Antiviral kinetics of tenofovir alafenamide and tenofovir disoproxil fumarate over 24 weeks in women of childbearing potential with chronic HBV.

BACKGROUND/
PURPOSE: Use of tenofovir disoproxil fumarate (TDF) improves patient outcomes in preventing mother-to-child transmission (pMTCT) of the hepatitis B virus (HBV) in mothers with chronic HBV and high viral loads. Given the lack of data for tenofovir alafenamide (TAF) in pMTCT, rates of early viral suppression with TAF and TDF were evaluated in women of childbearing potential (WOCBP) participating in 2 randomized, double-blind, Phase 3 studies in chronic HBV.
METHODS: In a patient subset meeting WOCBP criteria and with baseline HBV DNA >200,000 IU/mL, rates of viral suppression with TAF or TDF in achieving the target of HBV DNA <200,000 IU/mL at weeks 12 and 24 were assessed. Multivariate logistic regression was used to identify factors predictive of failure to suppress HBV DNA to the target level.
RESULTS: In 275 of 1298 (21%) patients meeting WOCBP criteria with high viral load, 93% and 96% had HBV DNA <200,000 IU/mL at weeks 12 and 24, respectively. Results for TAF (n = 194) vs TDF (n = 81) treatment were similar at weeks 12 and 24 (94% vs. 90% and 97% vs. 93%), respectively. High baseline HBV DNA level, genotype D infection, and prior interferon (week 24 only) were predictive of failure to achieve the target level. Both treatments were well tolerated with TAF showing less impact on renal and bone parameters.
CONCLUSIONS: In WOCBP with high VL, no differences were found between TAF and TDF in reducing HBV DNA to levels associated with lower transmission risk. These data support ongoing studies of TAF for pMTCT.

RCT Entities:   Population Interventions Outcomes

Introduction

Worldwide, up to 2 billion people worldwide have become infected with the hepatitis B virus (HBV), with 257 million individuals estimated to have chronic infection (i.e. hepatitis B surface antigen [HBsAg] seropositivity for >6 months) [1, 2]. Although a safe and effective vaccine has been available for many years, vertical transmission of HBV from mother to child still occurs, accounting for approximately 50% of new infections in highly endemic areas and one-third of new cases in areas of lower prevalence [3, 4].

Passive-active infant immunoprophylaxis with hepatitis B immune globulin (HBIG) and the hepatitis B vaccine has greatly reduced the risk of viral transmission from mother to child; however, failures still happen with this approach, particularly when maternal viral load is high at time of delivery [4-7]. In fact, a serum HBV DNA level of 200,000 IU/mL (2 × 105 IU/mL) or greater at delivery is identified as a major risk factor for vertical transmission of HBV [5-7]. Notably, in resource-poor settings where access to HBV DNA assays is unavailable, determination of HBeAg seropositivity can be used as an alternative means to determine eligibility for antiviral prophylaxis [8].

Several antiviral agents, notably lamivudine [9], telbivudine [9, 10], and tenofovir disoproxil fumarate [9, 11–13] have been shown to reduce rates of mother-to-child transmission (MTCT) in pregnant women with chronic HBV infection and high viral loads when given in addition to standard immunoprophylaxis. These findings have been confirmed in a recent systematic review and meta-analysis [14]. Of these, TDF is preferred for prevention of MTCT (pMTCT) in all treatment guidelines given its high potency, proven efficacy, and acceptable safety profile in controlled trials [4, 15–18]. For women with chronic active hepatitis B requiring treatment who are, or desire to become pregnant, TDF is the preferred choice by all major guidelines. For women with chronic HBV not meeting treatment criteria, recommendations for when to initiate TDF during pregnancy for prevention of HBV transmission, and what constitutes a high viral load, differs somewhat among guidelines. In pregnant women with HBV DNA >200,000 IU/mL, the American Association for the Study of Liver Diseases (AASLD) recommends TDF during the third trimester (weeks 28–32) with discontinuation at time of delivery or up to 4 weeks postpartum [15]. The European Association for the Study of the Liver (EASL) recommends TDF when HBV DNA is >200,000 IU/ml (or HBsAg >4 log10 IU/mL) starting at weeks 24–28 weeks of gestation and continuing for up to 12 weeks post-delivery [16]. The Asian Pacific Association for the Study of the Liver (APASL) recommends initiation of TDF (or telbivudine as an alternative) at 28–32 weeks of gestation if the mother’s HBV DNA level is >6-7 log10 IU/mL with continued treatment until birth [17], while the China algorithm for interrupting MTCT recommends TDF (or telbivudine) to be started at gestational week 24–28 in women with HBV DNA >2 x 106 log10 IU/mL and discontinuation of therapy at delivery [4]. The World Health Organization (WHO) recommends that pregnant women testing positive for HBV infection (HBsAg-positive) with an HBV DNA ≥ 5.3 log10 IU/mL (≥ 200,000 IU/mL) receive TDF prophylaxis from the 28th week of pregnancy until at least birth for pMTCT [18]. Taken collectively, currently available recommendations suggest use of TDF when maternal HBV DNA is 200,000 IU/ml (2 x 105 IU/mL) or greater, with treatment continuing for 12 to 24 weeks depending on timing of the infant birth and whether treatment is continued post-partum.

Tenofovir alafenamide, a novel prodrug of tenofovir, has demonstrated antiviral efficacy non-inferior to that of TDF in Phase 3 studies in chronic HBV patients with elevated HBV DNA and alanine aminotransferase (ALT) levels [19-21]. Because of its greater plasma stability, TAF is given in a lower dose than TDF (25 mg vs 300 mg) yet achieves high intracellular levels of the active form of tenofovir within hepatocytes, while plasma concentrations of tenofovir are approximately 90% lower than with TDF [22, 23]. The unique pharmacologic profile of TAF supports improved renal and bone safety when compared to TDF in comparative trials [19-21]. TAF was approved for treatment of chronic HBV in November 2016, by the US Food and Drug Administration, and is currently approved in over 70 countries worldwide. Although clinical data are limited, TAF has not shown adverse embryo-fetal effects in animals and would be predicted to have a safety profile comparable to that of TDF when used during pregnancy, given that these two prodrugs share the same active form [24]. During the clinical development of TAF, pregnant women were excluded from participation in clinical trials, and if pregnancy occurred, study treatment was discontinued. Several trials evaluating TAF for pMTCT of HBV are underway with results expected within a few years. Thus, to better understand the role for TAF in pMTCT, we performed an analysis of the kinetics of viral load reduction in women with chronic HBV and high baseline viral loads (>200,000 IU/mL) who were of child bearing potential (WOCBP) and treated with TAF or TDF in 2 Phase 3 studies [19, 20].

Methods

Study design and participants

This was a secondary analysis of patients enrolled in 2 prospective, randomized, double-blind, Phase 3 studies comparing TAF vs TDF in chronic HBV [19-21]. The original Phase 3 clinical trials were approved by all participating institutional review boards and ethics committees. One study (Study GS-US-320-0110; NCT0190471) enrolled a total of 873 HBeAg-positive patients [19], while the other (Study GS-US-320-0108; NCT01940341) included 425 HBeAg-negative patients, otherwise the studies were identical in design [20]. Detailed eligibility and exclusion criteria and design of these studies are available in the respective publications. In brief, males and nonpregnant females, 18 years of age or older with screening HBV DNA >20,000 IU/mL and ALT >60 U/L (males) or >38 (females) U/L, without cirrhosis or with compensated cirrhosis, and with estimated creatinine clearance (eGFR) >50 mL/min were eligible. Patients coinfected with hepatitis C, hepatitis D, or HIV, those with clinical or laboratory evidence of decompensated liver disease, or evidence of hepatocellular carcinoma were excluded.

WOCBP included females 18 to 49 years of age based on the World Health Organization definition of reproductive age [25] who were identified by investigators as being of childbearing potential, and without history of hysterectomy, bilateral oophorectomy, or ovarian failure. For assessment of viral kinetics during treatment, only WOCBP with HBV DNA levels ≥200,000 IU/mL (2 × 105 IU/mL) at baseline were included.

Endpoints

The primary effectiveness endpoints for this analysis were the proportions with HBV DNA <200,000 IU/mL at week 12 and week 24 by treatment assignment (TAF or TDF). Several secondary viral endpoints were explored including the proportions with viral decline to <20,000 and <29 IU/mL at weeks 12 and 24, the subsets within each group with baseline HBV DNA ≥ or <8 log10 IU/mL who achieved HBV DNA <200,000 IU/mL at weeks 12 and 24, and responses in HBeAg-positive vs HBeAg-negative WOCBP. Additionally, ALT normalization was assessed in patients with baseline ALT levels above upper limit of normal (ULN). Safety assessments included in the original study protocols were evaluated in WOCBP by reported adverse events, serious adverse events, and lab abnormalities through week 24. Other safety parameters were change in eGFR by the Cockcroft-Gault method, and percent change in bone mineral density (BMD) at week 24 by dual energy X-ray absorptiometry (DXA) scans performed at hip and lumbar spine.

Assessments

Blood samples were collected every 4 weeks and plasma HBV DNA was determined by COBAS Taqman HBV Test for Use with the High Pure System (Roche Molecular Diagnostics, Pleasanton, California, USA) having a lower limit of quantitation of 29 IU/mL. ALT normalization was assessed using ULN cutoffs for females by 2016 AASLD (≤19 U/L) and central laboratory (≤34 U/L) criteria.

Statistical analyses

Included in the statistical analyses were females that met WOCBP criteria with baseline HBV DNA levels ≥200,000 IU/mL who were randomized and had received at least 1 dose of study medication. Viral suppression and ALT normalization analyses were performed by modified intention-to-treat, missing equals failure. Differences in proportions with 95% confidence intervals (CI) were determined for the rates of viral suppression at the weeks 12 and 24 endpoints (HBV DNA <200,000, <20,000, and <29 IU/mL) by Mantel-Haenszel Tests adjusted by baseline viral load (≥ and < 8 log Evaluation of risk factors for failure to achieve the viral suppression target (<200,000 IU/mL at weeks 12 and 24) were determined by univariate (UV) and multivariate (MV) analyses. Variables included in the UV analysis are provided in S1 Table. Factors with a p-value <0.15 by UV analysis were included in the MV stepwise model. P-values for change in eGFR from baseline were determined from the 2-sided Wilcoxon ranked-sum test, and when comparing mean percent change in BMD, an ANOVA model was used which included treatment as a fixed effect. Only patients with non-missing data were included in eGFR and BMD treatment comparisons.

Results

Patient population

Of the total enrollment of 1298 patients across the 2 Phase 3 studies, 479 (37%) were female, and 275 (21%) patients met WOCBP criteria with HBV DNA >200,000 IU/mL. Of the 275 patients, 194 (71%) and 81 (29%) received TAF and TDF, respectively. At baseline the two groups of were generally well matched (), having a median age of 35 years, 81% Asian, and the majority (52%) were genotype C. Over 80% were HBeAg-positive, with a slightly higher proportion in the TDF vs TAF group. Mean HBV DNA levels were similar at baseline; 40% overall had a level ≥8 log10 IU/mL. A small percentage (<8%) had cirrhosis by history or Fibrotest score ≥0.75; nearly all had ALT >ULN by both criteria. Only a small proportion (<5%) had comorbidities of hypertension, cardiovascular disease, and/or diabetes mellitus.

Virologic responses at weeks 12 and 24

Overall, 93% (255/275) and 96% (264/275) of patients had HBV DNA <200,000 IU/mL at weeks 12 and 24, respectively. A slightly higher proportion of TAF- vs TDF-treated patients achieved this endpoint at week 12 (TAF 94% vs TDF 90%; difference in proportion [95% CI]: 2.1% [-5.7% to 9.8%]; p = 0.504) and week 24 (TAF 97% vs TDF 93%; difference in proportion 4.4% [-2.7% to 11.4%]; p = 0.105); but in general, the kinetics of viral suppression were similar between groups over this period (). Among patients with high baseline viral load (HBV DNA ≥8 log10 IU/mL), the proportion achieving HBV DNA <200,000 IU/mL was similar between treatments: 88% (65/74) with TAF and 86% (30/35) with TDF at week 12, and 96% (71/74) with TAF and 91% (32/35) with TDF treatment at week 24 ().

By week 12, 76% of patients overall (209/275) achieved HBV DNA < 20,000 IU/mL with similar response rates between treatments (TAF 76% vs TDF 75%; difference in proportion -1.9% [-12.9% to 9.1%]; p = 0.727); by week 24, 91% (251/275) achieved this endpoint with a higher response for TAF vs TDF (93% vs 86%; difference in proportion 4.7% [-4.0% to 13.5%]; p = 0.215) (Fig 1B). Full suppression rates (HBV DNA <29 IU/mL) were lowest overall and similar between groups: TAF 9% (18/194), TDF 9% (7/81) at week 12 (difference in proportion -0.6% [-8.7% to 7.5%]; p = 0.872), and TAF 42% (81/194), TDF 47% (38/81) at week 24 (difference in proportion -8.5% [-20.3% to 3.3%]; p = 0.162) (Fig 1C).

Fig 1

Rates of viral decline to <200,000 IU/mL, <20,000 IU/mL, and <29 IU/mL.

TAF, tenofovir alafenamide; TDF, tenofovir disoproxil fumarate.

Among HBeAg-positive patients, the target of HBV DNA <200,000 IU/mL was achieved in 92% (144/156) and 90% (64/71) of TAF and TDF patients at week 12, respectively; at week 24, 97% (151/156) and 93% (66/71) of TAF and TDF patients, respectively, met this endpoint (Fig 2C). In the smaller subset of HBeAg-negative patients (n = 48), nearly all TAF and TDF patients achieved the target level (98% at weeks 12 and 24) (Fig 2D).

Fig 2

Rates of viral decline to <200,000 IU/mL by select baseline characteristics.

Baseline Viral Load (A) ≥8 log10 IU/mL and (B) <8 log10 IU/mL, (C) HBeAg-positive and (D) HBeAg-negative patients. TAF, tenofovir alafenamide; TDF, tenofovir disoproxil fumarate.

UV analysis identified several factors meeting criteria for inclusion in the MV model (); however, higher baseline HBV DNA level and genotype D infection were the only predictors for early failure in achieving the target level (). Among patients with HBV DNA ≥8 log10 IU/mL at baseline, the week 12 response rate overall was 73% for those with genotype D infection compared to 94% for non-genotype D patients. At week 24, only prior interferon treatment was predictive of target failure ().

ALT normalization

Overall, ALT normalization was observed in 44% of patients at 12 weeks (TAF 45%, TDF 40%), and in 64% of patients at 24 weeks (TAF 65%, TDF 63%) by central lab criteria (). Rates of ALT normalization were lower and similar between treatments when assessed by the AASLD criteria ().

Safety

In the WOCBP population, similar proportions experienced at least 1 adverse event (TAF 71%, TDF 68%) over 24 weeks (). Most adverse events were mild or moderate in severity with no patients discontinuing study treatment for an advent event. The most common adverse events (≥6%) were nasopharyngitis, headache, upper respiratory tract infection, cough, upper abdominal pain, and dyspepsia.

Four patients in the TAF group (2%) experienced 1 serious adverse event each of anemia, appendicitis, pneumonia, and pyrexia; none of these events was treatment-related or required treatment interruption or discontinuation. The most commonly reported Grade 3 laboratory abnormalities included blood in the urine and elevations in serum ALT ().

Median eGFR remained stable over the treatment period in the TAF group, whereas declines were observed in patients receiving TAF (). In TDF-treated patients, median (Q1, Q3) eGFR declined at week 12 compared with an increase on TAF treatment (-6.0 [-13.2, 2.4] vs +1.8 [-7.6, 8.7] mL/min; p<0.001); however, at week 24, eGFR change was similar between groups. Mean (SD) percent changes in BMD from baseline showed greater decreases with TDF vs. TAF at spine (-2.33% [2.49] vs -0.38% [2.30]; p<0.001) and hip (-0.77% [1.98] vs -0.17% [1.75]; p = 0.017) at week 24.

Discussion

Antiviral therapy with TDF given during the third trimester of pregnancy has been shown to reduce the risk of HBV transmission from mother to infant in several controlled clinical trials [9, 11–13], and in a recent systematic review and meta-analysis [14]. Data regarding the efficacy and safety of TAF for pMTCT of HBV are currently lacking. In this secondary analysis of early viral kinetics in a well characterized cohort of WOCBP with chronic HBV and high baseline viral load, we found that treatment with TAF was similarly effective to TDF in reducing plasma HBV DNA to a threshold associated with minimal or low risk of mother-to-child transmission of the virus to infants who are also given passive-active immunoprophylaxis [12]. After 12 weeks of treatment with TAF or TDF, 90% or more had a reduction in plasma HBV DNA to <200,000 IU/mL, and by 24 weeks, an even higher proportion of patients achieved this goal. These results also suggest that at least 12 weeks treatment is optimal to ensure most pregnant women with high viral loads achieve the target HBV DNA level by the approximate time of delivery. This is of relevance given that certain guidelines for pMTCT of HBV recommend starting TDF later than week 28, and while 40 weeks is typical, the timing of delivery does vary with up to 12.5% of births occurring before gestation week 37 [26]. Our data are supportive of guidelines by EASL and China which recommend initiation of treatment earlier at gestational weeks 24–28 [4, 16]. Waiting to initiate treatment between gestational weeks 28–32, as per the APASL, AASLD, and recently issued WHO guidelines [15, 17, 18], might be suboptimal for certain women, particularly in the setting of very high HBV DNA levels and/or preterm delivery.

Several studies support the level of maternal viremia being a critical factor when considering the timing of antiviral therapy pMTCT of HBV. By multivariate analysis, we identified 2 factors—higher baseline HBV DNA level and infection with genotype D as predictors for failure to achieve the viral suppression target at 12 weeks. High baseline viral load is widely known to affect rates of HBV DNA suppression as previously reported for TDF and ETV in patients with chronic HBV [27, 28]. Although less is known about the impact of HBV genotype on kinetics of viral suppression, we have previously reported infection with genotype D is associated with a slower rate of viral decline with TAF and TDF treatment, although the reason for this finding is unclear [29]. There are many areas globally where genotype D infection is common and wherein perinatal transmission remains an important source of HBV acquisition (e.g. parts of Africa, the Indian subcontinent, and Southern Europe [30]).

The viral kinetic data from this analysis correlate well with published studies that evaluated the association between maternal HBV DNA decline and infant outcomes when TDF is used for pMTCT of HBV [11-13]. In a randomized, controlled study in HBeAg-positive pregnant women with high viral load, TDF treatment started at pregnancy week 30-32 significantly reduced the rate of viral transmission to infants compared with controls (i.e. same infant immunoprophylaxis regimen without antiviral use in mothers): 5% vs 18%, respectively; p = 0.007 [12]. Substantially more mothers given TDF achieved the suppression target of HBV DNA <200,000 IU/mL at delivery vs controls (68% vs 2%; p<0.001), while in TDF-treated mothers the proportion with HBV DNA > 200,000 IU/mL at delivery was 1.5-fold greater when baseline HBV DNA was higher (> vs ≤8 log10 IU/mL; 39% vs 25%; p = 0.19). In a randomized, controlled study which compared TDF to placebo for pMTCT of HBV (passive-active immunoprophylaxis regimen given to all infants), pregnant women randomized to TDF (median gestational age 28 weeks) showed no HBV transmissions compared to 2% (3 infants) with placebo (p = 0.12) [13]. At time of delivery, 12% of TDF-treated mothers had HBV DNA >200,000 IU/mL as compared with 90% in the placebo group. In a controlled study of TDF vs no TDF treatment along with infant passive-active immunoprophylaxis [11], of 62 mothers who received TDF (mean screening viral load 8.25 log10 IU/mL) starting at 30–32 weeks gestation, 61 (98%) had HBV DNA <6 log10 IU/mL at the time of delivery, with a good correlation (correlation coefficient 0.8098; p<0.0001) between duration of TDF treatment and log10 reduction in HBV DNA levels, while TDF treatment was associated with a lower risk of HBsAg seropositivity in infants at 6 months (1.54% vs 10.71%; p = 0.048).

In the present analysis, treatment with TAF or TDF was safe and well tolerated and no patients discontinued treatment for an adverse event. WOCBP who received TAF showed only minimal declines in estimated glomerular filtration rate compared to greater declines with TDF, particularly at 12 weeks. Those receiving TAF also showed smaller mean percent declines in hip and spine BMD at 24 weeks. Whether the better bone and renal safety profile we observed for TAF vs TDF in WOCBP is confirmed in clinical studies with TAF for pMTCT of HBV remains to be determined.

There are several limitations to the present analysis. This was a descriptive, secondary analysis of viral suppression rates and safety in a subset of WOCBP using data previously collected from 2 large Phase 3 studies for CHB [25]. In applying the WHO definition, it is possible some postmenopausal women may have been included, and conversely, some WOCBP not fitting the WHO definition, may have been excluded. Also, the median age of the women included in our analysis (35 years) may be higher than the age of most pregnant women in many countries. Further, the study population was drawn from studies of patients with chronic active HBV who met criteria to initiate oral antiviral treatment by all major guidelines. This study population differs from the population for antenatal antiviral prophylaxis to prevent mother-to-child transmission–untreated women who are HBeAg-positive in the immunotolerant or “chronic HBV infection” phase of chronic HBV [16]. Thus, given these differences, it is unclear whether the rates of viral suppression we observed here would directly apply to the population typically given antivirals for pMTCT of HBV. Lastly, the physiological state of pregnancy can alter pharmacokinetics of some drugs, including antivirals [30-32]. Studies in pregnant women monoinfected with HBV [30] or HIV [31] receiving TDF have reported approximately 20% lower tenofovir exposures, particularly during the third trimester. The impact of these small changes in tenofovir pharmacokinetics on therapeutic response in pMTCT is not expected to be important, while the impact of pregnancy on TAF pharmacokinetics is presently unknown.

In conclusion, in WOCBP with high viral load, treatment with TAF or TDF for 12 and 24 weeks showed similar effectiveness in reducing HBV DNA to levels associated with minimal viral transmission to infants who receive complete HBV passive-active immunoprophylaxis. Findings from this analysis will inform present and future studies evaluating TAF for the prevention of mother-to-child transmission of the hepatitis B virus.

Univariate analysis.

Predictors of HBV DNA ≥ 200,000 IU/mL at Week 12.

(DOCX)

Click here for additional data file.

Changes from baseline in estimated Glomerular Filtration Rate (eGFR) by the Cockcroft-Gault method.

SD, standard deviation; TAF, tenofovir alafenamide; TDF, tenofovir disoproxil fumarate. *p<0.05, †p<0.001.

(DOCX)

Click here for additional data file.

27 Jan 2021

PONE-D-20-22518

Antiviral Kinetics of Tenofovir Alafenamide and Tenofovir Disoproxil Fumarate over 24 Weeks in Women of Childbearing Potential with Chronic HBV

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Reviewer #1: PONE-D-20-22518: statistical review

SUMMARY. This is a secondary analysis of subjects with chronic hepatitis B, enrolled in two previous studies and respectively treated with tenofovir alafenamide (TAF) and tenofovir disoproxil fumarate (TDF). It compares the proportion of subjects who reach the viral supression target (HBV DNA <200,000 IU/mL) at weeks 12 and 24 of the follow-up period. Supplementary analyses are made by considering alternative viral supression targets in subgroups. In a second part of the study, logistic regression analysis is performed to detect significant risk factors for failure to achieve the viral suppression target. While I have nothing to say about the second part of the study (methods are correct and results are clearly described and appropriately interpreted), I have a couple of concerns about the first part of the analysis: see major issues 1 and 2 below.

MAJOR ISSUES

1) One main finding of the study is that TAF and TDF have a similar effect on viral suppression at weeks 12 and 24, as described at the beginning of page 11. However, it looks like the difference between proportions has not been tested (ot it has, but results have not been reported ...). A two-sample test for proportions can be performed to show that, separately at weeks 12 and 24, proportions are not significately different. Or, perhaps more elegantly, an ANOVA logistic model can be estimated to perform a simultaneous test on both the 12th and the 24th week.

2) Figures 1 and 2 display the cumulative proportion of subjects who reach the suppression target at several weeks. I wonder why we shoud limit our attention to these two weeks when we could compare the two curves as a whole. For example, taking the complementary cumulative proportions (e.g. 1-p_w, where p_w is the cumulative proportion at week w), we obtain two survival curves that can be compared by a nonparametric logrank test, without the need of specific parametric assumptions. The authors should either perform this analysis or, alternatively, explain why weeks 12 and 24 are the only relevant weeks in this study.

Reviewer #2: The manuscript entitled “Antiviral kinetics of TAF and TDF over 24 weeks in women of childbearing potential with chronic HBV” addresses an important topic in optimizing chronic HBV treatment among women of childbearing potential, as pregnant women were previously excluded from participation in clinical trials. The Introduction states that ‘several investigator-sponsored trials evaluating TAF for PMTCT of HBV are underway with results expected within a few years.” It is unclear to this reviewer why it takes several more years for these important trials to complete (given that the follow-up period for HBV is relatively short).

The present study provides reassuring data on early viral suppression in women of childbearing age and, as expected, TAF was well tolerated and showed less impact on renal and bone parameters than TDF. These findings are important and do support the ongoing clinical trials of TAF in pregnant women for HBV PMTCT. The reviewer notes that 4 coauthors are employed by the company (Gilead Sciences) and many coauthors have potential conflicts of interest; however, these potential conflicts have been transparently reported.

Following are several comments and suggestions for the authors:

(1) The authors mention current HBV care and treatment guidelines from the various professional liver societies around the world (with substantial variations noted in these regional guidelines). However, the authors fail to include the WHO HBV guidelines. On World Hepatitis Day (July 2020), WHO published updated guidelines for the PMTCT of HBV – which are highly relevant to the present article.

(2) Two systematic reviews prepared for the latest 2020 WHO HBV guidelines have appeared in Lancet Infectious Diseases (published online August 2020; in hard copy, current issue): Boucheron et al; and Funk et al. These 2 systematic reviews provide relevant summary data for the Introduction and Discussion sections of your paper.

(3) First sentence “Worldwide, over 2 billion are infected with HBV …” (note this doesn’t seem accurately worded; over 2 billion people have become infected at some time during their lifetime). Chronic HBV infections worldwide are estimated by WHO at 257 million persons (see World Hepatitis Report, WHO, Geneva, 2017). The cited paper by Polaris – your reference 2 - is a relative outlier (‘up to 292 million’) among several published HBV modeling papers from 4-5 different groups. Your reference 1 (Schweitzer et al) estimated the number of people living with HBV infection at 240 million. The WHO estimate was a consensus estimate (involving many of the same investigator groups) – this reviewer recommends using the WHO (consensus) estimate.

(4) Intro, page 6, last para: Could you mention anything on the price differential between TAF and TDF? In practice, this will be an important determinant of use.

(5) “Currently approved in over 70 countries worldwide, including China, Japan, S Korea, Taiwan, and Hong Kong.” This seems overly focused on one area of the world. How about Europe, Africa, S America? (or simply delete individual countries).

(6) Page 7, line 3: ‘…same active moiety”. It seems accurately worded in this context. Just keep in mind, many people - like this reviewer - may not be familiar with ‘moiety’.

(7) Page 7, line 5: “Investigator-sponsored trials …” - not sure I understand this. Do you mean “investigator-initiated”? Usually each trial has a financial sponsor (e.g., NIH, a company or a foundation).

(8) Page 8, line: “2 identically designed, prospective, randomized trials…” However, we learn that one trial focused on HBeAg-positive patients and the other on HBeAg-negative patients. Therefore, I think ‘identically designed’ is incorrect.

(9) The study design included women of childbearing potential as ‘females 18-49 years of age’ – with a median age of 35 years. Thus, the average age in your study is higher than the average age of pregnant women in most countries (around 25-26 years of age) though this is changing. Not sure if this 10-year age difference might change any of the findings – I suggest including it among the study limitations.

(10) Discussion: page 13, line 12-13: Does your study provide any data on HBV viral load suppression ‘less than 12 weeks’ after initiation of HBV treatment (either TAF or TDF)? Such data would be very helpful.

(11) Discussion, end of para 1: Again, it would be helpful to include the updated 2020 WHO HBV guidelines in your Discussion. In many countries (e.g., in Africa, Latin America, many countries in Asia outside China), these would be the ones used.

Overall, this is an important paper and the findings among women of childbearing potential appear quite unique in the HBV literature.

Here are a few minor issues:

1) Reference 3 lacks information on year and pages.

2) Table headings need more information: for example, they should include information about the study population/year of the study cohorts included. Currently, the table headings cannot be interpreted or ‘stand on their own’.

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24 Mar 2021

Manuscript ID PONE-D-20-22518: Responses to Editor and Peer Reviewer Comments

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We have included captions for supporting information at the end of the manuscript as instructed.

REVIEWER COMMENTS:

Reviewer #1: PONE-D-20-22518: statistical review

SUMMARY. This is a secondary analysis of subjects with chronic hepatitis B, enrolled in two previous studies and respectively treated with tenofovir alafenamide (TAF) and tenofovir disoproxil fumarate (TDF). It compares the proportion of subjects who reach the viral suppression target (HBV DNA <200,000 IU/mL) at weeks 12 and 24 of the follow-up period. Supplementary analyses are made by considering alternative viral suppression targets in subgroups. In a second part of the study, logistic regression analysis is performed to detect significant risk factors for failure to achieve the viral suppression target. While I have nothing to say about the second part of the study (methods are correct and results are clearly described and appropriately interpreted), I have a couple of concerns about the first part of the analysis: see major issues 1 and 2 below.

MAJOR ISSUES

1) One main finding of the study is that TAF and TDF have a similar effect on viral suppression at weeks 12 and 24, as described at the beginning of page 11. However, it looks like the difference between proportions has not been tested (or it has, but results have not been reported...). A two-sample test for proportions can be performed to show that, separately at weeks 12 and 24, proportions are not significantly different. Or, perhaps more elegantly, an ANOVA logistic model can be estimated to perform a simultaneous test on both the 12th and the 24th week.

Author’s Reply

We acknowledge that the differences in proportions (95% confidence intervals and p values) between the treatment groups were not computed, given the numerical similarity in responses. However, as requested, we have now performed a statistical comparison of treatment responses (TAF vs TDF) for both week 12 and week 24 endpoints (i.e. for % with HBV DNA <200,000, <20,000, and <29 IU/mL). Additional text describing the statistical methods employed has been added to the Methods on Page 9, and the results of the statistical analyses have been added to Results on Pages 12 and 13. There were no statistically significant differences seen in treatment responses for TAF vs TDF treatment at any of the 2 time points (i.e. weeks 12 and 24) or for any of the efficacy endpoints.

2) Figures 1 and 2 display the cumulative proportion of subjects who reach the suppression target at several weeks. I wonder why we should limit our attention to these two weeks when we could compare the two curves as a whole. For example, taking the complementary cumulative proportions (e.g. 1-p_w, where p_w is the cumulative proportion at week w), we obtain two survival curves that can be compared by a nonparametric logrank test, without the need of specific parametric assumptions. The authors should either perform this analysis or, alternatively, explain why weeks 12 and 24 are the only relevant weeks in this study.

Author’s Reply

For completeness and perspective, Figures 1 and 2 include comparative viral suppression rates over 48 weeks; however, as stated in the manuscript (and highlighted in the figures), the rates of suppression at weeks 12 and 24, which were derived from data generated from Studies GS-US-320-0110 (HBeAg-positive patients) and GS-US-320-0108 (HBeAg-negative patients), are considered most pertinent when applied to the setting of preventing mother-to-child transmission (MTCT) of HBV. Although current guidelines such as those from AASLD and APASL recommend 12 weeks of treatment starting at gestational weeks 28-32 for the prevention of MTCT in highly viremic mothers, the optimal duration of TDF (or TAF) treatment is largely unknown, and as pointed out in the manuscript, there are data from some studies to suggest 12 weeks of TDF treatment before delivery may not be sufficient for some patients (i.e. those with very high viral loads). In our analysis, by modeling the level of viremia reduction to <200,000 IU/mL at delivery, we aimed to investigate both a 12- and 24-week duration of TDF or TAF treatment to see whether a longer duration might represent a potential advantage for maternal viremia reduction. While the overall rates of viremia reduction to <200,000 IU/mL (Figure 1, Panel A), showed comparable results at weeks 12 and 24, we did demonstrate that for women of child bearing potential with baseline HBV DNA at or above 8 log10 IU/mL, higher proportions in both treatment groups achieved this end point at week 24 compared with week 12.

Reviewer #2: The manuscript entitled “Antiviral kinetics of TAF and TDF over 24 weeks in women of childbearing potential with chronic HBV” addresses an important topic in optimizing chronic HBV treatment among women of childbearing potential, as pregnant women were previously excluded from participation in clinical trials. The Introduction states that ‘several investigator-sponsored trials evaluating TAF for PMTCT of HBV are underway with results expected within a few years.” It is unclear to this reviewer why it takes several more years for these important trials to complete (given that the follow-up period for HBV is relatively short).

The present study provides reassuring data on early viral suppression in women of childbearing age and, as expected, TAF was well tolerated and showed less impact on renal and bone parameters than TDF. These findings are important and do support the ongoing clinical trials of TAF in pregnant women for HBV PMTCT. The reviewer notes that 4 coauthors are employed by the company (Gilead Sciences) and many coauthors have potential conflicts of interest; however, these potential conflicts have been transparently reported.

Following are several comments and suggestions for the authors:

(1) The authors mention current HBV care and treatment guidelines from the various professional liver societies around the world (with substantial variations noted in these regional guidelines). However, the authors fail to include the WHO HBV guidelines. On World Hepatitis Day (July 2020), WHO published updated guidelines for the PMTCT of HBV – which are highly relevant to the present article.

Author’s Reply

The authors respectfully acknowledge that it was an oversight on our part not to cite or discuss the recently released and very pertinent WHO guidelines on antiviral prophylaxis for the prevention of mother-to-child transmission of HBV, and greatly appreciate the suggestion by the Reviewer to include this document in our manuscript. As our paper was submitted to PLOS One at a time coincident with the release of these guidelines last summer, this likely explains its omission. We have now included the WHO HBV PMTCT guidelines in the Introduction section as an additional citation (new Reference 18) on Page 5 in the sentence “Of these, TDF is preferred for prevention of MTCT (pMTCT) in all treatment guidelines given its high potency, proven efficacy, and acceptable safety profile in controlled trials [4, 9, 15-18].” We have also inserted content from the WHO Guidelines into the first paragraph of the Introduction on Page 6 as follows: “The World Health Organization (WHO) recommends that pregnant women testing positive for HBV infection (HBsAg-positive) with an HBV DNA ≥ 5.3 log10 IU/mL (≥ 200,000 IU/mL) receive TDF prophylaxis from the 28th week of pregnancy until at least birth for pMTCT [18].” Finally, reference to the WHO guidelines for pMTCT have also been added as text in the Discussion on Page 16 as follows: “ Waiting to initiate treatment between gestational weeks 28-32, as per the APASL, AASLD, and recently issued WHO guidelines [15, 17, 18], might be suboptimal for some women, particularly in the setting of very high HBV DNA levels and/or preterm delivery.”

(2) Two systematic reviews prepared for the latest 2020 WHO HBV guidelines have appeared in Lancet Infectious Diseases (published online August 2020; in hard copy, current issue): Boucheron et al; and Funk et al. These 2 systematic reviews provide relevant summary data for the Introduction and Discussion sections of your paper.

Author’s reply

The authors thank the Reviewer for this recommendation and have now added content from these 2 timely and pertinent systematic reviews to the manuscript. In the Introduction on Page 4, a sentence has been added to point out the recent publication by Boucheron et al regarding use of HBeAg testing to determine eligibility for antiviral prophylaxis (new Reference #8), and also in the introduction we have included text regarding the systematic review and meta-analysis by Funk et al (new reference 14) as well as in the inserted text in the first paragraph of the Discussion.

(3) First sentence “Worldwide, over 2 billion are infected with HBV …” (note this doesn’t seem accurately worded; over 2 billion people have become infected at some time during their lifetime). Chronic HBV infections worldwide are estimated by WHO at 257 million persons (see World Hepatitis Report, WHO, Geneva, 2017). The cited paper by Polaris – your reference 2 - is a relative outlier (‘up to 292 million’) among several published HBV modeling papers from 4-5 different groups. Your reference 1 (Schweitzer et al) estimated the number of people living with HBV infection at 240 million. The WHO estimate was a consensus estimate (involving many of the same investigator groups) – this reviewer recommends using the WHO (consensus) estimate.

Author’s Reply

The authors appreciate the clarification provided by the Reviewer and we have revised the wording in the first sentence of the Introduction accordingly. Also, it is appreciated that the citation of the Polaris data is considered by some to be an outlier. Therefore, at the suggestion of the Reviewer we have revised the estimated number to 257 million (the WHO figure) and replaced the Polaris citation with the WHO Global Hepatitis Report (Reference 2).

(4) Intro, page 6, last para: Could you mention anything on the price differential between TAF and TDF? In practice, this will be an important determinant of use.

Author’s Reply

While drug pricing is an important issue from a social and public health perspective, the authors respectfully decline the suggestion to include this in the manuscript as this is a secondary scientific analysis. Upon approval of TAF (Vemlidy) by the US FDA in 2016 for the treatment of chronic HBV its pricing has been on equal parity with that of TDF (Viread), so price has not been a key differentiator for their use. Finally, neither TDF nor TAF are currently indicated in their prescribing information for use in preventing mother-to-child transmission of HBV (rather the use of TDF is considered a public health measure driven by scientific data and treatment guidelines), therefore, any mention of drug pricing in the context of this paper we feel would not be appropriate.

(5) “Currently approved in over 70 countries worldwide, including China, Japan, S Korea, Taiwan, and Hong Kong.” This seems overly focused on one area of the world. How about Europe, Africa, S America? (or simply delete individual countries).

Author’s Reply

The Reviewer’s point is valid and as suggested we have now deleted any reference to specific countries wherein TAF is approved (Introduction, Page 6).

(6) Page 7, line 3: ‘…same active moiety”. It seems accurately worded in this context. Just keep in mind, many people - like this reviewer - may not be familiar with ‘moiety’.

Author’s Reply

We have replaced the words “active moiety” with the term “active form” and included more detail as to what this refers to – specifically the active intracellular form of tenofovir which is tenofovir diphosphate.

(7) Page 7, line 5: “Investigator-sponsored trials …” - not sure I understand this. Do you mean “investigator-initiated”? Usually each trial has a financial sponsor (e.g., NIH, a company or a foundation).

Author’s Reply

As this descriptor is not necessarily needed in this context, we have deleted the term “investigator-sponsored” from the sentence. The authors intent in using this term was to point out that for these particular trials, Gilead Sciences, the manufacturer of TAF and TDF, is not the Sponsor of the studies, rather they are being conducted by various investigators who serve as the study sponsor while Gilead provides unrestricted grant funding for the research to be conducted.

(8) Page 8, line: “2 identically designed, prospective, randomized trials…” However, we learn that one trial focused on HBeAg-positive patients and the other on HBeAg-negative patients. Therefore, I think ‘identically designed’ is incorrect.

Author’s Reply

The only differences between the 2 trials in terms of study design were the study populations included (HBeAg-positive patients in Study GS-US-320-0110 and HBeAg-negative patients in Study GS-US-320-0108), and by extension, the number of patients enrolled in each study due to the estimated sample sizes required; however, we can appreciate that technically in this sense, the designs were not truly identical. Thus, the term “identically designed...” has been deleted from the sentence. Instead, we have added new text to the end of the following sentence stating the following point “otherwise the 2 studies were identical in design.”

(9) The study design included women of childbearing potential as ‘females 18-49 years of age’ – with a median age of 35 years. Thus, the average age in your study is higher than the average age of pregnant women in most countries (around 25-26 years of age) though this is changing. Not sure if this 10-year age difference might change any of the findings – I suggest including it among the study limitations.

Author’s Reply

We agree the age difference could be considered an issue as suggested, although we do not have any data to suggest that viral load responses for TAF and TDF are influenced by age, particularly in younger, pre-menopausal women. However, it is acknowledged that this should be mentioned as a potential study limitation. Thus, we have added a sentence to the study limitations paragraph in the Discussion to make this point.

(10) Discussion: page 13, line 12-13: Does your study provide any data on HBV viral load suppression ‘less than 12 weeks’ after initiation of HBV treatment (either TAF or TDF)? Such data would be very helpful.

Author’s Reply

As shown in Figure 1, HBV DNA suppression was also assessed earlier than Week 12 – at Weeks 4 and 8. As expected, the suppression rates are lower following initiation of treatment than those seen at Weeks 12 and 24, and the figure does provide the reader with a frame of reference for these earlier suppression rates which we believe is sufficient.

(11) Discussion, end of para 1: Again, it would be helpful to include the updated 2020 WHO HBV guidelines in your Discussion. In many countries (e.g., in Africa, Latin America, many countries in Asia outside China), these would be the ones used.

Overall, this is an important paper and the findings among women of childbearing potential appear quite unique in the HBV literature.

Author’s Reply

As discussed earlier in our response to the Reviewer we have now included reference to the updated WHO HBV guidelines in the Discussion.

Here are a few minor issues:

1) Reference 3 lacks information on year and pages.

Author’s Reply

Thank you for pointing this out. The journal citation has been corrected based on information found in PubMed to: PLoS One 2017 Jun 2;12(6):e0178671. doi: 10.1371/journal.pone.0178671.

2) Table headings need more information: for example, they should include information about the study population/year of the study cohorts included. Currently, the table headings cannot be interpreted or ‘stand on their own’.

Author’s Reply

We have now added information to all table headings to indicate data sources.

Submitted filename: PLOSone Author Responses 03 19 21.docx

Click here for additional data file.

29 Apr 2021

Antiviral kinetics of tenofovir alafenamide and tenofovir disoproxil fumarate over 24 weeks in women of childbearing potential with chronic HBV

PONE-D-20-22518R1

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Reviewers' comments:

4 May 2021

PONE-D-20-22518R1

Antiviral kinetics of tenofovir alafenamide and tenofovir disoproxil fumarate over 24 weeks in women of childbearing potential with chronic HBV

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Table 1

Patient demographics and baseline characteristics in the subset of WOCBP (studies GS-US-320-0110 and GS-US-320-0108; week 96 integrated safety analysis set; reference 21).

	TAF (n = 194)	TDF (n = 81)	Total (N = 275)
Median (range) age, yr.	35 (18−49)	36 (18−48)	35 (18−49)
Race, n (%)
Asian	158 (81)	65 (80)	223 (81)
Black or African American	1 (<1)	0	1 (<1)
White	35 (18)	16 (20)	51 (19)
Mean (SD) BMI, kg/m2	23.0 (4.45)	22.5 (3.96)	22.9 (4.31)
BMI ≥30 kg/m2, n (%)	12 (6)	3 (4)	15 (5)
HBeAg positive, n (%)	156 (80)	71 (88)	227 (83)
Mean (SD) HBsAg level, log10 IU/mL	3.99 (0.66)	4.03 (0.63)	4.01 (0.65)
HBsAg >4 log10 IU/mL, n (%)	99 (51)	40 (49)	139 (51)
Mean (SD) HBV DNA, log10 IU/mL	7.6 (1.05)	7.6 (1.04)	7.6 (1.05)
Baseline HBV DNA, n (%)
<7 log10 IU/mL	51 (26)	23 (28)	74 (27)
≥7 to < 8 log10 IU/mL	69 (36)	23 (28)	92 (33)
≥8 log10 IU/mL	74 (38)	35 (43)	109 (40)
Median (Q1, Q3) ALT, U/L	77 (50, 111)	67 (53, 116)	74 (51, 113)
ALT, central laboratory
≤ULN	17 (9)	3 (4)	20 (7)
>ULN to 5 x ULN	151 (78)	65 (80)	216 (79)
>5 x ULN to 10 x ULN	21 (11)	9 (11)	30 (11)
>10 x ULN	5 (3)	4 (5)	9 (3)
ALT, AASLDa
≤ULN	1 (<1)	0	1 (<1)
>ULN to 5 x ULN	131 (68)	51 (63)	182 (66)
>5 x ULN to 10 x ULN	42 (22)	20 (25)	62 (23)
>10 x ULN	20 (10)	10 (12)	30 (11)
HBV genotype, n (%)
A	10 (5)	2 (2)	12 (4)
B	42 (22)	16 (20)	58 (21)
C	99 (51)	44 (54)	143 (52)
D	42 (22)	19 (23)	61 (22)
E	1 (<1)	0	1 (<1)
Cirrhosis history, n (%)
Yes	9 (7)	5 (7)	14 (7)
No	118 (93)	64 (93)	182 (93)
Indeterminate/unknown	67	12	79
Fibrotest score ≥0.75b, n (%)	4/185 (2)	1/79 (1)	5/264 (2)
Prior treatment experience, n (%)
Oral nucleos(t)ide	34 (18)	28 (35)	62 (23)
Interferons	23 (12)	11 (14)	34 (12)
Median (Q1, Q3) eGFR,c mL/min	107 (94, 126)	103 (89, 127)	106 (92, 127)
Hypertension, n (%)	7 (4)	5 (5)	11 (4)
Cardiovascular disease, n (%)	1 (<1)	1 (1)	2 (<1)
Diabetes mellitus, n (%)	6 (3)	4 (5)	10 (4)

aULN, 19 U/L for females and 30 U/L for males.

bSuggestive of Metavir fibrosis stage F4 (cirrhosis)

eGFR, estimated glomerular filtration rate using the Cockcroft-Gault method.

ALT, alanine aminotransferase; TAF, tenofovir alafenamide; TDF, tenofovir disoproxil fumarate, ULN, upper limit of normal range.

Table 2

Multivariate analysis: Predictors of HBV DNA ≥200,000 IU/mL.

	Week
	12	24
Baseline HBV DNA (log10 IU/mL)	5.8 (1.9, 17.6) p = 0.0021	-
HBV genotype D	4.3 (1.3,14.7) p = 0.0203	-
Prior interferon treatment	-	14.5 (1.3, 164.5) p = 0.0309

Values are reported as odds ratio (95% CI).

HBV, hepatitis B virus.

Table 3

ALT normalization at weeks 12 and 24 in the subset of WOCBP (studies GS-US-320-0110 and GS-US-320-0108, week 96 full analysis set; reference 21).

	TAF	TDF	Total
Central laba, n/n (%)
Week 12	80/177 (45)	31/78 (40)	111/255 (44)
Week 24	115/177 (65)	49/78 (63)	164/255 (64)
AASLD,b n/n (%)
Week 12	16/193 (8)	7/81 (9)	23/274 (8)
Week 24	40/193 (21)	18/81 (22)	58/274 (21)

AASLD, American Association for the Study of Liver Diseases; ALT, alanine aminotransferase; TAF, tenofovir alafenamide; TDF, tenofovir disoproxil fumarate.

aUpper limit of normal (ULN), 34 U/L for females.

ULN, 19 U/L for females.

Table 4

Adverse events through 24 weeks in the subset of WOCBP (studies GS-US-320-0110 and GS-US-320-0108; week 96 integrated safety analysis set; reference 21).

	TAF (n = 194)	TDF (n = 81)
Any adverse event	138 (71)	55 (68)
Any study drug−related adverse event	38 (20)	17 (21)
Any Grade 3 or 4 adverse eventa	3 (2)	1 (1)
Any serious adverse eventa	4 (2)	0
Any serious adverse event related to study treatment	0	0
Any adverse event leading to discontinuation of study treatment	0	0
Deaths, n	0	0
Common AEs (≥6% in either group)
Nasopharyngitis	24 (12)	4 (5)
Headache	16 (8)	7 (9)
Upper respiratory tract infection	13 (7)	4 (5)
Cough	12 (6)	3 (4)
Nausea	12 (6)	6 (7)
Abdominal pain (upper)	11 (6)	2 (2)
Dyspepsia	8 (4)	5 (6)
Lab Abnormalities (≥2% in either group)
Occult Blood, Grade 3	35 (18)	16 (20)
Urine Erythrocytes, Grade 3	28 (14)	19 (23)
Alanine Aminotransferase, >5 × ULN	18 (9)	7 (9)
Aspartate Aminotransferase, >5 × ULN	5 (3)	6 (7)
Creatine Kinase, ≥10 × ULN	5 (3)	0
Urine Glucose, Grade 3	3 (2)	0
Hemoglobin, 7 to <9 g/dL	3 (2)	2 (3)

Data are presented as n patients (%). AE, adverse event; TAF, tenofovir alafenamide; TDF, tenofovir disoproxil fumarate.

aNo reported Grade 3 or 4, or serious AEs were considered to be related to study drug.