CLINICAL PHARMACOLOGY

Mechanism of Action

Hepatitis C virus is a single-stranded RNA virus, and its open-reading frame encodes ten structural proteins (viral capsid and envelope) and non-structural proteins (required for viral replication). NS5B is one of the non-structural proteins essential for viral RNA replication, and has been found
to be a valuable target for directly acting antiviral agents (DAAs). The uridine nucleotide analog sofosbuvir is a phosphoramidate prodrug that has to be triphosphorylated within the cells to produce its action. The required enzymes for its activation are present in the human hepatic cells, therefore, it is converted to its active metabolite during the first-pass metabolism, directly at the desired site of action: The liver. The metabolic pathway for activation of the prodrug is shown in Figure 1. This analog then mimics the physiological nucleotide and competitively blocks the NS5B polymerase, thus inhibiting the HCV-RNA synthesis by RNA chain termination. The catalytic site of the enzyme is also highly conserved across all the HCV genotypes, accounting for pan-genotypic efficacy of sofosbuvir.
Figure1

Pharmacodynamics

Effect on Electrocardiogram

The effect of sofosbuvir 400 and 1200 mg on QTc interval was evaluated in a randomized, single-dose, placebo-, and active-controlled (moxifloxacin 400 mg) four period crossover thorough QT trial in 59 healthy subjects. At a dose three times the maximum recommended dose, SOVALDI does not prolong QTc to any clinically relevant extent.

Pharmacokinetics

Absorption

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 ~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 sofosbuvir (N=838) and GS-331007 (N=1695) AUC0-24 were 828 ng•hr/mL and 6790 ng•hr/mL, respectively. Relative to healthy subjects administered sofosbuvir alone (N = 272), the sofosbuvir AUC0-24 was 39% 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.

Effect of Food

Relative to fasting conditions, the administration of a single dose of SOVALDI with a standardized high fat meal did not substantially affect the sofosbuvir Cmax or AUC0-inf. The exposure of GS-331007 was not altered in the presence of a high-fat meal. Therefore, SOVALDI can be administered without regard to food.

Distribution

Sofosbuvir is approximately 61-65% bound to human plasma proteins and the binding is independent of drug concentration over the range of 1 µg/mL to 20 µg/mL. Protein binding of GS-331007 was minimal in human plasma. After a single 400 mg dose of [14C]-sofosbuvir in healthy subjects, the blood to plasma ratio of 14C-radioactivity was approximately 0.7.

Metabolism

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.
After a single 400 mg oral dose of [14C]-sofosbuvir, sofosbuvir and GS-331007 accounted for approximately 4% and >90% of drug related material (sum of molecular weight-adjusted AUC of sofosbuvir and its

metabolites) systemic exposure, respectively.

Elimination

Following a single 400 mg oral dose of [14C]-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. The median terminal half-lives of sofosbuvir and GS-331007 were 0.4 and 27 hours, respectively.

Microbiology

Mechanism of Action

Sofosbuvir is an inhibitor of the HCV NS5B RNA-dependent RNA polyme-rase, which is essential for viral replication. Sofosbuvir is a nucleotide pro-drug that undergoes intracellular metabolism to form the pharmacologically active uridine analog triphosphate (GS-461203), which can be incorporated into HCV RNA by the NS5B polymerase and acts as a chain terminator. In a biochemical assay, GS-461203 inhibited the polymerase activity of the recombinant NS5B from HCV genotype 1b, 2a, 3a and 4a with IC50 values ranging from 0.7 to 2.6 µM. GS-461203 is not an inhibitor of human DNA and RNA polymerases nor an inhibitor of
mitochondrial RNA polymerase.

Antiviral Activity

In HCV replicon assays, the EC50 values of sofosbuvir against full-length replicons from genotype 1a, 1b, 2a, 3a and 4a, and chimeric 1b replicons encoding NS5B from genotype 2b, 5a or 6a ranged from 0.014 to 0.11 µM. The median EC50 value of sofosbuvir against chimeric replicons encoding NS5B sequences from clinical isolates was 0.062 µM for genotype 1a (range 0.029-0.128 µM; N=67), 0.102 µM for genotype 1b (range 0.045-0.170 µM; N=29), 0.029 µM for genotype 2 (range 0.014-0.081 µM; N=15) and 0.081 µM for genotype 3a (range 0.024-0.181 µM; N=106). In infectious virus assays, the EC50 values of sofosbuvir against genotype 1a and 2a were 0.03 and 0.02 µM, respectively. The presence of 40% human serum had no effect on the anti-HCV activity of sofosbuvir. Evaluation of sofosbuvir in combination with interferon alpha or ribavirin showed no antagonistic effect in reducing HCV RNA levels in replicon cells.

Resistance

In Cell Culture

HCV replicons with reduced susceptibility to sofosbuvir have been selected in cell culture for multiple genotypes including 1b, 2a, 2b, 3a, 4a, 5a and 6a. Reduced susceptibility to sofosbuvir was associated with the primary NS5B substitution S282T in all replicon genotypes examined. An M289L substitution developed along with the S282T substitution in genotype 2a, 5 and 6 replicons. Site-directed mutagenesis of the S282T substitution in replicons of 8 genotypes conferred 2- to 18-fold reduced susceptibility to sofosbuvir and reduced the replication viral capacity by 89% to 99% compared to the corresponding wild-type. In biochemical assays, recombinant NS5B polymerase from genotypes 1b, 2a, 3a and 4a expressing the S282T
substitution showed reduced susceptibility to GS-461203 compared to respective wild-types.

In Clinical Trials

In a pooled analysis of 982 subjects who received SOVALDI in Phase 3 trials, 224 subjects had post-baseline NS5B genotypic data from next generation nucleotide sequencing (assay cutoff of 1%).
Treatment-emergent substitutions L159F (n= 6) and V321A (n= 5) were de-tected in post-baseline samples from GT3a-infected subjects across the Phase 3 trials. No detectable shift in the phenotypic susceptibility to sofosbuvir of subject isolates with L159F or V321A substitutions was seen. The sofosbuvir-associated resistance substitution S282T was not detected at baseline or in the failure isolates from Phase 3 trials. However, an S282T substitution was detected in one genotype 2b subject who relapsed at Week 4 post-treatment after 12 weeks of sofosbuvir monotherapy in the Phase 2 trial P7977-0523 [ELECTRON]. The isolate from this subject displayed a mean 13.5-fold reduced susceptibility to sofosbuvir. For this subject,the S282T substitution was no longer detectable at Week 12 post-treatment by next generation sequencing with an assay cut off of 1%.
In the trial done in subjects with hepatocellular carcinoma awaiting liver transplantation where subjects received up to 48 weeks of sofosbuvir and ribavirin, the L159F substitution emerged in multiple subjects with GT1a or GT2b HCV who experienced virologic failure (breakthrough and relapse). Furthermore, the presence of substitutions L159F and/or C316N at baseline was associated with sofosbuvir breakthrough and relapse post-transplant in multiple subjects infected with GT1b HCV. In addition, S282R and L320F substitutions were detected on-treatment by next generation sequencing in a subject infected with GT1a HCV with a partial treatment response.
The clinical significance of these substitutions is not known.

Cross Resistance

HCV replicons expressing the sofosbuvir-associated resistance substitution S282T were susceptible to NS5A inhibitors and ribavirin. HCV replicons expressing the ribavirin-associated substitutions T390I and F415Y were susceptible to sofosbuvir. Sofosbuvir was active against HCV replicons with NS3/4A protease inhibitor, NS5B non-nucleoside inhibitor and NS5A inhibitor resistant variants.

Nonstructural protein 5B (NS5B) is a viral protein found in the hepatitis C virus (HCV). It has the key function of replicating the HCV's viral RNA by using the viral positive RNA strand as its template and catalyzes the polymerization of ribonucleoside triphosphates (rNTP) during RNA replication. Several crystal structures of NS5B polymerase in several crystalline forms have been determined based on the same consensus sequence BK (HCV-BK, genotype 1). The structure can be represented by a right hand shape with fingers, palm, and thumb. The encircled active site, unique to NS5B, is contained within the palm structure of the protein. Recent studies on NS5B protein genotype 1b strain J4’s (HC-J4) structure indicate a presence of an active site where possible control of nucleotide binding occurs and initiation of de-novo RNA synthesis. De-novo adds necessary primers for initiation of RNA replication. Current research attempts to bind structures to this active site to alter its functionality in order to prevent further viral RNA replication.

CLINICAL STUDIES

Phase III Trials of Sofosbuvir

Four phase III trials of sofosbuvir have been published to date, all evaluating sofosbuvir 400 mg plus ribavirin (weight-based dosing) for at least 12 weeks in patients with chronic HCV
In the FISSION study, 499 treatment-naïve patients with genotype 2 or 3 HCV were randomized to sofosbuvir 400 mg plus ribavirin for 12 weeks or peginterferon plus ribavirin for 24 weeks in a noninferiority trial. In this trial, despite a marked difference in the RVR rates (99% versus 67%, respectively), the SVR12 rates were identical at 67% [Lawitz et al. 2013d] . Notably, in this study there was a marked difference in response rates between patients with genotype 2 (97% SVR 12) and genotype 3 (56% SVR12) HCV, and between cirrhotic (47% SVR12) and noncirrhotic (72% SVR12) patients.

The NEUTRINO study was a 12-week open label study of sofosbuvir plus ribavirin in treatment naive patients with HCV genotype 1, 4, 5 or 6 (of whom 98% had genotype 1 or 4). SVR12 rates of 90% were observed overall, with 81% response rates in genotype 1 with cirrhosis. These cirrhotic patients also had extremely low rates of treatment discontinuation of only 2%, suggesting that this combination is safe and extremely well tolerated even in cirrhotic patients [Lawitz et al. 2013d].
POSITRON evaluated sofosbuvir plus ribavirin compared with placebo in patients with genotype 2 and 3 HCV in whom interferon was not an option (previously intolerant of interferon, unwilling or unable to take interferon). Overall SVR12 rates in the sofosbuvir/ribavirin group were higher than in FISSION (78%), possibly because of the higher proportion of genotype 2 pa-tients who again demonstrated significantly higher SVR12 rates (92%) than genotype 3 (68%). Degree of liver fibrosis did not appear to significantly influence outcomes in genotype 2 patients (94% SVR12 even in those with cirrhosis), but in genotype 3 patients, SVR12 rates were low at 21% in patients with cirrhosis [Jacobson et al. 2013].

The FUSION study was a blinded, active-control study involving patients with genotype 2 or 3 HCV who had not had a response to prior treatment with an interferon-containing regimen. Patients were randomly assigned to either 12 weeks of sofosbuvir and ribavirin followed by 4 weeks of matching placebo, or 16 weeksof sofosbuvir and ribavirin. One third of participants were cirrhotic. Overall rates of SVR were significantly higher in the 16-week arm (73%) than in the 12-week arm (50%), and again the genotype 2 patients had higher SVR rates (86% and 94% for 12 and 16 weeks, respectively) than genotype 3 patients (30% and 62% respectively for 12 and 16 weeks) [Jacobson et al. 2013]. Cirrhosis was associated with a decreased rate of SVR, particularly in genotype 3 receiving 12 weeks of therapy rather than 16 weeks (SVR 19% versus 61%). In genotype 2 patients SVR rates ranged from 60% to 78% in cirrhotic patients, and up to 100% in noncirrhotic patients without cirrhosis who were treated for 16 weeks.

Potential for Sofosbuvir in Liver Transplantation

One obvious clinical need is for data regarding safety and efficacy of sofos-buvir in patients who have decompensated chronic liver disease, are peri-transplant or post-liver-transplant. The excellent safety data to date and the lack of significant drug interactions makes sofosbuvir an appealing choice to be studied in these groups. To date there is one case report published of a patient with severe recurrent cholestatic hepatitis C, 6 months post-transplant, who was effectively rescued and achieved SVR with treatment with sofosbuvir and daclatasvir in combination [Fontana et al. 2013]. This is promising, and results of future trials of sofosbuvir in these types of patient groups are awaited with interest.

CURRENT STATUS

The US FDA has recently (6 December, 2013) approved sofosbuvir under the brand name Sovaldi for the treatment chronic HCV infection under a breakthrough therapy designation, because it has shown a substantial improvement over the other available therapies. The most interesting feature of this approval lies in the fact that this drug can be administered without the need of interferon therapy. On the basis of the type of HCV infection, the treatment regimen may include sofosbuvir and ribavirin/sofosbuvir, ribavirin, and Peg-interferon-alfa. Earlier it was under the FDA's priority review program (an expedited review of drugs useful for serious conditions, which, after their approval, would provide significant improvement in safety or effectiveness). Positive results from major clinical trials, plus a demonstration of efficacy in patients who cannot tolerate interferon-based regimens and in patients with liver cancer undergoing liver transplantation, make this drug a valuable and very useful therapy for these patient populations.

h2. CURIOSITY

Country

Price per 12 week treatment

Commentary

USA
$84,000
USA has the highest price in the world
Canada
$55,000
35% lower price than in USA
UK
£35,000 / $55,000
Readily available
Germany
€60,000 / $75,000
Readily available
France
€56,000 / $70,000
Readily available
Brazil
$7000*
*Government opted negotiation price
India
$900*
*Promised by Gilead Sciences; Sovaldi's producer
Egypt
$900 (with insurance)
$6,000 (without insurance)
Currently lowest available price in the world

CONCLUSIONS

Although sofosbuvir is by no means one of the first DAA drugs to reach phase III clinical trials, the collective trial data for sofosbuvir do represent a significant paradigm shift in the management of HCV infection. With sofosbuvir-based regimens, successful interferon-free treatment of HCV is now achievable across multiple genotypes, but different patterns of genotypic response to treatment have emerged compared with those seen in traditional interferon-based therapy. It is now clear that in the current DAA era, genotypes 2 and 3 should be evaluated quite separately, as SVR rates in genotype 3 patients are significantly lower than those seen in genotype 2 [Jacobson et al. 2013; Lawitz et al. 2013d]. Genotype 2 is a readily treatable strain of HCV, with superb SVR rates with sofosbuvir/ribavirin dual therapy even in patients with established cirrhosis. In contrast, the presence of advanced liver fibrosis has a significant detrimental effect on SVR rates in genotype 3 patients when a 12-week regimen is used. The FUSION data strongly suggest that for this group of patients, at least 16 weeks of therapy is needed for the dual sofosbuvir/ribavirin regimen, and the results of ongoing trials evaluating a 24-week duration of treatment are awaited with interest. The other strategy that could be considered is addition of another DAA to the regimen, but until these data are available to clarify the optimal interferon-free treatment of genotype 3 HCV patients with advanced fibrosis or previous treatment failure, there may be an initial role for continuing to use peginterferon together with sofosbuvir in this group of patients.
For genotype 1 HCV, the NEUTRINO data have shown the highest yet re-ported SVR rates for patients with genotype 1 HCV and cirrhosis when treated with sofosbuvir/ribavirin dual therapy, and this is particularly notable for the lack of significant adverse events, in contrast with the ongoing emerging data about safety of protease inhibitor/peginterferon-based triple therapy. However this dual therapy is not sufficient for the genotype 1 null responder group. Emerging phase II data in genotype 1 patients of triple therapy regimens of sofosbuvir plus ribavirin combined with an NS5A inhibitor, either daclatasvir or ledipasvir, demonstrate SVR rates close to 100%, in both treatment-naïve and null-responder patients, with potential to further shorten the duration of therapy to 8 weeks with this triple therapy approach, utilizing the fixed-dose once daily regimen of sofosbuvir/ledipasvir. Data on the effectiveness of this combination in cirrhotic genotype 1 null responders is awaited. A new era of successful interferon-free DAA therapy for HCV is emerging, with potential to broaden treatment of HCV to include patient groups who have either avoided or not been suitable for previous interferon- based therapy, and it is likely that sofosbuvir will form the backbone of this treatment approach.

During its October 2013 meeting, the CHMP gave an opinion on the conditions under which early access to sofosbuvir, in combination with other medicines, could be given in compassionate-use programmes, for patients with chronic hepatitis C infection before or after liver transplantation.
Such programmes, set up at the national level (in italy as well), are intended to give patients with a life-threatening, long-lasting or seriously disabling disease who have no available treatment options access to treatments that are still under development and that have not yet been authorised.
During its November meeting, the CHMP also provided an opinion on the use of a combination of sofosbuvir with the antiviral daclatasvir in certain patients with chronic hepatitis C virus (HCV) infection, in a compassionate-use programme.
From november 2014 this new weapon against hepatitis C it’s available also in Italy.

take home message

From the above discussion, it seems that sofosbuvir is a promising therapy for chronic HCV infection, as it offers several advantages over the existing therapies, particularly in dealing with patients with decompensated liver disease and patients who cannot tolerate interferon-containing therapies. On account of its excellent performance in clinical trials, this drug has got FDA approval on 6 December, 2013, under the breakthrough therapy designation. This drug is effective against all HCV genotypes, has a better safety profile, and low risk of development of resistance; however, careful clinical use and monitoring is still essential, to gather more data on this drug. Large post-marketing studies, including pharmacoepidemiological and pharmacovigilance studies, can solve many unanswered questions for the future of this novel drug. As of now, sofosbuvir is among the most promising agents available for the treatment of chronic HCV infection.

Sofosbuvir: A novel treatment option for chronic hepa-titis C infection

Harmeet Kaur Bhatia, Harmanjit Singh,1Nipunjot Grewal,2 and Navreet Kaur Natt3

Sofosbuvir: A novel treatment option for chronic hepatitis C infection

BIBLIOGRAPHY:
DISCOVER SOVALDI
European Medicines Agency recommends approval of

sofosbuvir for the treatment of chronic hepatitis C
Medscape
RNA polymerase NS5B

Capone Davide Michele, Fiore Paolo Ivan