Study Of Curative Effect And Mitochondrial Toxicity Of Telbivudine Treatment Within Chronic Hepatitis B Patients

Telbivudine（LDT) is the new type of sinistral nucleoside drugs which wasapproved for the treatment of chronic hepatitis B (CHB) in China in2007. The resultsof Ⅲ clinical trial showed that, the curative effect of telbivudine treatment for both1year and2years were significantly better than Lamivudine (LAM), on either HBeAgpositive patients or HBeAg negative patients. The#015data of this Ⅲ clinical trialwere obtained from China, the results showed that the HBV DNA undetection ratewas63%, HBeAg serological conversion rate was29%which was significantlyhigher than lamivudine, after2years treatment. As Telbivudine has just been used forclincal treatment in China in recent years, few medical evidences can indicateTelbivudine’s efficacy and safety of the long-term application in Chinese population,study focusing on the long-term treatment of telbivudine on chronic hepatitis Bpatients to figure out the curative effect, has great significance for the clinicalmedication in China.Although large-scale clinical trials had confirmed that, the anti-HBV effect oftelbivudine was great, but along with the increase in cases of clinical application,telbivudine related adverse reaction reports also gradually increased., such as themyopathy, toxic renal damage, peripheral neuropathy, hepatic steatosis, lactic acidosis,and so on, and the severe adverse reactions can lead to peripheral hypodermic adiposeatrophy, lactic acid poisoning and acute pancreatitis. A number of studies have shownthat the nucleoside drugs’ adverse reactions is associated with mitochondrial toxicity, and mitochondrial toxicity may appear in the early treatment period.In recent years, many investigations focused on the mitochondrial damage ofnucleoside reverse enzyme inhibitors (NRTIs),and the mitochondrial DNA (mtDNA)content in PBMCs was used as a biomarker of mitochondrial dysfunction, but thosestudies were mainly set up in HIV/AIDS population. Chronic hepatitis B patientsalways got antiviral treatment with single nucleoside drug and much less dosage,,comparing with HIV/AIDS patients. so the mitochondrial toxicity should not be much.So far, few studies had focused on the PBMCs mtDNA content in chronic hepatitis Bpatients treated with nucleoside analogues (acid),.One study had indicated that, the R964C mutation in mitochondrial DNApolymerase gamma (pol gamma) gene had responsibility to mitochondrial toxicity inpatients which accepted NRTIs treatment，potentially because that R964C mutationdecreased the activity of DNA polymerase gamma.Current genetic pharmacologyresearch also suggests that NRTIs toxicity may be associated with geneticsusceptibility, but frequency of the mutation and its potential impacts are not yetclear. Figuring out these genetic factors is critical for clarifying the mechanism ofNRTIs mitochondrial toxicity in human, and is also very important for developing thedrugs with high-efficiency and low-toxicity.The clinical effects and safety of LDT were observed by putting it into use on23CHB patients for104weeks. In order to explore the relationship between the level ofmtDNA before treatment and mitochondrial toxicity, and to make clear the predictivevalue of the decline in mtDNA copises, we detected the mtDNA copy number atdifferent periods in treatment and analyzed the R964C mutation of DNA polymerasegamma. Based on this study, we hope it can provide the theoretical foundation forindividual medication in future.Objectives:To investigate clinical efficacy and safety of Telbivudine treatment withinchronic hepatitis B patients which accepted104weeks treatment; To study thepossibility of mitochondrial toxicity caused by long-term Telbivudine treatmentwithin. chronic hepatitis B patients. Components:1. Detecting rate of Transaminase recovery, serum biomarkers of hepatitis B,clearance of HBV-DNA, fibrosis levels, before treatment and at different time pointsduring the treatment of, hepatitis B virus (HBV), and analyzing the results of livertisusse pathological detections before and after treatment, to evaluate the clinicalcurative effect of Telbivudine.2. Using the fluorescence PCR to detect the mitochondrial DNA level in PBMCsbefore treatment and at different time points during the treatment,,to explore themitochondrial damage caused by Telbivudine treatment; And detecting R964Cmutation of DNA polymerase gamma gene in those patients, to investigate therelationship between the R964C mutation and mitochondrial toxicity3. Observing the clinical adverse reactions and changes of lab indexes during thetreatment, to explore the safety of Telbivudine treatment.Methods:1. Human subjects: Chronic hepatitis B patients which accepted antiviral treatmentwith Telbivudine. The inclusion criteria included that: chronic hepatitis B wasdiagnosed according to diagnostic criterias from “the chronic hepatitis B preventionguide” which was confirmed by both Chinese medical association and the infectiousdiseases society in2010, age of subjects were from16to60years, and gender was notlimited.Exclusion criteria: subjects co-infected with HAV, HCV, HDV or HEV,subjects co-infected with HIV or had drug resistance, autoimmune or alcoholic liverdamage.2. Treatment: The patients administered Telbivudine600mg, oral, once every day,combined with adefovir (10mg, oral, once every day) if drug resistance or virologicalbreak emerged, the treatment went through104weeks.3. The test items:1) testing biochemical，virological and serological response,Fibrosis level at the baseline and at time points of antiviral24W、52W、76W and104W,2) liver pathological detection results before and after the treatment;3)Detecting mitochondrial DNA level in PBMCs at the baseline and at time points of antiviral24W、52W、76W and104W,4) detecting R964C mutation of the DNApolymerase gamma gene in patients.Results:1.Subjects general situation:23individuals including18males and5females wereinvolved in, average age was26.3±4.59years, the oldest one was35years old, andthe youngest one was19years old.2. Indexes of virology, serology and liver function: In CHB patients(n=23) treatedby Telbivudine, at24w treatment timepoint, the undetectable HBV DNA rate and ALTrecovery rate were34.8%and74.0%respectively,and the HBeAg elimination rate andHBeAg seroconversion rate were13.0%and0.0%respectively; At the52th week, theundetectable HBV DNA rate and ALT recovery rate were87.0%and91.3%respectively, HBeAg seroconversion rate is2.3%; At the76th week, the undetectableHBV DNA rate and ALT recovery rate were87.0%and91.3%respectively, HBeAgseroconversion rate is8.7%; While at the104th week, the undetectable HBV DNArate and ALT recovery rate were91.3%and100.0%respectively, HBeAgseroconversion rate is8.7%; and no patient was found with HBsAg elimination.3. Liver stiffness measurement(LSM): Statistical analysis result of total LSM datasindicated that LSM level decreased following the treatment (F=6.35, P=0.000). Thescore at baseline was higher than that at24W,52W,76W and104W timepoints,respectively,(P=0.000). But no significant difference was found between24w,52w,76w,104w.4. Pathological detection:1) Before treatment, the degree of liver inflammation ofthese patients were mainly in G2stage (11cases,49%) and G3stage (10cases,43%),the liver fibrosis degree were mainly in S2(14cases,61%) and S3(5cases,22%),2)After104ws treatment, the degree of liver inflammation were mainly in G1stage(13cases,58%) and G2stage (7cases,32%) and the liver fibrosis degree weremainly in stage S1(15cases,66%) and stage S2(,5cases,22%).5. Clinical adverse reactionsSo forth, no any myopathy, toxic renal injury, peripheral neuropathy, hepaticsteatosis, or lactic acidosis was found in those23patients. Serum creatinine (Cr) and uric acid (UA): No significant changes of Cr or UA wasfound during the treatment (P>0.05). During the process of trentment,there was nopatient has higher creatinine; At the treatment of24w,52w,76w,104w, there were5cases (21.7%) patients,2cases (8.7%),5cases (21.7%),6cases (26.1%) had elevatedblood uric acid, respectively.Creatine kinase (CK) results: At the treatment of0w,24w,52w,76w, and104w,patients with elevated creatine kinase were5cases (21.7%),13cases (56.5%),15cases (65.2%),15cases (65.2%),15cases (65.2%), respectively. Comparion betweeneach timepoint showed that the difference was statistically significant (F=3.56, P=0.009);0w CK was lower than24w,52w,76w,and104w, respectively,(P values were0.056,0.001,0.006,0.018,respectively). But no significant difference was foundbetween24w,52w,76w and104w, respectively (P>0.05).6. The result:(1) the mitochondria DNA testing results:20cases of healthy volunteers wereinvolved in this experiment as control group, the real-time PCR datas were analyzedby2CTmethod. The analysis results showed that on0W and24W,52W,76W and104W,the mtDNA relative expression level were1.16±1.37,1.79±2.23,2.33±2.31,2.28±2.94and1.96±2.09respecively, No significant mtDNA content decreasingwas found between24w,52w,76w and104w, respectively (P>0.05)(2) R964C mutation detection: sequencing result showed that no R964Cmutation was found in those23individuals.Conclusions：1. Through the clinical observation lasting for104weeks, LDT was indicated tohave a good antiviral effect, with a strong inhibition of HBV replication.The clinicalobservation of104weeks showed LDT has better antiviral effect,and strongerinhibition of HBV replicate.2. The possibility of adverse reactions by LDT is small during the first2years, so itis a relatively safe and effective antiviral drug.3. No obvious damage of mitochondria in PBMCs was found in CHB patientstreated by LDT in the first2years, we speculate that LDT is unlikely to lead tomitochondrial toxicity. 4. Because we have not found any mutations in the R964C of DNA polymerase gene and any patients with mitochondrial toxicity, so it is unable to figure out therelationship between the gene mutation and mitochondrial toxicity at present.