Effective Treatment of Lymphogranuloma venereum Proctitis With Azithromycin

Abstract

Background

Lymphogranuloma venereum (LGV) is a sexually transmitted infection caused by Chlamydia trachomatis (CT) serovars L1, L2, and L3 and is endemic among men who have sex with men (MSM) in Europe. We evaluated weekly oral azithromycin 1 g for 3 weeks as a treatment for LGV proctitis.

Methods

This is an open clinical trial with convenience allocation according to treating physician preferences. Adults with clinical proctitis received a single dose of 1 g of intramuscular ceftriaxone and were subsequently allocated to receive (i) doxycycline 100 mg twice daily for 21 days (Doxycycline group) or (ii) azithromycin 1 g orally once weekly for 3 weeks (Azithromycin group). LGV cure (primary endpoint) was defined as resolution of symptoms at week 6 (clinical cure, LGV-CC), with an additional supporting negative rectal polymerase chain reaction (PCR) at week 4 (microbiological cure, LGV-MC), if available.

Results

One hundred and twenty-five individuals with LGV clinical proctitis were included. All were MSM, and 96% were living with human immunodeficiency virus (HIV). Eighty-two were in the Azithromycin group, and 43 were in the Doxycycline group. LGV cure on a modified intention-to-treat analysis (primary endpoint), occurred in 80 of 82 (98%) in the Azithromycin group versus 41 of 43 (95%) in the Doxycycline group (treatment difference [95% confidence interval {CI}] 2.2% [−3.2, 13.2]). LGV-MC occurred in 70 of 72 (97%) vs 15 of 15 (100%) in the Azithromycin group and Doxycycline group, respectively (treatment difference [95% CI] −2.8% [−9.6; 17.7]). Adverse events were similar in both treatment groups.

Conclusions

Our findings support extended azithromycin dosing as an alternative treatment option for symptomatic LGV proctitis and provides the rationale for future randomized trials.

Lymphogranuloma venereum, proctitis, azithromycin, men who have sex with men (MSM), prospective observational trial

There is little well-conducted research published on the treatment of Lymphogranuloma venereum (LGV) proctitis. LGV has historically been an endemic sexually transmitted infection (STI) in Southeast Asia, the Caribbean, Latin America, and regions of Africa. Early in the twenty-first century it appeared in industrialized countries in the form of self-limiting epidemic outbreaks, particularly among men who have sex with men (MSM), and more recently it has become hyperendemic in that population. A recent meta-analysis [1] showed that the recommended first-line treatment with 100 mg of doxycycline twice daily for 21 days was highly effective for the treatment of symptomatic rectal infection. However, this treatment is challenging for patients due to the extended dosing schedule, frequent side effects such as nausea, in addition to being contraindicated in some populations such as pregnant and lactating women. It is clear that an alternative, simpler yet similarly effective treatment option would be of benefit to individuals with LGV proctitis and sexual health physicians.

Added Value of This Study

To date, the only prospective trial addressing LGV treatment dates from 1957 that included only 43 individuals and compared drugs that are currently not in use [2]. This current large prospective study demonstrates comparable efficacy, safety, and tolerability of an extended azithromycin regimen for LGV proctitis to standard treatment in HIV-infected MSM.

Implications of All Available Evidence

Extended dosing of azithromycin has potential as an alternative treatment for symptomatic rectal LGV infection. The findings of our study support further randomized clinical trials to confirm these results and provide supporting data for azithromycin as an alternative LGV treatment in the future STI guidelines.

BACKGROUND

Lymphogranuloma venereum (LGV) is an ulcerative and invasive sexually transmitted infection (STI) caused by Chlamydia trachomatis (CT) serovars L1, L2, and L3 [3]. It is endemic in tropical regions and, since 2003, has become hyperendemic among men who have sex with men (MSM) in western countries [4, 5]. Sexual behaviors such as unprotected anal intercourse, having numerous sexual partners, and sex under the influence of recreational drugs have been associated with this infection [6].

The most common clinical manifestation reported in MSM is an anorectal syndrome that is characterized by pain, tenesmus, bloody discharge, and inguinal lymphadenopathy.

Doxycycline 100 mg twice a day orally for 21 days is the first-line treatment for this infection [1, 3, 7–10] although the quality of evidence for this treatment is low. A small comparative nonrandomized treatment study in 43 subjects diagnosed with LGV was published in 1957 [2] and showed no significant differences in response to chloramphenicol, tetracyclines, or sulfadiazine. Later, various tetracycline formulations, such as doxycycline and minocycline, showed effectiveness in small, uncontrolled, clinical studies [11, 12]. As doxycycline has pharmacokinetics similar to those of tetracycline, a more convenient dosing, and lower toxicity, it was chosen and remains the recommended current first-line treatment. Erythromycin 500 mg 4 times daily orally for 21 days [3, 7, 8] is an alternative treatment regimen when doxycycline is contraindicated.

Major international [7–9, 13] guidelines suggest azithromycin treatment as an alternative regimen for LGV infections based on favorable pharmacokinetic studies. However, limited clinical data are currently available on the effectiveness of this antibiotic as alternative treatment for LGV [3, 12, 14–16].

The Hospital Clinic’s STI Unit protocol included the option of an extended azithromycin (EAz) dosing regimen (1 g orally once a week for 3 weeks) as an alternative treatment for rectal LGV. The main objective of this study was to compare the effectiveness of EAz versus the standard doxycycline regimen to treat symptomatic rectal LGV infection.

METHODS

Study Design and Subjects

This prospective non-randomized open clinical trial with convenience allocation according to treating physician preferences for the initial dose of empirical proctitis treatment was done in the STI Unit of a tertiary hospital in Barcelona, Spain. Subjects were recruited between 15 June 2010 and 30 June 2017.

We enrolled adults ≥18 years of age who presented with an anorectal syndrome that included at least one of the following symptoms: rectal pain, anorectal bleeding, constipation, inflammation, tenesmus, bloody or mucoid discharge with or without perianal ulcerative lesions, and/or local lymphadenopathy. All had a history of recent unprotected receptive anal intercourse. Only those with a positive rectal DNA-LGV swab result were included in this study (see Procedures and Figure 1). Study exclusion criteria included non-azithromycin or doxycycline use as empirical proctitis treatment, pregnancy or breastfeeding, allergy or intolerance to tetracyclines or macrolides, or receipt of an antibiotic with anti-chlamydial activity within 4 weeks of clinical presentation. Those individuals with a microbiological diagnosis of LGV infection but without clinical proctitis were not included in this study. Ethics approval was obtained from the local Research Ethics Committee (HCB/0875). The study adhered to ethical principles as set forth in the Declaration of Helsinki and followed all principles of good clinical practice. All individuals provided oral informed consent prior to their inclusion as described below.

Figure 1.

Trial profile. Abbreviations: CT, Chlamydia trachomatis; GN, Neisseria gonorrhea; LGV, Lymphogranuloma venereum; PCR, polymerase chain reaction; STI, sexually transmitted infection.

PROCEDURES

All individuals with clinical proctitis had, as part of the routine clinical workup, the following: (i) a questionnaire (visit 0; day −7) collecting epidemiological, clinical, and sexual behavior data (see Appendix 1); (ii) physical examination; and (iii) biological sample collection for a complete STI screening (see Appendix 2). According to local guidelines [8], subjects who consulted with clinical proctitis could be empirically treated with a single dose of 1 g intramuscular ceftriaxone plus either oral doxycycline 100 mg orally twice a day for 7 days or azithromycin 1 g single dose (AzSD) according to the physician preference (visit 0; day −7).

Seven days after empirical treatment initiation (study initiation: visit 1; day +1), individuals with positive rectal LGV-polymerase chain reaction (PCR) were provided standardized information about the different treatment options for LGV and, after giving their consent, were included in the study as follows: (i) subjects who were initially treated with doxycycline (Doxycycline group) maintained this drug to complete a 21-day course and were clinically assessed at day +14 (visit 3) and then to ensure the clinical cure (CC) (end of study: visit 5, day +35); and (ii) subjects who had been initially treated with azithromycin were offered continuing treatment with azithromycin (Azithromycin group) after agreeing to additional weekly follow-up visits for microbiological assessment by combined Neisseria gonorrhea (GN) and CT-PCR (LGV-PCR was performed when dual GN-CT test was positive for CT) (visit 1, day +1; visit 2, day +7; visit 3, day +14; visit 4, day +21). All subjects were requested to refrain from sexual contact until the end of the study (visit 5, day +35), which was done either face-to-face or by phone with a comprehensive questionnaire of the previously anorectal symptoms (Figure 1). Other nonrectal site STI coinfections (GN, syphilis, herpes simplex virus, Hemophilus, hepatitis C virus, and human immunodeficiency virus [HIV]) were treated according to the standard of care.

Outcome

The primary objective of the study was to assess the efficacy of Azithromycin group compared with Doxycycline group for the treatment of LGV proctitis. The primary efficacy endpoint (LGV cure) was defined as a sustained clinical response (clinical cure [CC]) and microbiological cure (MC), if performed. The MC or test of cure (TOC) was only recommended in those individuals not treated with the standard of care [7, 9, 17]. Regarding to the treatment of LGV-PCR missing data, a “not performed” test was considered negative if it was in the middle of 2 negative tests and positive if the next was positive. We defined CC as the sustained resolution of any defining symptoms of anorectal syndrome evaluated in a clinic appointment at least 6 weeks after the start of treatment. Microbiological cure was defined as a negative rectal LGV-PCR on/or before week 4 after the start of treatment plus no recurrence of symptoms for at least 6 weeks after the start of treatment. Secondary endpoints were rate of CC by on-treatment (OT) analysis; rate of MC; time to resolution of symptoms; time to negative LGV-PCR after Azithromycin-group treatment; and tolerance of treatments recorded at every follow-up visit by a questionnaire that included assessed of gastrointestinal symptoms, rash, and photosensitivity.

Statistical Analysis

The primary efficacy analysis was done on a modified intention-to-treat (mITT) population that included all individuals with confirmed LGV who came to their first clinical control 1 week after the administration of empirical treatment and agreed to participate in the study. For this mITT analysis, subjects lost to follow-up were considered nonresponders. In the OT analysis, we excluded those subjects who were lost to follow-up. Subjects were defined as lost to follow-up when no clinical or microbiological data were available 6 weeks after the LGV diagnosis.

The descriptive analysis was based on absolute frequencies and proportions for categorical variables and mean and standard deviation (SD) or median and interquartile range (IQR) for quantitative variables. The analysis of the contingency tables was carried out using the χ ² or the Fisher exact test. Student t test or Mann-Whitney test was used to compare quantitative variables between groups. The primary endpoint was presented as percent with the 95% confidence interval (CI) and with the risk difference CI based on Newcombe method 10 [18]. Due to the limited sample size and the very low number of nonrespondent subjects, no inferential analysis was considered. The Hodges-Lehmann median difference of time to clinical symptom resolution and its 95% CI was reported.

All tests were considered 2-tailed, and the level of significance was set at <.05. Data were collected in Excel files, and the statistical analyses were performed with Stata (Stata Release 15, statistical software 2015, StataCorp, College Station, Texas, USA).

RESULTS

Study Subjects

Out of 152 microbiological diagnoses of LGV during the study period, 136 individuals presented with clinical proctitis; the other 16 were asymptomatic or had LGV in another clinical location, and 125 of the 136 LGV proctitis fulfilled the inclusion criteria and were included in the study (study initiation, day +1). Eleven individuals who initially presented with LGV proctitis did not fulfill inclusion criteria: initial empirical treatment was modified by physicians outside the study in 6 individuals, and 5 individuals were initially treated outside the STI Unit (emergency department) with other alternative treatments: AzSD [3], levofloxacin [1], or minocycline [1].

All subjects were MSM with median age at the time of diagnosis of 38 years (IQR, 33, 44), and 120 (96%) were living with HIV. Baseline demographic data are presented in Table 1. Subjects presented a median of 21 days (IQR, 8–45) after onset of symptoms. Proctitis was reported as a unique form of presentation in 103 (76%) individuals; proctitis with a perianal ulcer in 22 (16%); proctitis with inguinal lymph node in 7 (5%); proctitis with anal canal mass in 3 (2%); and proctitis accompanied by a perianal ulcer and inguinal adenopathy in 1 subject. No differences in clinical presentation severity or baseline demographics were found across treatment study groups (Table 1).

Table 1.

Baseline Characteristics on the Participants and Subjects in the Modified Intent to Treat

Characteristics	All Subjects	Doxycycline Group	Azithromycin Group	P value
n	125	43	82
Age, years	40 (33–44)	38 (32–43)	40 (34–45)	.340
Male, no. (%)	125 (100%)	43 (100%)	82 (100%)
MSM	125 (100%)	43 (100%)	82 (100%)
Nationality (%)				.345
Spanish (%)	58 (47%)	23 (53%)	35 (43%)
Not Spanish (%)	66 (53%)	20 (47%)	46 (57%)
Median no. sexual partners, no. (IQR)	2 (1–3)	2 (1–3)	2 (1–3)	.983
In the previous 3 months	4 (1–10)	4 (1–10)	4 (2–10)	.436
In the previous 12 months	10 (4–40)		10 (4–35)	.543
No condom use	125 (100%)	43 (100%)	82 (100%)
HIV, no. (%)
Positive	120 (96%)	40 (93%)	80 (97%)	.338
UVL at the time of diagnosis	97 (82%)	30 (75%)	67 (85%)	.193
STI coinfections at the time of Dx
Any	35 (28%)	15 (33%)	20(25%)	.319
Prior non-STIs
Any	90 (70%)	31 (69%)	58 (70%)	.984
LGV presentation
Proctitis alone	90 (72%)	27 (63%)	63 (77%)	.141
Presence of inguinal lymph nodes	8 (6%)	3 (7%)	5 (6%)	1.000
Time to the onset of symptoms to Dx  Median (IQR) days	21 (8–45)	30 (15–60)	15 (7–34)	.016

n: Data obtained from this number of subjects.

Abbreviations: Dx, diagnosis; HIV, human immunodeficiency virus; IQR, interquartile range; LGV, Lymphogranuloma venereum; MSM, men who have sex with men; STI, sexually transmitted infection; UVL, undetectable viral load.

Eighty-two subjects received azithromycin (Azithromycin group), and 43 received doxycycline (Doxycycline group). No one who initiated AzSD treatment declined to continue the extended azithromycin regimen.

Efficacy

In the mITT population, for the primary efficacy endpoint, 80 of 82 individuals (98%; 95% CI: 91%, 100%) in the Azithromycin group versus 41 of 43 (95%; 95% CI: 84%, 99%) in the Doxycycline group achieved LGV-cure, with a treatment difference (95% CI) of 2.2% (−3.2, 13.2) (Table 2, Figure 2). In the second efficacy endpoint, LGV-cure by OT analysis, there was no loss of follow-up in either of 2 groups or subjects in whom treatment was changed because of intolerance or side effects, so according to the predefined criteria for the mITT and OT analysis, results were identical in both analyses. Microbiological cure was achieved in 70 of 72 individuals (97%; 95% CI: 90%, 100%) vs 15 of 15 (100%; 95% CI: 78%, 100%) in the Azithromycin group and Doxycycline groups, respectively (Table 2, Figure 2).

Table 2.

Primary and Secondary Outcomes

	Azithromycin Group	Doxycycline Group	Treatment Differences (95% CI)
Primary end point
LGV cure (mITT)	80 of 82 (98%)	41 of 43 (95%)	2.2% (−3.2, 13.2)a
Secondary end points
LGV cure (OT)	80 of 82 (98%)	41 of 43 (95%)	2.2% (−3.2, 13.2)a
LGV microbiological response (mITT)	70 of 72 (97%)	15 of 15 (100%)	−2.8% (−9.6, 17.7)a
Time to resolution of symptoms
Median time (IQR), days	8.0 (−5, 15)	9 (−5, 21)	1 (−5, 2)b
Tolerance
No. (%) adverse events	11 (13%)	6 (14%)	−.5% (−15.0, 11.1)

Abbreviations: CI, confidence interval; IQR, interquartile range; mITT, modified intention to treat; OT, on-treatment analysis.

^a95% CI using the Newcombe Method 10.

^b95% CI of the Hodges-Lehmann median difference.

Figure 2.

Difference in the efficacy end-points analysis at the end of the study between groups (Forrest Plot). Abbreviations: Dox, doxycycline; EAz, azithromycin; LGV, Lymphogranuloma venereum; MC, microbiological cure; mITT, modified intention to treat; OT: on-treatment analysis.

Two individuals from the Doxycycline group (LGV010 and LGV028) and 1 (LGV079) from the Azithromycin group were considered as failure of cure by clinical criteria. By microbiological criteria 2 individuals from the Azithromycin group (LGV032 and LGV079) were treatment failures (Appendix 3).

The median time to clinical symptom resolution in mITT was 8 days (IQR, 5–15) for those in the Azithromycin group and 9 days (IQR, 5–21) for individuals in the Doxycycline group; treatment difference was −1 (−5, 2) days.

Microbiological Results

Eighty-seven subjects were tested for MC. Seventy-two of 82 individuals in the Azithromycin group and 15 of 43 in the Doxycycline group were tested for LGV-PCR throughout the study. In the Azithromycin group, LGV-PCR was negative in 55% (12/22), 91.5% (22/24), 92.5% (25/27), and 96.5% (55/57), at days + 1, +7, +14 and + 21, respectively. According to the protocol analysis of LGV-PCR missing data, the results were as follows: LGV negatives: 52.1% (12/23), 90.3% (28/31), 95.3% (41/43), 97.2% (70/72) at days +1, +7, +14, and +21, respectively. In the Doxycycline group, LGV-PCR were performed in 2 individuals at day +1 and in 13 at day +21, and all were negative (see Appendix 4).

Safety

No severe adverse events (AEs) occurred in any group, and no subjects discontinued participation in the study because of an AE. AEs were mild and similar in both groups: 11 of 82 (13%) versus 6 of 43 (14%) in the Azithromycin group and Doxycycline group, respectively; treatment difference was −0.5% (−15.0, 11.1). Gastrointestinal symptoms and rash were the only drug-related AEs reported in both groups.

DISCUSSION

In this study, the efficacy and safety of azithromycin was similar to doxycycline for the treatment of symptomatic LGV proctitis. The 98% efficacy of LGV-cure (97% in the analysis by MC) in the Azithromycin group in our study cannot be compared to any other large series because, to date, the only data about the efficacy of any azithromycin regimen in symptomatic LGV proctitis is from case reports [5, 19] or very small series [15].

It is unclear if the 2 individuals in the Doxycycline group (Appendix 3) considered as treatment failure in LGV-cure analysis were true treatment failures as doxycycline was not administered under direct observation. However, our subjects confirmed perfect adherence with no missed doses. In addition, the absence of a LGV-PCR result after 21 days of doxycycline treatment and the absence in our study of a specific algorithm to classify treatment failure also made it difficult to distinguish treatment failure from reinfection [20]. Nevertheless, it is important to point out that several failures of a 21-day course of doxycycline in LGV infections have been reported. Although most failures were in individuals with severe inguinal lymph node involvement [21, 22], others have been also reported in individuals with only clinical proctitis. These treatment failures were not only clinical failures [21–23] but also microbiological [24–26]. One subject with a positive LGV-PCR at visit 4 (day +21) was restarted on treatment despite absence of clinical symptoms despite the possibility of a false positive result from remnant LGV DNA [27]. In contrast to DNA, Chlamydia RNA is only produced by the metabolic active bacteria and has a shorter half-life; it is more reliable to detect viable bacteria and treatment failures, but unfortunately that test was not available in our laboratory. A second subject had a reappearance of symptoms 3 weeks after the positive LGV-PCR on visit 4 (day +21) and was regarded as a true treatment failure, although this was in the context of having a sex partner who refused LGV testing and was untreated during the study (Appendix 3).

Similar to that reported in many other developed countries [26, 28, 29], our STI Unit has seen an increase in the number of cases of LGV proctitis over the last few years. It is important to note that in our STI Unit, from 2014 to 2017, more than half of the diagnosed proctitis (53%, 123 of 233) have been caused by LGV (Appendix 5), and as previously reported, we have seen a strong association with MSM [30]. The high proportion of LGV in the population living with HIV has been suggested to be due to an increase in susceptibility due to a weaker mucosal barrier and/or immunosuppression [31].

The rate of LGV proctitis associated in this study of 96% was higher than the 82% in the United Kingdom [28], or the 64% in the European Centre for Disease Prevention and Control report [30] This may be explained by proximity of our STI Unit to a popular MSM neighborhood in the center of the city of Barcelona, and the large cohort of MSM living with HIV who attend the Hospital Clinic.

The only difference in baseline characteristics between the Doxycycline group and the Azithromycin group was 30 days versus 15 days in the time from the onset of the symptoms at the initiation of treatment. This may be explained because in the first 3 years of the study there were more individuals who started doxycycline than azithromycin, and at that time, the suspicion of LGV in a patient with proctitis was lower than in the preceding years leading to a delay in the referral of subjects to the STI Units. It is important to point out that none of 28% of subject who were diagnosed with a concomitant sexually transmitted disease (STD) required any treatment with activity against CT.

Our results with an 88% (72/82) of MC (9 of the 10 individuals who refused the test of cure were followed clinically after the end of the study as part of the hospital HIV cohort, without clinical recurrence) in individuals who received azithromycin, along with a similar time for resolution of the clinical symptoms, would support the option of azithromycin as an alternative treatment for LGV proctitis.

To support the efficacy of azithromycin treatment for LGV, it is important to highlight the rate of LGV-DNA clearance in the Azithromycin group (Table 2). De Vries et al [32] reported in a study in 20 individuals with both symptomatic and asymptomatic LGV proctitis, a rate of clearance of Chlamydia DNA of 70.5% (12/17) at day +7 after starting doxycycline regimen and 89.5% (17/19) at day +14, which are similar to our results: 52% (12/23) and 90.3% (28/31) at days +7 and +14, respectively.

Although acceptability was not formally assessed in this study, the similar adverse event profile and adherence to medication between both treatment allocations suggests a similar profile. The long half-life of azithromycin (68 hours) that allows single weekly dosing in contrast to twice daily dosing of doxycycline is likely to promote acceptability and facilitate adherence in these subjects.

Despite interest in the use of azithromycin for LGV proctitis, doubts remain as to its efficacy [7, 9]. However, it is important to highlight that the main reason for this concern comes from several observational studies [33, 34] a systematic review, and meta-analysis that showed a higher rate of treatment failures in non-LGV CT rectal infections with a single dose of azithromycin compared to 100 mg of doxycycline twice daily for 7 days [35]. Another concern about the use of azithromycin if prescribed before having all the microbiological results is the possible presence of a coinfection with Mycoplasma genitalium (MG) [36]. If there is an undiagnosed coinfection with MG, the organism may be exposed to suboptimal macrolide concentrations, potentially selecting macrolide resistances [37]. Currently it is not known whether a single dose of azithromycin, considering its PK/PD characteristics, would be adequate to cure a macrolide sensitive MG or whether it is able to select resistance to MG in <3–7 days (the time needed to have the MG PCR results). This dilemma may be abrogated by point-of-care tests in the near future that avoid empirical treatment of STIs [38].

This study has some limitations. Although in the study prospective there was no formal randomization procedure, it was carried out at a single site, which limits the generalization of results. Furthermore, TOC was performed only 2 weeks after the last dose of azithromycin and with a DNA test that cannot exclude a false-positive result due to remnant trace DNA. The more recent version of the European guidelines on the management of LGV and other international guidelines advise to perform a TOC 4–6 weeks after the end of treatment [7, 10]. We did not identify the specific variant of LGV serovars in all cases (the few ones identified were all L2b); we cannot extrapolate or results to all different variants of LGV serovars [22, 29].

In conclusion, our findings suggest that extended azithromycin 1 g orally once weekly for 3 weeks can be a treatment option for LGV proctitis and supports its evaluation further in a larger phase 3 randomized clinical trial.

Notes

Potential conflicts of interests. J. L. B. has received honoraria for lectures or advisory boards and travel support and his institution research grants from Abbie, Bristol-Myers Squibb, MSD, ViiV Healthcare, Gilead, and Janssen. E. M. has received honoraria for lectures or advisory boards and travel support, and his institution research grants from Abbie, Bristol-Myers Squibb, MSD, ViiV Healthcare, Gilead, and Janssen. J. M. G. has received honoraria for lectures or advisory boards and travel support and his institution research grants from MSD, ViiV Healthcare, Gilead, and Janssen. Since May 2018 he is a full-time employee of ViiV Healthcare. All other authors report no potential conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.

Contributors. J. L. B. designed the study. J. L. B., I. F., and M. A. recruited subjects. J. L. B., I. F., M. A., A. B. A., J. M., and A. G. C. recruited data. E. D. L. and J. L. B. analyzed data. E. D. L., R. D. C., E. M., J. M. G., T. E., J. L. B., and I. F. interpreted data. J. L. B., I. F., R. D. C., J. M. G., and E. M. wrote the report. All authors had access to the clinical study report, reviewed, and approved the submission of the article.

Author notes

J.L. B. and I. F. contributed equally to this work.