Abstract
Adefovir, a nucleotide analog developed by John Martin, was a major breakthrough in the treatment of chronic Hepatitis B. Prior to adefovir, Hepatitis B treatment was limited to two therapeutic modalities, either interferon, which carried significant side effects and was efficacious in a minority of patients, or lamivudine which showed no durable effects with short-term use and a high rate of resistance with long-term use. Adefovir was found to be effective in suppressing viral replication and in resolving the hepatic inflammation associated with hepatitis B with only rare instances of resistance. In this article, we appreciate John Martin’s contribution to science and medicine as we review the landmark trials of adefovir that brought forth a new era of treatment of Hepatitis B.
Introduction
The goal of treatment of chronic hepatitis B is viral (HBV) suppression. The optimal result of treatment would be seroconversion or Hepatitis B Surface Antigen (HBsAg) loss. Viral suppression reduces damage to the liver which, if left untreated, could result in cirrhosis or hepatocellular carcinoma (HCC). Viral suppression offers the potential to improve overall survival rates by reducing comorbidities if left untreated. We can thank John Martin, a man known for being a brilliant scientist, manager, businessman, and CEO for bringing us the first nucleotide medication for the treatment of hepatitis B. Adefovir, a nucleotide analog, saved the lives of thousands of people who were resistant to lamivudine. Douglas Dieterich vividly remembers interacting with John Martin. “Whenever I saw him at the American Association of the Study Liver Diseases conferences or the European Association of the Study of Liver Disease conferences, he was studying science. You could always find John at the posters. Despite being the CEO of a global company, he was down to earth, a consummate scientist who was incredibly personable and driven to find solutions for patients ailing from viral diseases.”
There are seven medications currently approved by the FDA for treatment of hepatitis B [1,2]. The first medication approved for hepatitis treatment was interferon and shortly after, pegylated interferon, which allowed for easier patient use. Interferons work through immunodulatory and antiviral mechanisms to allow for the clearance of hepatitis B DNA and clearance of HBsAg in a significant minority of patients (3–8%). However, it is administered by subcutaneous injections, is only efficacious in a subset of patients (low viral loads, elevated ALT greater than 3 times the upper limit of normal, and high degree of histologic activity), and has significant side effects of flu like symptoms [1,2]. Nucleotide analogs were a promising new treatment as they had relatively mild side effects and easier oral administration (Figure 1). Timeline of the development of HBV treatment.
Lamivudine as HBV treatment
Lamivudine was the first nucleoside analog to be used against HBV, largely replacing the use of interferon since lamivudine has very few side effects, is orally administered, and acts rapidly. Lamivudine is an oral cytidine analog that causes termination of HBV DNA chain synthesis by inhibiting HBV DNA polymerase. Initial treatment with lamivudine showed a remarkable response with HBV DNA levels falling, transaminases normalizing, and liver histology improving compared with placebo [3]. Two significant issues arose with lamivudine. Those issues were centered around the discontinuation of lamivudine and prolonged daily use. Upon discontinuation of lamivudine, even after sustained viral suppression, the virus returned to its baseline levels of replication often accompanied by a dangerous alanine transaminase (ALT) flare. With prolonged daily use, lamivudine resistance was inevitable. In the first year of lamivudine treatment, close to a third of treated patients (32%) developed resistance [3] and by year four up to nearly three quarters of patients developed resistance (71%) [4].
Hepatitis B has a high mutation rate because its reverse transcriptase does not have a proofreading function and so is not able to remove erroneously inserted bases, leading to many mutations [5]. Resistance to lamivudine primarily occurs with the in rtM204V/I mutation where isoleucine or valine replaces methionine in codon 204 of the HBV DNA polymerase. This mutation changes the nucleoside binding site of the RNA-dependent DNA polymerase so that lamivudine no longer binds with the same affinity due to steric hindrance between the sulfur atom of lamivudine and the Beta-branched side chain in isoleucine or valine [6]. This alteration in the nucleoside binding site also affects binding of cytidine but not to the same degree. The viral load is initially lower with this resistance mutation than the wild type because the mutation decreases the virus's ability to replicate due to this decreased binding of cytidine [6–8]. However, with the additional mutations of rtL180M, the virus regains its replication competency by reshaping the catalytic site so that cytidine can bind with greater affinity [6,7] and viral loads increase along with transaminase levels [8,9].
In some instances, the resulting hepatitis flare caused by resistance leads to hepatic decompensation and death [9–11]. In a retrospective study, Chen et al. reported on a population of patients who developed biochemical breakthroughs due to M204V/I mutations while on lamivudine. They found that 4 months after biochemical breakthrough 56% of patients had acute exacerbations of hepatitis, 30% had hepatic decompensation, and six died. In this study, they compared a group of patients who stopped taking lamivudine at the onset of biochemical breakthrough to another group of patients who continued. They found that those who stopped lamivudine had more frequent exacerbations and higher ALTs during the hepatitis flares, but that there was no difference in hepatic decompensation or death between the two groups, though approximately half of the patients in the group that stopped lamivudine resumed it during a hepatitis flare in this retrospective study. They found that in the group that stopped lamivudine, the virus reverted to wild type causing the exacerbations. The viruses in the group that continued lamivudine developed a second mutation that led to increased fitness of the virus prior to the flares [10]. The authors conclude that additional agents are needed to rescue patients who develop recurrent hepatitis flares with lamivudine-resistant hepatitis B and suggest that adefovir could be that agent. Adefovir efficacy was not affected by the rtM204V/I mutation since adefovir does not contain the L nucleoside ring containing sulfur which is the cause of steric hindrance against lamivudine binding [7].
Adefovir
Summary of adefovir clinical trials.
Adefovir in HIV
The landmark placebo-controlled trial for adefovir in HIV compared the effects of adefovir 120 mg to placebo on HIV viral load and CD4 count. The study found that HIV RNA fell significantly in the adefovir group but not in the placebo group. Neither arm showed an improvement in the CD4 count at 24 weeks. The side effects of adefovir were hepatic and renal toxicity, with a significant elevation in transaminases and creatinine in those receiving adefovir compared with the placebo group. Many of the patients in this study were treatment experienced and had resistance mutations to zidovudine and lamivudine. This study found that adefovir was efficacious in the treatment of HIV even in those patients with resistance [12]. Due to significant renal toxicity seen in 33% of patients given adefovir 60 mg and 42% given adefovir 120 mg, adefovir was discontinued as HIV treatment in November of 2000. A later open label pilot study found that adefovir was efficacious in treating HBV in patients co-infected with HIV and lamivudine-resistant HBV. This 48-week open label trial gave HIV and lamivudine-resistant HBV coinfected patients adefovir 10 mg daily and found a reduction in HBV DNA for all patients while on adefovir. Utilizing a reduced dose of adefovir (10 mg daily), they did not find any signs of significant renal toxicity. Interestingly, there was no effect on the HIV for these patients at this lower dose of adefovir [13].
Adefovir in HBV
In 2003, the New England Journal of Medicine published two landmark trials on the efficacy of adefovir on Hepatitis B e Antigen (HBeAg)-negative and HBeAg-positive hepatitis B. The study for e antigen negative chronic hepatitis B, titled Adefovir Dipivoxil for the Treatment of Hepatitis B HBe Antigen–Negative Chronic Hepatitis B, randomized patients with chronic HBeAg-negative hepatitis B to adefovir 10 mg or placebo for 48 weeks. The results found histologic, virologic, and biochemical improvement. In the adefovir group, 64% of patients had a clinically significant improvement in their histology compared to 33% in the placebo group. In the adefovir group, there was a significant decrease in HBV DNA levels with 51% of the adefovir group having undetectable DNA levels compared to none in the placebo group. The median change in HBV DNA levels was approximately negative three log copies/mL by week 12. Finally, 72% in the adefovir group normalized their ALT compared to only 29% in the placebo group. They found no difference in adverse events between the two groups and no adefovir-resistance mutations in any patient at the end of the 48-week trial [14].
In the same issue, the New England Journal of Medicine published a trial by Marcellin et al. studying the efficacy of adefovir in HBeAg-positive chronic hepatitis B. This study randomized patients to three arms, adefovir 10 mg, adefovir 30 mg, and placebo. They similarly found histologic, virologic, and biochemical improvements in both of the adefovir arms when compared with placebo. Histologic improvement was found in 53% of patients on adefovir 10 mg, 59% on adefovir 30 mg, and 25% in placebo after 48 weeks of treatment. There was a significant decrease in the HBV DNA levels in both the adefovir 10 mg and adefovir 30 mg groups (3.52 log and 4.76 log median decrease, respectively) compared to placebo (0.55 log decrease) and a significantly higher number of patients in the adefovir group had undetectable HBV DNA by the end of the study period; 21% in the adefovir 10 mg group, 39% in the adefovir 30 mg group, and none in the placebo group had undetectable HBV DNA. A significantly higher proportion of patients in the adefovir groups normalized their ALT (48% in the 10 mg group, 55% in the 30 mg group) compared with placebo (16%). They did find an increase in nephrotoxicity, defined as an increase in creatinine greater than 0.5 mg/dL over baseline levels, in the adefovir 30 mg group. There was no difference in adverse events between the adefovir 10 mg and placebo groups. There were no adefovir-resistance mutations at the end of the 48-week study period [15].
The same year, an open label trial on the use of adefovir in pre- and post-transplant patients with chronic lamivudine-resistant hepatitis B was published. While the previous trials included those with compensated liver disease, this trial included patients with decompensated disease who were awaiting liver transplantation. While some patients in the previous trials were lamivudine experienced (∼8% in the HBeAg-negative trial, not reported in the HBeAg-positive trial), this trial specifically studied the effect of adefovir in lamivudine-resistant HBV. They found that adefovir 10 mg led to a reduction in the HBV DNA of approximately −3.3log and normalization of ALT in 76% of pre-transplant patients and 49% of post-transplant patients. Since this was an open label study, all post-transplant patients received adefovir 10 mg. Previously, HBV was treated with Hepatitis B immunoglobulin (HBIg) and lamivudine in post-transplant patients. There was still a relapse rate of 11–19% at 2 years with HBIg use and virologic breakthrough from lamivudine resistance was seen up to 45% of patients by 36 months. This study found that adefovir conferred a survival benefit to post-transplant patients with chronic hepatitis B compared with historical controls. No patients had developed adefovir resistance at 48 weeks [16].
Subsequent to this open label study, two randomized trials were published in Gastroenterology in 2004. The first, titled Adefovir dipivoxil added to ongoing lamivudine in chronic hepatitis B with YMDD mutant hepatitis B virus, added adefovir to lamivudine in patients with the M204V/I resistance mutation and followed them for 52 weeks. There were two arms of this study. In the first arm, patients were HBV HBeAg-positive and had compensated liver disease. These patients were randomized to receive adefovir 10 mg or placebo. They found that 85% of patients in the adefovir group had a clinically significant reduction in their HBV DNA compared to only 11% in the placebo group. They found a significant reduction of the ALT in the adefovir group, with a median change of ALT level of −48 IU/L in those taking adefovir compared to a median change of just −7.5 IU/L in the placebo group. 31% of patients normalized their ALT in the 52-week study period in the adefovir group compared to 6% in the placebo group. The authors reported no difference in adverse events between the two groups, specifically no evidence of renal toxicity in the adefovir group. The second arm included patients with decompensated liver disease or recurrence of HBV post-transplant. In this arm, all patients received adefovir open label and 92% had a significant reduction of HBV DNA from their baseline. They also found that 53% normalized their ALT and there were significant reductions in bilirubin and increase in albumin [17].
The second paper, titled Adefovir dipivoxil alone or in combination with lamivudine in patients with lamivudine-resistant chronic hepatitis B, was a multicenter study that randomized lamivudine-resistant chronic hepatitis B patients to one of three groups, adefovir plus lamivudine, lamivudine monotherapy, or adefovir monotherapy. They found that the HBV DNA decreased significantly and to a similar degree in the adefovir plus lamivudine group and the adefovir monotherapy group. They found no change from baseline levels of HBV DNA in the lamivudine monotherapy group. Similarly, they found that a similar proportion of patients normalized their ALT in the adefovir plus lamivudine group (53%) and adefovir monotherapy group (47%) but not in the lamivudine group (5%). They found no difference in adverse events across all groups [18].
Based on these studies, it was not known if patients should switch to adefovir once they develop lamivudine resistance or if they should continue lamivudine and add adefovir to the regimen. A 2007 randomized clinical trial examined this question. All patients had lamivudine-resistant HBeAg-negative chronic hepatitis B. Half were randomized to switch from lamivudine to adefovir 10 mg while the other half added adefovir 10 mg to their ongoing lamivudine treatment. The study followed the patients for 1 year and found that ALT and HBV DNA declined in both arms, but there was a significantly greater number of patients who had a relapse to abnormal ALT in the adefovir monotherapy arm. In line with this, they found that 21% of patients in the adefovir monotherapy arm developed adefovir resistance compared to none in the adefovir–lamivudine combination arm [19].
The following year Yatsuji et al. published a two-year follow-up on an open label study of treating lamivudine-resistant HBV by adding adefovir to lamivudine. They found that with combination therapy, 81% of patients had undetectable HBV DNA, 99% had normalization of their ALT, and 24% had HBeAg seroconversion at 24 months. They did find that 2 of the 129 patients developed adefovir resistance and breakthrough hepatitis. Adefovir resistance was associated with rt181 mutation also seen in lamivudine resistance. There did not appear to be significant adefovir resistance in combination therapy even at 2 years of follow-up [20].
Long-term adefovir data
Long-term adefovir data found that adefovir resistance, though much less common than lamivudine resistance, does occur. The long-term follow-up studies from the landmark adefovir in hepatitis B trials were published giving up to 4 years of follow-up data. The follow-up of the HBeAg-negative trial, titled Long-Term Therapy with Adefovir Dipivoxil for HBeAg-Negative Chronic Hepatitis B, published in the New England Journal in 2005 found that adefovir had good long-term efficacy and safety. In this trial, they randomized those who had received adefovir in the original trial to either continue adefovir or switch to placebo. The entire placebo group was switched to adefovir. They found that those who continued adefovir for the entire 144 weeks continued to have suppressed HBV DNA levels with a median reduction from baseline of 3.63 log copies/ml and a majority continued to have normal ALT. In those who were switched from placebo to adefovir, they unsurprisingly were found to have a reduction of their DNA levels to undetectable in 76% of patients and normalization of their ALT in 80% of patients in this group. Of those who were switched from adefovir to placebo, there was an increase in their viral load and in their ALT. Only 8% had undetectable HBV DNA at the end of the trial and 32% had normal ALT at week 96. It took a median of 8 weeks to see an increase in ALT to the pretreatment levels once these patients were switched from adefovir to placebo. None of them developed hepatic decompensation despite 32% of this group having significant elevations of ALT greater than 10 times the upper limit of normal. Importantly, they found no increase in creatinine in any patients who continued adefovir to week 144. Six patients in the continued adefovir group developed one of two resistance mutations, rtN236T and rtA181V. Four developed at week 96 and two developed at week 144. They found the rate of resistance to adefovir increased over time. The rate of resistance was 0% at week 48, 3% at week 96, and 5.9% at week 144 [21].
The follow-up data for the clinical trial of adefovir for treatment of HBeAg-positive hepatitis was reported in a 2008 paper in Hepatology titled Long-term efficacy and safety of adefovir dipivoxil for the treatment of hepatitis B e antigen–positive chronic hepatitis B. Unfortunately, the data is hard to interpret as the study drugs were misallocated in year 2 with significant crossover prompting the researchers to convert to an open label trial. They experienced significant attrition of subjects, and their long-term data is based on 64 patients who agreed to continue in the open label trial. However, within the limitations of this, they reported a continued decline in HBV DNA and continued normalization of ALT in these patients who continued adefovir. As this was an open label trial, there was no control group. Six of the patients developed renal toxicity defined as a sustained creatinine increase of greater than 0.5 mg/dL. Four were able to resume adefovir, but two had to permanently discontinue due to renal side effects. They found that 20% of their patients developed one of the same two previously reported resistance mutations, rtN236T and rtA181V. The viral load rebounded in 12 of 13 patients who developed these resistance mutations [22].
Two case reports of a total of five patients described an additional resistance mutation, rtI233V. This mutation was present prior to the initiation of adefovir in three patients [23] and arose de novo from adefovir selection pressure in an additional two patients [24]. All five patients with the rtI233V mutation responded to subsequent tenofovir therapy [23,24]. Two trials of entecavir for chronic HBV found a 2% rate of the rtI233V mutation and found that entecavir was effective in suppressing the viral load in patients with this mutation [25–27].
Discussion
The clinical effect of John’s discovery and approval of adefovir for hepatitis B and lamivudine-resistant hepatitis B cannot be overestimated. Lamivudine was approved for HIV in 1995 but not for hepatitis B until 2001. Most clinicians treating hepatitis B at the time were using lamivudine off-label for hepatitis B by the late 1990s since there really was no other treatment. Initially those patients did well, but as time went by nearly all of them became resistant to lamivudine and began to decompensate their liver disease. Many of these were Asian patients. Adding adefovir to the lamivudine absolutely saved their lives. It was approved in 2002. I still see patients that were decompensated cirrhotics in 2002 with lamivudine-resistant hepatitis B who are alive now in 2021 and in very good health due to John’s discovery of nucleotide analogs. One of the medical miracles that astounded clinicians then was how rapidly the liver recovered when we were able to treat the virus. I discussed this with John on many occasions and he was absolutely delighted to hear how well the patients were doing. Of course, this was only one nucleotide and there are several others that had similar effect on HIV and hepatitis B.
John Martin brought forth a new era of treatment for Hepatitis B patients. He was a man driven to find solutions for complex problems in viral disease. John advanced science that transformed the lives of large numbers of hepatitis B patients. He was always concerned about the patients. Thank you, John Martin. Thank you for helping us save the lives of thousands of patients.
Footnotes
Declaration of conflicting interests
The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.
Funding
The author(s) received no financial support for the research, authorship, and/or publication of this article.
