UDP-glucuronosyltransferases (UGT) prior to elimination. Both temsavir and its two predominant metabolites are known to inhibit BCRP.

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Biological Half-Life
The half-life of temsavir is approximately 11 hours. Fostemsavir is generally undetectable in plasma following oral administration.

Hepatotoxicity
In registration clinical trials, fostemsavir was associated with alanine aminotransferase (ALT) elevations in up to 25% of patients, but levels above 5 times the upper limit of normal (ULN) arose in only 4% of subjects. Most ALT elevations were transient, asymptomatic, and did not require dose modification or discontinuation. The more marked ALT elevations were usually attributable to other conditions or complications of HIV infection. No convincing cases of fostemsavir induced liver injury were observed in preregistration trials. Since approval of fostemsavir for use as a part of a multidrug therapy of HIV, there have been no published case reports of clinically apparent liver injury attributed to its use.
Interestingly, in the large preregistration trial of fostemsavir, elevations in serum aminotransferase levels were particularly noted in patients with coinfection with either hepatitis B virus (HBV) or hepatitis C virus (HCV). The deaths from liver disease in this trial appeared to be due to worsening of the coinfection during therapy. Clearly, patients with HBV or HCV coinfection should be treated for those viral infections before or concurrent with antiretroviral therapy with fostemsavir.
Likelihood score: E* (unproven but suspected cause of clinically apparent liver injury).

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Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
No information is available on the use of fostemsavir during breastfeeding. Because the drug and its active metabolite temsavir are over 80% protein bound, the amounts in milk are likely to be low. Achieving and maintaining viral suppression with antiretroviral therapy decreases breastfeeding transmission risk to less than 1%, but not zero. Individuals with HIV who are on antiretroviral therapy with a sustained undetectable viral load and who choose to breastfeed should be supported in this decision. If a viral load is not suppressed, banked pasteurized donor milk or formula is recommended.
◉ Effects in Breastfed Infants
Relevant published information was not found as of the revision date.
◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date.

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Protein Binding
Temsavir is approximately 88.4% protein-bound in plasma, primarily to serum albumin.

[1]. In vitro antiviral characteristics of HIV-1 attachment inhibitor BMS-626529, the active component of the prodrug BMS-663068. Antimicrob Agents Chemother. 2012 Jul;56(7):3498-507.

Fostemsavir (brand name: Rukobia) is a prescription medicine approved by the U.S. Food and Drug Administration (FDA) for the treatment of HIV infection in treatment-experienced adults who meet certain requirements, as determined by a health care provider. Fostemsavir is always used in combination with other HIV medicines.
Fostemsavir is the phosphonooxymethyl prodrug of temsavir, a novel HIV-1 attachment inhibitor. It binds to and inhibits the activity of gp120, a subunit within the HIV-1 gp160 envelope glycoprotein that facilitates the attachment of HIV-1 to host cell CD4 receptors - in doing so, temsavir prevents the first step in the HIV-1 viral lifecycle. The discovery of gp120 as a potential target of interest in the treatment of HIV-1 infection is relatively recent, and was born out of a desire to find alternative target proteins (i.e. mechanistically orthogonal therapies) for the treatment of HIV-1 patients with resistant infections. Fostemavir is the first attachment inhibitor to receive FDA approval, granted in July 2020 for use in combination with other antiretrovirals in highly treatment-experienced patients with multidrug-resistant HIV-1 infection whom are failing their current therapy. Targeting gp120 subunits is a new and novel therapeutic approach to HIV-1 infection, and the addition of attachment inhibitors, like temsavir, to the armament of therapies targeted against HIV-1 fills a necessary niche for therapeutic options in patients left with few, if any, viable treatments.
Fostemsavir is a unique antiretroviral agent that binds to an envelope antigen of the human immunodeficiency virus (HIV) inhibiting its attachment to cell surface receptors of CD4+ lymphocytes. It is used to treat patients with multidrug resistant infection and inadequate viral suppression despite optimized background therapy. Fostemsavir has been linked to a low rate of serum aminotransferase elevations during therapy but has not been linked convincingly to episodes of clinically apparent liver injury.

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Drug Indication
Fostemsavir is indicated, in combination with other antiretrovirals, for the treatment of multidrug-resistant HIV-1 infection in heavily treatment-experienced adults failing their current antiretroviral therapy due to resistance, intolerance, or safety concerns.
Rukobia, in combination with other antiretrovirals, is indicated for the treatment of adults with multidrug resistant HIV-1 infection for whom it is otherwise not possible to construct a suppressive anti-viral regimen.

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Mechanism of Action
The gp120 subunit within the gp160 envelope glycoprotein of HIV-1 is a new and novel target in the treatment of HIV-1 infection. These subunits are responsible for facilitating the first step in the viral life cycle, attachment, by mediating the interaction between the virus and host cell CD4 receptors. Following attachment, HIV-1 undergoes assembly, budding, and maturation within the host cell, after which mature viral particles are released to continue the viral life cycle. Fostemsavir's active metabolite, temsavir, is an HIV-1 attachment inhibitor. It binds directly to the gp120 subunit to inhibit viral interaction with host CD4 receptors, thereby preventing the initial attachment required for viral replication. It has also been shown to inhibit other gp120-dependent post-attachment steps required for viral entry.

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Pharmacodynamics
Temsavir inhibits the first stage in the HIV-1 viral lifecycle: attachment. It has a moderate duration of action necessitating twice-daily dosing. Fostemsavir, administered at roughly 4x the recommended human dose, has been observed to significantly prolong the QTc-interval. Patients with a history of QTc-prolongation, those receiving other QTc-prolonging medications, and/or those with pre-existing cardiac disease should use fostemsavir with caution, and should be monitored at baseline and throughout therapy for signs or symptoms suggestive of QTc-prolongation. Fostemsavir should also be used with caution in patients with hepatitis B or C co-infection as elevations in hepatic transaminases were observed in greater proportions in these populations in clinical trials.

C25H26N7O8P

583.48984

583.158

C, 51.46; H, 4.49; N, 16.80; O, 21.94; P, 5.31

864953-29-7

Temsavir;701213-36-7;Fostemsavir Tris;864953-39-9; Temsavir;701213-36-7; 864953-29-7(free base); 864953-39-9 (tromethamine) ; 864953-31-1 (disodium); 942117-71-7 (dihydrate)

11319217

White to off-white solid powder

1.6±0.1 g/cm3

904.1±75.0 °C at 760 mmHg

500.6±37.1 °C

0.0±0.3 mmHg at 25°C

1.723

-2.98

2

11

8

41

1020

0

O=C(C1C2C(=C(N3C=NC(C)=N3)N=CC=2OC)N(COP(O)(O)=O)C=1)C(N1CCN(C(C2C=CC=CC=2)=O)CC1)=O

SWMDAPWAQQTBOG-UHFFFAOYSA-N

InChI=1S/C25H26N7O8P/c1-16-27-14-32(28-16)23-21-20(19(39-2)12-26-23)18(13-31(21)15-40-41(36,37)38)22(33)25(35)30-10-8-29(9-11-30)24(34)17-6-4-3-5-7-17/h3-7,12-14H,8-11,15H2,1-2H3,(H2,36,37,38)

(3-(2-(4-benzoylpiperazin-1-yl)-2-oxoacetyl)-4-methoxy-7-(3-methyl-1H-1,2,4-triazol-1-yl)-1H-pyrrolo[2,3-c]pyridin-1-yl)methyl dihydrogen phosphate

BMS-663068; BMS663068; BMS-663068 dihydrate; BMS 663068; Fostemsavir; 864953-29-7; BMS-663068 free acid; Fostemsavir [USAN]; Fostemsavir(BMS-663068); 97IQ273H4L;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~100 mg/mL ( ~171.38 mM )
H2O : ~20 mg/mL (~34.28 mM)

Solubility in Formulation 1: 8.33 mg/mL (14.28 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication (<60°C).

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 (Please use freshly prepared in vivo formulations for optimal results.)