Bene t of Adjuvant Radiotherapy for Gallbladder Cancer: A Comparability-Based Metaanalysis


 Background & purpose: The benefits of adjuvant radiotherapy (ART) in gallbladder cancer (GBC) treatment remain inconclusive owing to the rarity of GBC and lack of randomized studies.Methods: PubMed, Medline, Embase, and Cochrane Library were systematically searched until March 2021. The primary endpoint was overall survival (OS). Comparative clinical studies that reported survival outcomes in GBC patients treated with or without ART were included. The comparability of each study was assessed by considering all possible clinical indicators (group 2: ART arm with poor clinical profile; group 1: ART arm with statistically similar profile or no evidence of having inferior clinical factors compared to non-ART arm).Results: Twenty-one studies involving 6876 GBC patients were reviewed. In pooled analyses of OS, the odds ratio (OR) was 1.26 ( p =0.111). In subgroup analyses considering comparability, the OR significantly favored the ART arm (1.92, p =0.008) among comparability group 1 studies, whereas it was 1.03 ( p =0.865) in comparability group 2 studies. The pooled rate of 5-yr OS in the ART vs. non-ART arms was 44.9% vs. 20.9% in group 1 and 34.1% vs. 40.0% in group 2. With ART, significant reduction in locoregional recurrence (OR: 0.21, p =0.001) but not in distant metastasis (OR: 1.32, p =0.332) was noted.Conclusion: ART not only showed benefits in patients with a similar clinical profile to those treated without ART but also yielded comparable survival in patients with an inferior clinical profile. Our results strongly support the more active application of ART in GBC treatment.Protocol registration: This study is registered in PROSPERO (CRD4202124062, available at: https://www.crd.york.ac.uk/).


Introduction
Gallbladder cancer (GBC) is an extremely aggressive malignancy of the biliary tract, and surgical resection remains the only potentially curative approach. However, only 10-30% of patients present with resectable disease, and high recurrence rates are reported even after curative resection [1][2][3][4]. Unfortunately, evidence supporting the role of adjuvant therapies in GBC is scarce, and adjuvant treatment is not yet considered standard practice. After one prospective randomized study showed survival benefits with adjuvant chemotherapy in 2002, [5] several phase II and retrospective studies have explored adjuvant strategies after resection of GBC. Owing to concerns about considerable locoregional recurrence, some researchers have attempted to investigate the benefits of adjuvant radiotherapy (ART).
However, because of selection bias and heterogeneity in studies, the role of ART remains unclear.
Two meta-analyses evaluating the clinical benefits of ART for GBC have been published [6,7].
Both studies provided supporting evidence for the survival benefits of ART, especially in the subgroup of patients with positive nodes or margins. However, these meta-analyses did not consider bias due to comparability issues but assumed that the ART arm had an inferior clinical profile, which may not always be true (e.g., ART could be omitted in patients with a short life expectancy). Furthermore, clinical prognosticators other than node positivity and positive margins were not considered, and subgroup analyses for these factors included only two to five studies.
Presently, established data from randomized studies are unavailable, and clinical decisions are made on the basis of findings from observational studies. Furthermore, with developments in radiotherapy techniques, which further reduced the obstacles to ART application, evidence supporting the application of ART for GBC in real clinical situations is needed. Therefore, this meta-analysis aimed to evaluate whether ART for GBC yields oncologic benefits. We attempted to enhance the study's reliability by comprehensively considering the comparability between arms on an individual study basis.

Study searching and inclusion
We adhered to the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines [8] for conducting this meta-analysis and referenced the Cochrane Handbook (version 6.2) for methodology [9]. The population, intervention, comparison, and outcome (PICO) question of the hypothesis was as follows: "Does ART confer a survival benefit in treating GBC?" The following inclusion criteria were used for recruiting studies: 1) comparative clinical studies involving GBC patient cohorts treated with and without ART, 2) studies with at least five GBC patients in each arm, and 3) studies that provided any measurement of overall survival (OS). We searched four databases (PubMed, Embase, Medline, and Cochrane Library), as recommended by the Cochrane Handbook [10], until March 2, 2021.
The search terms and strategies according to the databases are described in Supplementary Data 1. No language restriction was applied. Chinese articles were translated by a professional Chinese-English translator, and Japanese and Korean articles were translated and interpreted by one of the authors (CH Rim). Conference abstracts were included if they fulfilled the inclusion criteria. To reduce the risk of missed articles, the reference lists of the included articles and previous meta-analyses were reviewed. Studies from the same institution, with possible duplication, were filtered using the following criteria in order: 1) study providing target information and relevant clinical information, 2) study with a larger number of patients, and 3) more recent publication (when the number of patients was similar). Studies based on the National Cancer Database (NCDB) were also filtered using the same criteria when the possibility of duplication existed. All processes of study searching and inclusion were performed by two independent researchers; any disagreement was resolved through discussion and repeated searching.

Data collection processes and items
The primary endpoint was OS. Locoregional recurrence, distant metastases, grade ≥3 complications, and disease-free survival (DFS) were assessed as secondary endpoints. We used a standardized form to collect clinical and general information, including endpoints, as follows: 1) general information including author name, patient recruitment period, publication, institution where the study was performed, and conflicts of interest (commercial, academic, or none), and 2) clinical information including dose and modality of ART, application of concurrent chemotherapy, median OS, 3-and 5-yr OS, 3-and 5-yr DFS, grade ≥3 complications, and locoregional and distant recurrence rate. Data collection was performed by two independent researchers; any disagreement was resolved through a discussion based on a literature re-review.

Study quality and bias assessment
As most related studies were nonrandomized, possible confounders were carefully discussed, as recommended by the Cochrane Handbook. Considering the rarity of GBC, similarity of treatment modalities (surgery and/or ART), and reliability of the primary endpoint (OS), a meta-analysis might be feasible. However, according to our preliminary literature analysis, the most significant potential confounder was comparability. Most studies were not prospectively controlled; they retrospectively assessed the treatment outcomes. Previous meta-analyses assumed that ART was performed in patients with inferior clinical profiles. However, ART may have been omitted in patients with a short life expectancy. Additionally, some studies suggested that clinical indicators were similar between groups treated with and without ART, based on statistical analyses. Some meta-analyses attempted to resolve bias in comparability through subgroup analyses, according to clinical indicators such as lymph node (LN) metastases and resection margin (RM). Although these are known clinical indicators, few studies provided comparative data to allow such analyses, and clinical decisions and patient prognoses were also affected by other clinical factors (e.g., T stage, performance status, age, and surgical extent).
The clinical indicators provided by studies significantly varied in terms of details and types of indicators. Therefore, we assessed the comparability of candidate studies via discussion between two independent clinical oncologists. Studies in which the ART arm had clearly unfavorable clinical factors were categorized into comparability group 2. In group 2 studies, one or more of the following factors had significantly higher rate in ART arms: rates of nonradical surgery, advanced stage, LN+, and positive RM (defined as having either a statistically significant or >20% difference). Studies with patient distribution in a direction unfavorable to ART arms with clinical discretion (e.g. patients with status of positive RM or LN+ underwent ART in a certain institution) were also categorized into group 2. On the other hands, studies with no evidence that the clinical profiles of the ART arms were inferior to non-ART arms into group 1. Considering that most related studies were nonrandomized, we used the Newcastle-Ottawa scale [11]. We investigated the studies recruited to be included in three categories; score of 8 to 9 was regarded as high quality, 5 to 7 as medium quality, and less than 5 as low quality.
Low quality studies were discussed among authors to be excluded in pooled analyses, according to the advice of the Cochrane handbook that only observational studies with moderate to low risk of bias should be pooled analyzed [12]. The above comparability and quality assessments were performed by two independent researchers; any disagreement was resolved through discussion.

Statistics
A random-effects model was used for pooled analyses, considering possible heterogeneity in clinical profiles, treatment details among institutions, and the Cochrane Handbook [13] recommendation that a random-effects model should be used as the default choice for nonrandomized studies. Pooled odds ratios (ORs) were calculated for OS, DFS, and locoregional and distant recurrence. Moreover, 3-and 5-yr OS percentiles were also pooled.
Grade ≥3 complications were subjectively assessed. For studies with more than three arms (e.g., surgery only, surgery and chemotherapy, and surgery and chemoradiotherapy), two arms (the ART arm and the other arm most comparable to the ART arm) were chosen for analyses.
Heterogeneity assessment was performed using I 2 statistics [14] and the Cochran Q test, [15] in which p<0.1 and I 2 >50% were considered to indicate significant heterogeneity. Subgroup analyses were performed for OS (the primary endpoint) using the above comparability categories. Publication bias assessments including >10 studies were performed using visual funnel plots and quantitative Egger's test [16]. Possible publication bias was deemed to be present if the funnel plots showed asymmetry and the Egger's test p value was <0.1. Duval and Tweedie's trim-and-fill method [17] was performed for analyses with possible publication bias.

Results
In total, 2491 studies were initially obtained from the search. Studies with irrelevant formats and duplicated records among the databases were filtered using automated processes.  Table   1 shows the general information of the included studies (full version in Supplement Table 1).

Quality and bias assessments
Study quality was assessed using the Newcastle-Ottawa scale. Considering the low incidence of GBC, specificity of treatment modality, reliability of treatment records, and lack of the possibility of the outcome of interest being present at study initiation, most studies received full points for the selection criteria. Regarding outcome criteria, since all studies involved outcome data obtained from hospital records or national databases, they received points according to follow-up period and quality. Regarding comparability, two points were given if there was comparable data on two or more clinical factors, one point was given to studies that had information on a single clinical factor or institution's treatment policy. Studies without relevant information had zero point on comparability. Since all studies analyzed were to have medium or high qualities, all recruited studies were pooled analyzed for endpoints of interest.
The details and results of quality assessment are shown in Supplement Table 2.

Synthesis of clinical endpoints
The median survival of all cohorts was 23 months (range, 7-58 months). In the ART arm, the median survival was 25 months (range, 13-58 months), whereas it was 15 months (range, 7-50 months) in the non-ART arm. The median 5-yr OS percentile was 39% (range, 0-81%) in the ART arm and 23% (range, 0-73%) in the non-ART arm.
In pooled analyses of the primary endpoint (i.e., OS), the OR was 1 Table 3. Forest plots of each clinical endpoint (OS, DFS, locoregional recurrence, and distant metastasis) are shown in Figure 2. The detail of comparability assessment categorizing the comparability group is described in Supplement Table 3.

Heterogeneity analyses and publication bias assessment
Significant heterogeneity was found in pooled analyses of OS among all included cohorts and cohorts in comparability group 2, as well as in pooled analyses of DFS among the available studies. These heterogeneities may be influenced by differences in patient characteristics, differences in surgical extent, and simple subgrouping (e.g., owing to the limited number of studies, especially in comparability group 1, subgroup analyses were performed with two groups). Publication bias was assessed in pooled analyses including >10 studies, and no possible publication bias was detected.

Complications
Most studies did not report numerous or specific ART-related complications. Yang et al. [38] reported an overall incidence of nine cases (21%) of grade 3 complications in which the symptoms were reversible. Gu et al. [23] reported that although grade 1 or 2 hematologic and gastrointestinal toxicities were common in the ART arm, no grade ≥3 complications occurred.
Mondragón-Sánchez et al. [33] reported three cases of chronic abdominal pain that were difficult to control. Okamoto et al. [35] reported one case of postoperative mortality in the non-ART arm and two cases of mortality due to subphrenic abscess in the ART arm, of which one case was related to intraoperative radiotherapy-related stump injury.

Discussion
Most of the studies on adjuvant treatment for GBC to date are retrospective case series. As the field of radiation oncology is relatively small with less established research capacity, fewer related studies exist than that in medical and surgical oncology. To our knowledge, no randomized studies have verified the benefits of ART for GBC [39] Therefore, in the current clinical situation, therapeutic decisions are made by interpreting information obtained from existing retrospective studies.  1.92, p=0.008). Additionally, in the group of studies with an ART arm having an inferior clinical profile (group 2), the OR was 1.03 (p=0.865), and the 5-yr OS rate was 34.1% vs. 40% (ART vs. non-ART). These results suggest that ART might yield comparable oncologic outcomes even in patients with an inferior clinical profile. As the reduction in locoregional recurrence with ART (OR: 0.21, locoregional recurrence rate: 19% vs. 51.7%) but not in distant metastasis (OR: 1.32, p=0.332) was significant, the OS benefit in group 1 and the comparable results in group 2 seemed to be affected by the reduction in locoregional recurrence.
Only few studies have reported ART-related complications, partly because most included studies were conducted by surgeons or medical oncologists. We investigated the included studies to identify complications, especially grade ≥3 gastrointestinal toxicities, but none of the studies clearly reported such complications. Okamoto et al. [35] reported a fatal case after intraoperative radiotherapy; however, intraoperative radiotherapy is not a common modality in current clinical practice. From the perspective of radiotherapy techniques, as conventionally fractionated schemes of 45-50 Gy (below the tolerance dose of the small bowel) [41] were mostly used and only a small portion of the normal liver was irradiated, the possibility of serious complications might be insignificant. Additionally, although three-or two-dimensional multibeam treatment was used in studies included in the present meta-analysis, intensitymodulated radiotherapy has been increasingly used in clinical practice in the recent decade [42].
Sun et al. [43] reported that the application of intensity-modulated radiotherapy significantly reduced the irradiated volume of the normal liver and kidney. Among the 20 patients treated, no grade ≥3 complications occurred; grade 2 nausea and diarrhea were the most common complications. Bittner et al. [44] also reported that the application of intensity-modulated radiotherapy significantly reduced the incidence of grade ≥3 diarrhea (2% vs. 11.6%, p<0.001) and late gastrointestinal toxicity (5% vs. 10.6%, p=0.017) compared with the application of conventional three-dimensional radiotherapy in treating pancreatic cancer, which has a similar radiotherapy target to GBC.
Thus, the results of this meta-analysis support more active ART application in GBC treatment.
Particularly, ART should not be limited to LN+ or RM+ patients, but clinical decisions should be made after a comprehensive consideration of various clinical factors and situations.
Our study had some limitations. Meta-analyses of observational studies have been controversial because the pooled effect can be affected by heterogeneity among studies and uncontrolled confounders [13]. However, narrow subjects, such as adjuvant treatment of rare cancers such as GBC, may inevitably be supported by observational studies [45,46]. In this situation, a meta-analysis is one of the few methods available to aid clinical decisions. Notably, to enhance the reliability of the meta-analysis, we performed statistical complements, including heterogeneity and publication bias assessments and subgroup analyses, as well as comparability assessment on an individual study basis. Another limitation is that this study had no specific indication for ART, compared with previous studies and meta-analyses. However, this study aimed to support the more active application of ART by interpreting the literature, while providing reliable synthesized information. Similarly, our study is expected to stimulate future studies investigating more specific and personalized indications that are not limited to the LN+ or RM+ status.

Conclusion
This study comprehensively considered comparability between arms on an individual study basis and confirmed that ART has relevant oncologic benefits. Our results support the clinical application of ART in practice, and provide a basis for future research toward the creation of    *Comparability group 2: studies in which the adjuvant radiotherapy (ART) arm had an inferior clinical profile than the non-ART arm; group 1: studies in which ART arm had statistically similar profile or no evidence of having inferior clinical factors compared to non-ART arm.  Abbreviations: ART, adjuvant radiotherapy; OS, overall survival *Comparability group 2: studies in which the adjuvant radiotherapy (ART) arm had an inferior clinical profile than the non-ART arm; group 1: studies in which ART arm had statistically similar profile or no evidence of having inferior clinical factors compared to non-ART arm.