Absorption, Distribution and Excretion
Sofosbuvir is approximately 61-65% bound to human plasma proteins and the binding is independent of drug concentration over the range of 1 ug/mL to 20 ug/mL. Protein binding of GS-331007 was minimal in human plasma. After a single 400 mg dose of (14)C-sofosbuvir in healthy subjects, the blood to plasma ratio of (14)C-radioactivity was approximately 0.7.
The pharmacokinetic properties of sofosbuvir and the predominant circulating metabolite GS-331007 have been evaluated in healthy adult subjects and in subjects with chronic hepatitis C. Following oral administration of SOVALDI, sofosbuvir was absorbed with a peak plasma concentration observed at approximately 0.5-2 hour post-dose, regardless of dose level. Peak plasma concentration of GS-331007 was observed between 2 to 4 hours post-dose. Based on population pharmacokinetic analysis in subjects with genotype 1 to 6 HCV infection who were coadministered ribavirin (with or without pegylated interferon), geometric mean steady state AUC0-24 was 969 ng*hr/mL for sofosbuvir (N=838), and 6790 ng*hr/mL for GS-331007 (N=1695), respectively. Relative to healthy subjects administered sofosbuvir alone (N = 272), the sofosbuvir AUC0-24 was 60% higher; and GS-331007 AUC0-24 was 39% lower, respectively, in HCV-infected subjects. Sofosbuvir and GS-331007 AUCs are near dose proportional over the dose range of 200 mg to 1200 mg.
Following a single 400 mg oral dose of (14)C-sofosbuvir, mean total recovery of the dose was greater than 92%, consisting of approximately 80%, 14%, and 2.5% recovered in urine, feces, and expired air, respectively. The majority of the sofosbuvir dose recovered in urine was GS-331007 (78%) while 3.5% was recovered as sofosbuvir. These data indicate that renal clearance is the major elimination pathway for GS-331007.
Studies in pregnant rats showed that sofosbuvir crossed the placenta. Fetal blood and brain sofosbuvir derived radioactivity was higher than in dams, but fetal liver and kidney had lower levels than corresponding organs in dams. Sofosbuvir-derived radioactivity was also quantifiable in milk from day 2 postpartum rats, but nursing pups did not appear to be extensively exposed to drug-derived radioactivity. Milk to plasma ratios were 0.1 at 1 hour and 0.8 at 24 hours.
For more Absorption, Distribution and Excretion (Complete) data for Sofosbuvir (6 total), please visit the HSDB record page.

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Metabolism / Metabolites
In vitro studies in human liver microsomes showed that sofosbuvir was an efficient substrate for Cathepsin A (Cat A) and carboxyl esterase 1 (CES1). There were no indications of metabolism via urdine diphosphate glucuronosyltransferases (UGTs) or flavin-containing monooxygenase (FMO). Sofosbuvir was cleaved by CatA and CES1 and subsequent activation steps included amino acid removal by histidine triad nucleotide-binding protein 1 (HINT1) and phosphorylation by uridine monophosphate-cytidine monophosphate (UMP-CMP) kinase and nucleoside diphosphate (NDP) kinase. In vitro data indicated that Cat A preferentially hydrolysed sofosbuvir (the S-diastereomer) while CES1 did not exhibit stereoselectivity. This would be consistent with studies using GS-9851 showing a less efficient metabolism to the triphosphate in the hepatically-derived cell line containing the Clone A replicon and shown to exhibit low CES 1 activity, but high Cat A activity compared with primary human hepatocytes. Following incubation of hepatocytes from rat, dog, monkey and human GS-9851 was converted to the triphosphate GS-461203 in all species, most efficiently in human. Sofosbuvir was also readily converted to the triphosphate in dog liver after oral doses and was the dominant metabolite at all time points assessed with a long half-life of approx. 18 hours. The active metabolite GS-461203 could not be detected in monkey. Further while GS-461203 was detected in rat liver, it could not be measured in liver from mouse.
Sofosbuvir is extensively metabolized in the liver to form the pharmacologically active nucleoside analog triphosphate GS-461203. The metabolic activation pathway involves sequential hydrolysis of the carboxyl ester moiety catalyzed by human cathepsin A (CatA) or carboxylesterase 1 (CES1) and phosphoramidate cleavage by histidine triad nucleotide-binding protein 1 (HINT1) followed by phosphorylation by the pyrimidine nucleotide biosynthesis pathway. Dephosphorylation results in the formation of nucleoside metabolite GS-331007 that cannot be efficiently rephosphorylated and lacks anti-HCV activity in vitro.
GS-331007 and GS-566500 were detected in all species with GS-331007 being the major drug related material in all species and all matrices. In plasma, urine and feces of all species administered sofosbuvir the primary metabolite detected was GS-331007 accounting for >80% of total exposure. In rat liver and plasma GS-566500 was also detected. The metabolite profile was overall comparable between non-pregnant, pregnant and postpartum rats and in milk of postpartum rats with GS-331007 and 2 sulfate conjugates of GS-331007 being the major metabolites.
In dog following a single oral dose of 20 mg/kg of sofosbuvir three metabolites in plasma were identified, GS-331007, GS-566500 and M4 (proposed glucuronidation product of GS-606965), accounting for 93.4%, 1.6% and 0.5%, respectively of total plasma AUC. Parent compound amounted to 4.5%. In dog (and mouse) the majority of a radioactive dose was recovered in urine within 8 to 12 hours.
For more Metabolism/Metabolites (Complete) data for Sofosbuvir (7 total), please visit the HSDB record page.

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Biological Half-Life
The median terminal half-lives of sofosbuvir and GS-331007 were 0.4 and 27 hours, respectively.

Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
Sofosbuvir has not been well studied in nursing mothers being treated for hepatitis C infection, although one infant was breastfed for 3 weeks and had no developmental abnormalities. If sofosbuvir alone or in combination with ledipasvir (Harvoni) is required by the mother, it is not a reason to discontinue breastfeeding. Some sources recommend against breastfeeding when sofosbuvir is used with ribavirin.
Hepatitis C is not transmitted through breastmilk and breastmilk has been shown to inactivate hepatitis C virus (HCV). However, the Centers for Disease Control recommends that mothers with HCV infection should consider abstaining from breastfeeding if their nipples are cracked or bleeding. It is not clear if this warning would apply to mothers who are being treated for hepatitis C.
Infants born to mothers with HCV infection should be tested for HCV infection; because maternal antibody is present for the first 18 months of life and before the infant mounts an immunologic response, nucleic acid testing is recommended.
◉ Effects in Breastfed Infants
An infant was breastfed (extent not stated) for 3 weeks postpartum by a mother who took sofosbuvir 400 mg plus ledipasvir 90 mg daily for 12 weeks beginning at 31 weeks of gestation for her chronic hepatitis C infection. The infant was followed for 1 year and found to have normal growth and development.
◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date.

Mechanism of Action
Sofosbuvir is a direct-acting antiviral agent (pan-genotypic polymerase inhibitor) against the hepatitis C virus. HCV RNA replication is mediated by a membrane-associated multiprotein replication complex. The HCV polymerase (NS5B protein) is an RNA-dependent RNA polymerase (RdRp). It is the essential initiating and catalytic subunit of this replication complex and is critical for the viral replication cycle. There is no human homolog for HCV NS5B RdRp. Sofosbuvir is a monophosphorylated pyrimidine nucleotide prodrug that undergoes intracellular metabolism to form the pharmacologically active uridine analog triphosphate (GS-461203). GS-461203 competes with natural nucleotides for incorporation (by HCV NS5B) into the nascent RNA strand during replication of the viral genome. GS-461203 differs from endogenous pyrimidine nucleotides in that it has been modified at the 2' position with the addition of a methyl and a fluoro functional group. Incorporation of GS-461203 into nascent RNA strongly reduces the efficiency of further RNA elongation by RdRp, resulting in premature termination of RNA synthesis. The stopping of viral replication leads to a rapid decline of HCV viral load and clearing of HCV levels in the body.

C22H29FN3O9P

529.452530622482

529.162

1496552-28-3

Sofosbuvir;1190307-88-0;PSI-7976;1190308-01-0;Sofosbuvir-d6;1868135-06-1;Sofosbuvir-13C,d3

90055712

White to off-white crystalline solid

1.41±0.1 g/cm3 (20 ºC 760 Torr)

1

3

11

11

36

913

6

O=[P@@](N[C@H](C)C(OC(C)C)=O)(OC[C@@H]1[C@@H](O)[C@](F)(C)[C@H](N2C=CC(NC2=O)=O)O1)OC3=CC=CC=C3

TTZHDVOVKQGIBA-UHFFFAOYSA-N

InChI=1S/C22H29FN3O9P/c1-13(2)33-19(29)14(3)25-36(31,35-15-8-6-5-7-9-15)32-12-16-18(28)22(4,23)20(34-16)26-11-10-17(27)24-21(26)30/h5-11,13-14,16,18,20,28H,12H2,1-4H3,(H,25,31)(H,24,27,30)

propan-2-yl 2-[[[5-(2,4-dioxopyrimidin-1-yl)-4-fluoro-3-hydroxy-4-methyloxolan-2-yl]methoxy-phenoxyphosphoryl]amino]propanoate

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)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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