Looming Hepatitis C Epidemic Sparks New Research

A virus unidentified until 1989 now threatens to outstrip HIV as a killer by threefold unless interventions emerge.

Around the world, an epidemic is quietly building that has the potential to dwarf AIDS in the number of people affected. The culprit, hepatitis C virus (HCV), is not as consistently lethal as HIV but nevertheless can kill. Development of a broadly useful vaccine seems unlikely, and the only therapeutic agents currently approved by the Food and Drug Administration (FDA) are recombinant interferons, which are ineffective in the majority of patients. However, biotechnology and pharmaceutical companies involved in developing HIV drugs are turning their efforts toward creating similar new compounds for HCV treatment. This bodes well for the career prospects of scientists with skills in antiviral drug research.

Photo: Christian Handel

BETTER YET: Schering-Plough's Greg Reyes believes that increased information about HCV will lead to new and better drugs.

The Atlanta-based Centers for Disease Control and Prevention (CDC) estimates that almost 4 million people in America are HCV-infected-quadruple the number of domestic HIV infections. The World Health Organization puts the global incidence of HCV at 170 million. Already, hepatitis C is responsible for 8,000 to 10,000 deaths annually in the United States, by liver disease or by cancer, according to CDC. Without effective intervention, mortalities are expected to triple within the next 10 to 20 years. By the turn of the century, HCV-related deaths in America will exceed those related to AIDS.

Current annual federal government funding for HCV research is only about $11.9 million, according to the Cedar Grove, N.J.-based American Liver Foundation (ALF). In comparison, federal funding for AIDS research in 1994 was $1.4 billion. Up to 14,000 calls monthly from the public concerning HCV are taken by ALF, which reports that the virus already is the leading reason for liver transplantation in the U.S.

BAD RAP: Many people with HCV who did not engage in high-risk behavior are worried about potential stigma, says ALF's Adrian DiBisceglie.

"This is sort of a sleeper of a disease. It usually takes 10 to 20 years before it causes damage," says Adrian DiBisceglie, medical director of ALF and a professor of internal medicine at the St. Louis University School of Medicine. "So, even though the incidence of hepatitis C infection in the United States is now declining, the peak of the epidemic is still to come."

Until HCV was cloned and sequenced in 1989 by researchers primarily funded by Emeryville, Calif.-based Chiron Corp., it was simply lumped in with "non-A, non-B" hepatitis types. The incidence of HCV infection from transfusion has become extremely low since 1989, but many people who received blood products or had transplants before then are still at risk of developing complications. Likewise, anyone who used injected drugs could be at risk. The virus also is thought to be transmissible through straws used to sniff cocaine. In addition, there is thought to be a risk, although relatively low, of transmission among those with multiple sexual partners.

ALF is worried that a pejorative label could be attached to HCV sufferers. "Many of our constituency don't know how they got hepatitis C, because they never injected drugs or sniffed cocaine or had a transfusion," DiBisceglie points out.

American Liver Foundation 1425 Pompton Ave. Cedar Grove, N.J. 07009 (800) 456-4837 http://www.liverfoundation.org Hoffmann-La Roche Inc. 340 Kingsland St. Nutley, N.J. 07110 (201) 235-5000 Fax: (201) 235-7034 http://www.roche.com Schering-Plough Corp. One Giralda Farms Madison, N.J. 07940-1010 (973) 822-7000 Fax: (973) 822-7447 http://www.sp-research.com ViroPharma Inc. 76 Great Valley Parkway Malvern, Pa. 19355 (610) 651-0200 Fax: (610) 651-0588 http://www.viropharma.com

Approximately 85 percent of individuals with acute HCV infection develop chronic disease, but until then, most carriers don't know they have the virus. In at least one-fifth of carriers, mild forms of hepatitis will progress to cirrhosis of the liver within two decades and often, only then do patients become symptomatic. In up to 20 percent of those who develop cirrhosis, hepatocellular carcinoma (HCC) will follow, which usually is fatal within months of diagnosis.

A Consensus Development Conference on Management of Hepatitis C was hosted in March by the National Institutes of Health. "There is an urgent need for effective antiviral therapeutics capable of inhibiting virus replication and stopping or delaying the progression of liver disease," the conference's consensus statement declared.

Currently, three recombinant forms of interferon, a natural antiviral glycoprotein, have been approved by FDA for HCV treatment in the U.S. These drugs are produced by Swiss multinational Hoffmann-La Roche Inc.; by Thousand Oaks, Calif.-based Amgen Inc.; and by the market leader, Schering-Plough Corp. of Madison, N.J. In September, Schering-Plough announced that two Phase III clinical studies had shown a tenfold increase in eradication of the detectable virus in patients treated with a combination of injected interferon and oral ribavirin, compared with those treated only with interferon. The trials of 345 patients indicated that six months after a 24-week course of treatment with the combination therapy, 48.6 percent of patients still did not have the virus, compared with viral eradication in only 4.7 percent of patients treated with interferon plus a placebo. The results, being prepared now for publication, prompted the company to announce that it plans to file a New Drug Application with FDA by the year's end.

Greg Reyes, Schering-Plough's vice president in charge of infectious disease and tumor research, notes that interferon and ribavirin are not therapies specific to hepatitis C alone. His implication is that more selective drugs remain to be developed. "We know what causes hepatitis C, we recognize various genes encoded and understand the pathology. We can now target a portion of the HCV genome for expression of genes, and we can set up high-speed assays."

The emerging targets for such new drugs arise from the fact that, like HIV, hepatitis C is an RNA virus; the core of its genetic material is RNA rather than DNA. Among major pharmaceutical companies attempting to discover and develop compounds that can inhibit RNA replication are Schering-Plough; Hoffmann-La Roche; Merck & Co. Inc. of Whitehouse Station, N.J.; Glaxo Inc. of Research Triangle Park, N.C.; and Eli Lilly and Co. of Indianapolis. A number of biotechnology companies also are in the hunt.

Spurred by successes with HIV protease inhibitors, companies are applying their knowledge and infrastructure to discovering and developing compounds that will inhibit HCV protease, an enzyme that breaks up polyprotein as a key step in genomic replication. Reyes says Schering-Plough is focusing on a particular nonstructural gene for viral-encoded protease.

Another potential target is the RNA helicase, an enzyme specifically encoded by the virus. During replication, the single-stranded RNA makes a mirror image of itself and becomes double-stranded. The helicase is responsible for unwinding this double strand into single strands that subsequently go on to yield new infectious viruses. A small-molecule drug that blocks RNA helicase activity could inhibit this process. Recently, a Schering-Plough study used X-ray crystallography to document the three-dimensional structure of the helicase (N. Yao et al., Nature Structural Biology, 4:463-7, 1997).

Reyes declares, "We can easily envision a combination of interferon and ribavirin with either an inhibitor of helicase or protease, or perhaps with some other compound that prohibits RNA replication."

Whaijen Soo, vice president of clinical sciences at Hoffmann-La Roche, says his company also is working on HCV protease inhibitors, but adds, "This proteinase [protease] is much more difficult to target than HIV, and that has taken researchers a little bit by surprise." A major mystery of HCV's enzymology is how it maintains itself in the host cell without the retroviral activity of HIV. The latter virus has enzymes that catalyze the formation of DNA, but the RNA of hepatitis C never integrates into the host cell's genome.

Another problem that has dogged viral disease drug therapy has been that antiviral agents are usually toxic not only to the pathogen but also to the normal intracellular system. "So, viral diseases pretty much eluded antiviral approaches, until we recognized proteinease as specific to the virus," Soo explains, "and we realized that if you could target that enzyme, your drug could be effective without necessarily being toxic to normal human cells." He foresees increasingly more focused HCV drug research industry-wide. "I think you'll see a shift in emphasis to follow emerging opportunities," he says, "changes because of available genomes and platform technologies."

Soo also is enthusiastic about current trials at Roche using a process called "pegylation." It involves introducing a bulky polyethylene glycol onto Roche's interferon molecule. The objective is to decrease administration of the drug from three times weekly to once per week. "The polyethylene glycol shields the interferon, so it can stay much longer in the bloodstream," he explains.

Just-completed Phase II trials also suggest that the new molecule is more potent than the regular interferon. Those data now will be confirmed with Phase III trials under preparation, along with trials of the new molecule on patients with cirrhosis, who usually are not responsive to interferon.

ViroPharma Inc., a publicly held biotechnology firm in Malvern, Pa., that specializes in RNA antiviral drug discovery and development for a variety of ailments, is working on RNA replication inhibitors for HCV. Marc Collett, vice president of discovery research, indicates that such drugs are highly specific and selective, and therefore likely to be safe. This is because the process of RNA replication is unique to RNA viruses; DNA viruses, retroviruses, and human cells do not replicate RNA. Moreover, hepatitis C has its own specific replication process.

ViroPharma's HCV research is being done in part with the Richmond, Conn., and Montreal facilities of the German company Boehringer Inghelheim Pharmaceuticals Inc. Several molecular targets have been defined, one of which is the RNA helicase. Attempts are now being made to achieve "chemical optimization" of inhibitors. "We don't believe what we're doing is competitive [with protease inhibitors], we believe it's complementary," says Collett. He explains that because of hepatitis C's viral diversity and the chronic nature of the disease, it is very likely to be better managed by a combination of two or more drugs than by a single agent. Moreover, drugs attacking multiple targets will be more effective than those attacking single targets.

Perhaps the most difficult challenge is to develop a broadly effective vaccine. Whereas therapeutics can directly interact with proteins encoded by relatively stable or conserved portions of the genome, a vaccine usually neutralizes antigens on the surface of the hepatitis C virus, which is covered with an "envelope" that contains the most genetically mutable portion of the genome. The wide genetic diversity of HCV is thought to be the main reason why the virus can escape the host's immune surveillance and cause chronic disease.

SHIFTY VIRUS: Hepatitis C is so mutable, it has a virtual continuum of genetic diversity, according to Robert Purcell of NIH.

HCV strains are divided into six major genotypes, provisionally classed as separate species. Within these genotypes are more than 100 subtypes and within them, the virus mutates into isolates that are specific to individual carriers, creating innumerable quasispecies or swarms of closely related but different viruses. Development of a broadly effective vaccine is "maybe impossible," according to Robert Purcell, head of the hepatitis section in NIH's National Institute of Allergy and Infectious Diseases (NIAID). However, he adds that serum pooled from a number of donors may contain a mixture of antibodies capable of neutralizing a broad array of HCV genetic sequences. Therefore, despite what he calls "a continuum of genetic diversity" in hepatitis C, "these sequences may fall into a manageable number of shapes" for a polyvalent vaccine to achieve cross-reactivity.

To test this idea, "we would like to try to map out the neutralizing epitopes a little better. To do that, we need better in vitro culturing, and we're working on that."

It is a major research stumbling block that no one has managed to cultivate fully infectious HCV in vitro. However, Purcell and others have produced a monoclonal or molecularly identical HCV in chimpanzees by introducing the virus in the liver. He now plans to monitor development of an HCV quasispecies in animals, observing where mutations occur, developing antibody markers, and characterizing the activity of the neutralization epitopes. The goal is to help give the private sector "a better idea of which way to go" in developing a vaccine.

ANOTHER WAY: Herbert Bonkovsky of the University of Massachusetts believes that reducing iron levels could help in HCV treatment.

There might be yet another alternative to combating HCV. Herbert Bonkovsky, director of the division of digestive disease and nutrition at the University of Massachusetts, believes that reduction of iron, an element required for replication of virtually all organisms, could be a useful treatment. His team conducted 12 studies involving 306 patients to confirm Japanese research that iron reduction by repeated phlebotomy or bloodletting can significantly improve the levels of liver enzymes. When these biochemical markers for hepatitis C are brought to a normal level, the virus clears in the majority of patients. Although iron reduction alone does not cause a decrease in viral levels, Bonkovsky's studies show that iron and interferon lead to significant improvement in biochemical and virological responses (H.L. Bonkovsky et al., Hepatology, in press).

Bonkovsky believes many HCV patients are trying folk remedies such as extracts of snap cucumber and of milk thistle. "My own impression is that it probably isn't making a big difference," he says, "but there hasn't been a real trial." Of HCV research efforts in general, he believes, "It's growing fairly substantially this year at the government level. My guess is, in terms of the pharmaceutical companies, the amounts spent for HIV research are many fold greater than for HCV. My impression, too, is that is changing."