Treatment of Helicobacter pylori infection 14‐day concomitant quadruple therapy versus triple therapy: A parallel double‐blind randomized controlled trial

Abstract Background and Aims Successful Helicobacter pylori (Hp) eradication with the traditional 7‐day course of proton pump inhibitor triple therapy is declining. Prolonging therapy to 14 days is associated with better eradication rates. Most learned societies recommend concomitant quadruple therapy (QC) as a first‐line alternative therapy for this bacterial infection. The aim of this study is to compare the efficacy and safety of triple therapy (TT) and QC for the eradication of Hp infection. Methods A parallel double‐blind randomized controlled trial was conducted. The diagnosis of Hp infection was made by pathological examination of gastric biopsies. Patients were randomly assigned to two treatment groups: either QC (esomeprazole 80 mg, amoxicillin 2000 mg, clarithromycin 1000 mg, and metronidazole 1000 mg daily) or triple therapy (esomeprazole 80 mg, amoxicillin 2000 mg, and clarithromycin 1000 mg daily in divided doses) for 14 days. The efficacy of the treatment is defined by Hp eradication attested by a negative breath test performed 6 weeks after the completion of treatment. Treatment outcomes were compared using the chi‐square test, while binary logistic regression identified predictors of treatment failure. Results Ninety‐two patients were included. Forty‐two patients belonged to the QC group and 50 to the TT group. No significant difference was noted between the two groups concerning the rate of Hp eradication either by intention to treat (81% vs. 72% respectively, p = 0.31) or per protocol (81.6% vs. 76.1% respectively, p = 0.54). Likewise, there was no difference between the two groups in terms of tolerance to treatment (59.5% for QC vs. 58% for TT, p = 0.88). No factor has been associated with treatment failure. Conclusion There was no significant difference in the rate of HP eradication between the QC and the 14‐day triple therapy. Neither regimen should be used topically because of their low eradication rates.


| INTRODUCTION
Helicobacter pylori (Hp) infection is one of the most common chronic bacterial infections worldwide.In Tunisia, the seroprevalence of Hp has been estimated at 64% among blood donors 1 and at 51.4% among children attending their first year of primary school. 2e involvement of Hp in the occurrence of several pathologies of the gastroduodenal mucosa, ranging from chronic gastritis to more serious conditions such as gastric adenocarcinoma and lymphoma of mucosa associated lymphoid tissue (MALT), 3 as well as in the pathogenesis of several extra-digestive pathologies, justifies the initiation of treatment when it is demonstrated.
[7] Given the increasing rates of resistance of Helicobacter pylori to triple therapy, the majority of learned societies around the world currently recommend quadruple therapy as a first-line treatment. 4,8e former Tunisian consensus on the treatment of Hp infection, in 2006, has not been updated.Thus, gastroenterologists tend to follow European recommendations by widely prescribing concomitant 14-day quadruple therapies.Given the high resistance to metronidazole in Tunisia, it is not possible to know if the same combination could be as satisfactory without metronidazole.Indeed, it is recommended, when using triple therapy, to extend the treatment to 14 days because prolonging its duration increases its effectiveness. 4However, there is no study in Tunisia that compares triple therapy to quadruple therapy.The objective of this study is to compare the efficacy and safety of two protocols in the first-line treatment of Hp infection in Tunisian adults: the 14-day clarithromycin-based triple therapy and the 14-day concomitant quadruple therapy (QC).This study includes male and female patients aged between 18 and 65 with a documented Hp infection.The diagnosis of Hp infection was made through an anatomopathological study. 9Gastric biopsies were taken according to the Sydney protocol: two fundics in one pot, two antrals, and one at the angle of the lesser curve in a second pot. 10 The biopsies were fixed in formalin and then sent to the pathology department of Fattouma Bourguiba Monastir Hospital where they were studied by an experienced anatomopathologist.Also, pregnant or breastfeeding women were not included.

| Context and study population
The included patients were randomly divided into two treatment groups according to a 1:1 ratio: the first group received concomitant quadruple therapy (QC) combining a double dose proton-pump inhibitors (PPI) (esomeprazole: 40 mg × 2 per day) with amoxicillin (1 g × 2 per day), metronidazole (500 mg × 2 per day), and clarithromycin (500 mg × 2 per day) for 14 days.The second group received triple therapy (TT) combining a double dose PPI (esomeprazole: 40 mg × 2 per day) with amoxicillin (1 g × 2 per day) and clarithromycin (500 mg × 2 per day) for 14 days.
The randomization was carried out through a platform that is available online (https://secure.dacimasoftware.net/medis/View/Login.aspx) and that allows the giving of a treatment box number to every randomized patient.The treatment boxes were prepared in advance by a doctor who had a table assigning each box number to its corresponding therapy (QC or TT).To preserve the double-blind nature of the study, two measures were essential: placing a placebo which has the same visual characteristics as metronidazole, in the boxes of the triple therapy and having a second doctor responsible for providing and explaining the treatment to patients.
The main aim of the study was to assess the efficacy of the two treatment regimens.To achieve this goal, a breath Test was performed with a minimum time period of 6 weeks after the end of treatment.The negativity of the breath Test signals the success of the eradication.
To assess the safety of used therapies, adverse events have been carefully researched.The tolerance of the treatment is judged on the absence of adverse events or the occurrence of mild adverse events that do not affect compliance with the treatment.
To assess treatment compliance, patients were instructed to return all unused medications.A pill intake of more than 90% was considered good compliance with the treatment. 11

| Course of the study
After approval, the patient was informed of the progress of the study and of the need to attend scheduled visits.Figure 1 illustrates the schematization of the course of the study.

| Sample size
The sample size was calculated using a sample size calculator (BiostatTGV).A dropout rate of 20% was considered, and the sample size was chosen to detect a 20% difference in proportions of participants who achieved Hp eradication in both treatment arms using the Pearson's chi-squared test assuming Hp eradication rate of 70% among triple therapy group 12 versus 90% among quadruple therapy group. 4These calculations were made based on the assumption that study power (1 − β) is 80%, and type I error (alpha) probability is 0.05.One-sided binomial proportion test was used, so we obtained an estimated minimum sample size of 92 participants.
We estimated the rate of loss of follow-up at 20%, which makes the number of participants to be recruited at around 111.The analytical study included the intention-to-treat (ITT) analysis which consists of analyzing all the patients in the group where they were randomized and the per-protocol analysis (PP), which consists of analyzing a subgroup of the population, including only patients in full compliance with the protocol For the comparison of the percentages, the Pearson's χ 2 test was used or the Fisher test when appropriate.For the means, the Student's t-test was used.

| Statistical study
The significance level was set at p < 0.05 for all statistical tests.Variables significant at 20% in univariate analysis were introduced into binary logistic regression for a multivariate analysis.

| RESULTS
As shown in the flow diagram (Figure 2), a total of 121 patients were screened for Hp infection.Out of 100 patients, 50 and 50 were randomized into triple and concomitant therapies, respectively.Eight patients in the triple therapy group were lost to follow-up.Sociodemographic characteristics and lifestyle habits were comparable between the QC and TT groups (Table 1).
Endoscopic findings, dominated by erythematous gastropathy, were comparable between patients who received QC and those who received TT (Table 2).
In the intention to treat, the Hp eradication rate was 81% for the patients who received QC versus 72% for those who were treated with TT, with no significant difference between the two (Table 3).
In multivariate analysis, no predictor of Hp eradication failure, such as age, gender, smoking, overweight, and therapeutic compliance, was found in this study (Table 4).
The occurrence of adverse effects was noted in 59.5% of patients in the QC group versus 58% in the TT group, with no F I G U R E 1 Course of the study.significant difference between the two (p = 0.88).Treatment adherence was also comparable between the two QC and TT groups (90.5% vs. 92%, p = 0.36).

| DISCUSSION
Different countries around the world have conducted studies to estimate the rate of Hp eradication according to the recommended treatment protocol QC versus TT.The results of this work are summarized in Table 5.
[21] The data from our study, which showed that the eradication rate was statistically similar between TT and QC, are inconsistent with most other works that have shown that the efficacy of QC is better when compared to that of the TT.The differences between the results of previous studies and this one could be explained by the fact that the rate of antimicrobial resistance to metronidazole in Europe, which is 32%, 1 and in South Korea, 22 is significantly lower than that of Tunisia which is estimated at 51.3%. 5 The concordance between  the present results and the two Singaporean 13 and Taiwanese 15 studies supports this hypothesis because these countries belong to the Asian continent where antimicrobial resistance to metronidazole is high and close to that of the Tunisian population (44%). 6veral predictors of the efficacy of eradication treatment have been reported. 7,23Among them, three were the most reported: primary resistance of Hp to antibiotics, degree of treatment adherence, and duration of treatment.
The emergence of primary resistance is currently a preeminent and universal problem with dangerously high levels, mainly for clarithromycin, which is a pillar of anti-Hp therapy. 6,24,25In Tunisia, resistance rates are lower compared with neighboring countries 26 and relatively stable over time 5,27 These data are very probably falsely reassuring and should be interpreted with prudence because they were not updated and they included low effective.In addition, Tunisia was ranked among the six countries with the highest consumption of antibiotics in the world. 28erefore, although the study of sensitivity was not carried out in this study, the existence of high rates of primary resistance in Tunisia is very likely.This may explain the low rate of Hp eradication, clearly below 90%, observed in our study for the two protocols.
Finally, eradication regimens should be based on the best locally effective regimen, ideally using individual susceptibility testing, or community antibiotic susceptibility, or antibiotic consumption data and clinical outcome data.The agents available differ in different regions, and this, in part, dictates what regimens are possible. 21ong the host-related predictive factors of eradication failure, adherence is the most important. 8Recent studies suggest that adherence to prescribed treatment greater than 90% results in better eradication rates, regardless of the treatment prescribed.In this study, there was no statistically significant difference in the rate of Hp eradication between the concomitant quadruple therapy and the 14-day triple therapy.These eradication rates are well below 90%, which is the lower limit adopted by several guidelines as the optimal threshold for the effectiveness of anti-Hp cure.Neither regimen should be used topically because of a likely high rate of primary resistance to clarithromycin.

A 13 -
month, parallel double-blind, randomized controlled clinical trial was conducted from February 2019 to February 2020 in the hepatogastroenterology department of Fattouma Bourguiba Hospital in Monastir, Tunisia.
All patients have been informed in advance and have given informed consent to join the study.The study has excluded patients: − With cirrhosis.− With renal failure.− Having complicated peptic ulcer (stenosis or hemorrhage or perforation) in an acute phase.− Having severe psychiatric disorders.− Having history of gastric surgery.− Having already received an Hp eradication treatment.− Having received an antibiotic within the last 2 weeks.− Who are allergic to one of the antibiotics used in the anti-Hp cure.− Who are drug addicted.
Data were captured and analyzed using IBM Statistical Package for the Social Sciences (SPSS) version 21.0 software.The descriptive study involved a calculation of simple frequencies and relative frequencies (percentages) for the qualitative variables and a calculation of means and standard deviations for the quantitative variables or medians with interquartile ranges if the normality was not assumed (the normality was assessed by the Kolmogorov-Smirnov test).
This protocol was carried out in accordance with the Declaration of Helsinki on good clinical practice after informed and signed consent by all participants before their inclusion in the protocol.This project was exposed and approved by the Committee for the Protection of Persons of the University of Monastir-Tunisia on February 19, 2019 with the following registration number: TN2018-INT-INS-14.

F I G U R E 2
Flow diagram of the study.A total of 121 patients participated in the study, of which 100 were included in the analysis.ITT, intention to treat; PP, per-protocol.T A B L E 1 Demographic characteristics of patients in the two groups.
Helicobacter pylori eradication rate in the two groups according to the type of analysis.
Abbreviations: BMI, body mass index; M, mean; QC, concomitant quadruple therapy; SD, standard deviation; TT, triple therapy.T A B L E 2 Endoscopic characteristics of patients in the two groups.Abbreviations: QC, concomitant quadruple therapy; TT, triple therapy.
Predictors of Helicobacter pylori eradication failure in the two treatment protocols.Summary of results of the randomized controlled trials concerning the rate of Helicobacter pylori eradication according to the treatment protocol: Concomitant quadruple therapy (QC) versus triple therapy (TT).
− This is a single-center study with a limited number of patients.−Previous exposure of patients to antibiotics that may raise the problem of secondary resistance has not been studied.− The antibiotic sensitivity testing was not carried out because it was not available in the study center.T A B L E 4 T A B L E 5