Antiviral research has been stymied for a long time by the fact that viruses, as recently as 10 years ago, were difficult to grow outside the human body. "It has really only been the last couple of years that the aspect of limiting technology for HCV has been resolved," said Joanna Boerner, scientific associate with Novartis (www.novartis.com). "Getting the virus to replicate outside of a host is a tricky thing."
At the ൛th International Conference on Antiviral Research (ICAR)," held recently in Puerto Rico, some of the best breakthroughs in the field of antiviral research were presented. Important progress that has been made in the therapeutic areas of hepatitis C was detailed. Worldwide, more than 170 million people are infected with this potentially fatal disease. The hepatitis C virus (HCV) causes liver inflammation, and when the disease becomes chronic, it can lead to cirrhosis, liver failure, and cancer. Liver failure due to chronic hepatitis C is the leading cause of liver transplantation today.
Virtually Trolling for Targets
Migenix (www.migenix.com) develops drugs to treat or prevent infectious, metabolic, or degenerative diseases. In addition to the development of celgosivir for HCV, currently in Phase II, Migenix is making use of computational-based screening to cut down the time and expense of identifying novel HCV leads.
Dale Cameron, director of research (anti-infectives) and manager, intellectual property, presented a poster at ICAR, explaining how he identified two novel HCV inhibitors using virtual screening. "The work I presented was actually a small, back-up project related to our main nonnucleoside project," Cameron said. "Using OpenEye Scientific Software screening tools together with publicly available HCV inhibitor structural information and commercial databases proved to be an effective way to identify novel HCV leads."
Cameron chose NS5B, an RNA-dependent RNA polymerase that has been demonstrated to be essential for viral replication and has also been validated in the clinic, as his target. He also took advantage of publicly available coordinates for ligand protein complexes, published structure-activity-relationships, and the availability of an in-house assay for biological screening of the virtual hits. And he was amazed and impressed with the results.
"It resulted in two reasonable hits out of only 32 compounds," he reported. "And virtual screening itself goes relatively fast. The full screening process is more time consuming when you follow up with the real screening, although if publicly available compounds are utilized, only assaying capabilities are required to verify the virtual hits. More extensive infrastructure, such as that available in larger companies, is typically required to further develop a compound once found. For a smaller company, or academia for that matter, to be able to play in this field, there needs to be cooperation and a way to manage the business end.
"Virtual screening is not the answer to all bottlenecks but it is another tool in the technology arsenal," Cameron concluded.
"HCV is an area in which Roche is highly interested," stated David Smith, a principal research scientist at Roche (www.roche.com). Currently, Roche has several products for HBV and HCV on the market: Pegasys (peginterferon alfa 2a, 40KD) for hepatitis B and C; Pegasys plus Copegus (peginterferon alfa-2a,40KD, plus ribavirin) for hepatitis C, which includes patients with normal ALT or co-infected with HIV; and Roferon (A-interferon alfa 2a) for hepatitis B and C.
"What is available now is effective in 80 percent of people who have genotypes 2 and 3 of the disease," said Smith. However, when you are looking at those who have genotype 1maybe 50 percent of all 170 million or so people infectedyou are only looking at about 40 percent effectiveness. We need to do better than that."
To that end, Roche is currently working on a polymerase inhibitor R1479 and its prodrug R1626. "Nucleosides are being shown to be a highly effective form of treatment," Smith said. "Using computational models, we targeted specific areas of the virus. We noticed that elongation occurs at the three-prime position, so we decided that the four-prime position was the area we wanted to target."
Roche took the prodrug R1626 into a Phase Ib, a two-week multiple ascending-dose study in HCV genotyope 1 patients. "We saw an increase in the amount of compound circulating and as a result, a clinically significant viral load reduction. We are encouraged by the results," commented George Hill, senior director of clinical research.
In this Phase I study, patients were randomized to receive either oral treatment with R1626 or placebo for 14 days with 14 days of follow-up. The preliminary data obtained from the 500-mg and 1,500-mg twice-daily doses were presented at the ൱st Annual Meeting of the European Association for the Study of the Liver (EASL)," held in April. The study found that at the 1,500-mg twice-daily dose, R1626 was associated with clinically significant reductions from baseline in serum HCV RNA (a measure of how much virus is in the blood) of 1.2 log10 (group mean). In addition, at both 500-mg and 1,500-mg twice-daily doses, R1626 was well-tolerated in patients with no serious adverse events and no premature withdrawals.
The study is ongoing and higher doses of R1626 are being evaluated. Future Phase II studies with R1626 in combination with Pegasys, with or without Copegus, are also planned. "We certainly need a better treatment option for our patients. The new oral polymerase inhibitor R1626 shows promising results and provides the opportunity of new combination therapy with Pegasys," Hill said.
"Eventually, we hope to be able to achieve a shorter treatment duration and better safety and efficacy."
The Proof is in the Concept
Currently, there are drugs on the market to treat HCV. However, they are said to be not very effective or well tolerated. The good news is that improved antiviral research is taking hold, as technology makes it possible to replicate the virus for study outside the body.
Novartis’ research efforts are focused on hepatitis C, and scientific approaches include well-known viral targets, such as the NS3 serine protease, as well as other Novartis-specific compound opportunities. Joanna Boerner presented a paper at ICAR on NIM811, an HCV replication inhibitor that exhibits potent antiviral activities by itself, or in concert with a nonnucleoside HCV polymerase inhibitor.
"One of the main viruses we work with is HCV. We are finding that NIM811 has good anti-HCV activity with favorable pharmacokinetics and safety profiles," said Boerner. "In vitro, our compound produces significant decreases in viral RNA levels. This effect is enhanced when NIM811 is used in combination with interferon or inhibitors of viral protease or polymerase. Furthermore, the combination of NIM811 with a viral inhibitor suppresses the emergence of resistant virus."
Boerner added that based on animal studies, therapeutic concentrations of NIM811 will be achieved in liver tissue where HCV replicates. "What we have in NIM811 is an effective antiviral for treatment of HCV infection."
Achillion Pharmaceuticals’ (www.achillion.com) lead candidate, GS 9132, also known as ACH-806, has demonstrated potent antiviral activity in both primary infection models and cell-based replicon assays across multiple genotypes. Genetic-resistance studies have demonstrated that the compound is compatible with HCV protease and polymerase inhibitors. The compound has demonstrated good oral bioavailability and safety pharmacology and is currently in a Phase I trial, which is scheduled to be completed imminently.
Mingjun Huang, senior director, antiviral drug discovery, presented a poster detailing work that Achillion did with NS5B polymerase nonnucleoside inhibitors (NNIs).
"NS5B is a primer target in searching for inhibitors of HCV replication. Although a variety of in vitro assays have been used successfully to identify NNIs, IC50 assays often differ with the recombinant NS5B form and the template and/or primer concentration used in the assays, raising concerns about the validity of the assays.
"In the study, we explored the replication complexes in vitro," said Huang. "In the poster it was shown that NNIs specifically block the synthesis of single-stranded HCV RNA catalyzed by the replication complexes in vitro although they have no inhibitory effect on synthesis of double-stranded HCV RNA."
According to Huang, the assay will be useful in guarding structure and activity relationship analysis during optimization of NNIs since it uses NS5B and template in its natural context and is, at the same time, independent of the cellular penetration and toxicity of the compounds.
Making Sense of the Genome
AVI BioPharma (www.avibio.com) presented on AVI-4065, an antisense approach to active HCV infection, looking at both preclinical and clinical evaluations. AVI’s antisense compounds are made up of repeating subunits linked together to form a polymer. This is referred to as the antisense backbone. Each antisense subunit carries a genetic letter that matches with its pair on the gene target. Although genetic letters are common to all antisense compounds, the chemical structure of the subunits and the linkages that string them together vary widely.
Early antisense backbones were made from natural genetic materials and linkages. These compounds were easily degraded or broken down by enzymes in the blood and within cells and had difficulty crossing cellular membranes to enter the cells that contained their genetic target.
"HCV is a virus composed of RNA, so what we do is make our antisense complementary to the virus," said Pat Iversen, senior vp of R&D. "Specificity is determined by dialing in the right numbers in the right arrangement. Since our compound is based on the same naming convention of the genes in the human genome, and since we have the sequence of the human genome, we can do initial toxicology studies in silico."
Finding compounds that fight HCV has been troublesome. "Problems with current therapy include short half-life and high rates of adverse events, and if a patient misses a dose, viral resistance emerges quickly," said Iversen. "Our preliminary results using AVI-4065 have been promising. We haven’t yet seen a serious drug-related adverse event and we are seeing a high degree of efficacy and a relatively long half-life. We have designed the drug so that it will be active across genotypes, which is very important, since the biggest gap right now is effective treatment for Genotype 1, which makes up approximately half of all HCV cases in North America and Europe.
"The results are preliminary," concluded Iversen, "but with the extraordinary safety profile and initial signs of efficacy that we have seen, it is promising."