Intrapleural Administration With Rh-Endostatin and Chemical Irritants in the Control of Malignant Pleural Effusion: A Systematic Review and Meta-Analysis

Introduction A modified and recombinant human endostatin (Rh-endostatin) is often used in the control of malignant pleural effusion (MPE) through intrapleural infusion. Objectives To demonstrate the clinical response, survival, and safety of Rh-endostatin plus chemical irritants, their optimal combinations, treatment threshold, and optimal usage, we performed a new systematic review and meta-analysis. Methodology All randomized controlled trials (RCTs) were collected from Chinese and English electronic databases (from inception until August 2020). We pooled the data using a series of meta-analyses and summarized the evidence quality following the Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach. Results We included 75 RCTs recruiting 4,678 patients, which reported six combinations for Rh-endostatin plus chemical irritants. Among the six combinations, only Rh-endostatin plus cisplatin (DDP) with enough trials might improve the complete response [2.29 (1.93, 2.71)] and quality of life [3.01 (2.49, 3.63)] and reduce treatment failure [0.29 (0.25, 0.33)] and progressive disease [0.27 (0.22, 0.34)]. It might not increase the risk of adverse drug reactions. For patients with lung cancer, moderate to massive effusion, initial treatment, Karnofsky Performance Status (KPS) score ≥60, or anticipated survival time ≥3 months, Rh-endostatin (30–45 mg each time, once or twice a week 3–4 times) plus DDP (30–60 mg/m2) obtained a significant improvement in clinical response and a reduction of failure and progressive disease. Most results had good robustness and moderate quality. Conclusions Current evidence suggests that Rh-endostatin with DDP may be an optimal combination, which may improve clinical response and reduce failure and progressive disease with good safety. Rh-endostatin (30–40 mg each time, once or twice a week 3–4 times) with DDP (30–40 mg/m2) may be an optimal usage for achieving an ideal response.


INTRODUCTION
Malignant pleural effusion (MPE) is a common clinical problem in patients with malignant tumors, with an estimated annual incidence of at least 150,000 in the USA (1). Based on postmortem records, MPE was found in 15% of patients who died with malignant tumors (2). Most patients often suffered from breathlessness and chest pain. The quality of life (QOL) was poor, and the median survival time was only 3-12 months (2,3). Chemical pleurodesis is a first-line treatment for symptomatic patients with MPE and suspected expandable lung (4,5) and a procedure performed to obliterate the pleural space to prevent recurrent MPE using a chemical irritant as platinum, bleomycin (BLM), tetracycline, doxycycline, or silver nitrate, among others (3)(4)(5)(6). However, these strategies are mostly of palliative value and focus on the control of symptoms and improvement of QOL and fail to improve survivals. So, new control strategies are urgently needed.
Proangiogenic factors have been implicated as a critical cytokine in the occurrence, development, and transferring of MPE (7)(8)(9)(10). Endostatin, a 20-kDa C-terminal fragment of type XVIII collagen, is one of the most potent inhibitors of angiogenesis (11). Endostatin and its derivatives have been reported to be more effective when combined with chemotherapy, radiotherapy, or gene transfer in the treatment of malignant tumors (12,13). Endostar, a modified and recombinant human endostatin (Rh-endostatin), was the approved regimen in non-small-cell lung cancer (NSCLC) by the State Food and Drug Administration of China in 2005 (14). The expert consensus also recommends Rh-endostatin plus first-line chemotherapy to treat stage III/IV NSCLC (15,16). Interestingly, eight systematic reviews (SRs)/meta-analyses had reported that intrapleural administration of Rh-endostatin with platinum (17,18), cisplatin (DDP) (19)(20)(21)(22)(23), or chemotherapeutic agents (24) might improve the objective response rate [complete response (CR), partial response (PR)], disease control rate [CR + PR+ no response (NR)/stable disease (SD)], and QOL, without an increase in the incidence of adverse drug reactions (ADRs) in MPE. Three meta-analyses (25)(26)(27) had reported that Rh-endostatin with DDP also might obtain the same effects in MPE from lung cancer. Based on the above evidence, Rh-endostatin alone or plus chemical irritants was recommended in the control of MPE by expert consensus from China (28). However, strong clinical heterogeneity was found in the patient features, types, combinations, and usages of Rh-endostatin/chemical irritants. The drug usages are complex, diverse, and even inappropriate. Obviously, the current studies ignored clinical heterogeneity. Current evidence (17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27) failed to conclusively demonstrate whether Rh-endostatin plus chemical irritants improves clinical response, survival, and safety. Their optimal combinations, therapeutic threshold, and optimal usage remain unclear. In addition, no evidence revealed their thoracentesis-related adverse events (TRAEs). All these have become the new bottleneck of rational drug use decision.
Recently, many new trials (29)(30)(31) have been published. So, we performed a new SR and meta-analysis to further demonstrate the clinical response, survival, and safety of Rh-endostatin with chemical irritants, reveal their optimal combinations, therapeutic thresholds, and optimal usage for achieving a desired response, and provide evidence for developing an optimal control strategy of MPE.

Selection of Studies
Two evaluators (C-QW and MH) were asked to collect the qualified trials about Rh-endostatin plus chemical irritants for MPE according to the preestablished inclusion and exclusion criteria.

Assessment of Methodological Bias Risk
Two evaluators (X-RH and QC) were asked to assess the bias risk of methodology using the Cochrane Collaboration's risk of bias assessment tool for RCTs (33). The bias risk was assessed as a judgment (high, low, or unclear) for individual elements of five domains (selection, performance, attrition, reporting, and other).

Indicator Definition
The clinical responses were evaluated using CR, treatment failure, and progressive disease (PD). Based on previous studies (34)(35)(36)(37), we integrated all the criteria as follows: (i) CR, (ii) PR, (iii) NR or SD; and (iv) PD (Supplementary Material S2). Treatment failure was defined as NR/SD plus PD (38). Survival was defined as overall survival (OS) rate, progression-free survival (PFS) rate, or hazard ratio (HR) of the OS and PFS. Using the Karnofsky Performance Status (KPS) scale, if the KPS score increased ≥10 after perfusion, the QOL was improved.
The secondary outcomes were ADRs and TRAEs. According to the World Health Organization (WHO) (39) or Common Terminology Criteria for Adverse Events (CTCAE) standards (40), ADR was defined as neutropenia, thrombocytopenia, anemia, cardiotoxicity, hepatotoxicity, nephrotoxicity, gastrointestinal reactions, alopecia, peripheral neuritis, chest pain, and fever, among others. TRAE was defined as treatment-related mortality (TRM) and a series of clinical symptoms such as respiratory failure, pneumothorax, cutaneous emphysema, or catheter-related infection/ chest infection, among others.

Data Collection
Two evaluators (X-TZ and T-yF) collected all the data using a predesigned data extraction form. The data included the first author, year of publication, and demographic information of patients; baseline characteristics such as primary tumors, pleural fluid volume, KPS score, treatment history (initial treatment, retreatment, or both), anticipated survival time (AST), sample size, drainage methods [indwelling pleural catheters (IPCs) or thoracocentesis]; combinations and usages of Rh-endostatin and chemical irritants; evaluation time and follow-up protocols; and outcomes including CR, treatment failure, PD, OS, PFS, QOL, ADRs, and TRAEs. Additionally, we contacted the corresponding author to obtain the available survival data. If the authors were unavailable, we adopted the Engauge Digitizer 4.1 to transform the Kaplan-Meier survival curves into available data (41,42).

Statistical Analysis
According to the data features, the odds ratio (OR) or hazard ratio (HR) and their 95% CI were used to quantify the CR, treatment failure, PD, OS, PFS, QOL, ADRs, and TRAEs, and p < 0.05 was considered a statistical significance. Two evaluators (C-QW and X-RH) conducted a series of meta-analyses using the Review Manager 5.4.1 (as recommended by the Cochrane Collaboration). The Cochran's c 2 test and I 2 statistic were conducted to analyze the potential statistical heterogeneity. If p ≥ 0.1 and I 2 ≤ 50%, a fixed-effects model (FEM) was used to pool the OR or HR and their 95% CI. Otherwise, a randomeffects model (REM) was used. If the number of trials was larger than 10, a funnel plot and Egger/Begg's test were used to examine the potential publication bias.
When at least one item was considered a high risk, the trial was defined as poor quality. When the result was statistically different and beneficial to Rh-endostatin infusion, the trial was defined as an underestimated or overestimated trial following our experiences (38,43,44). According to the principle of underestimating efficacy and overestimating risk, we established a sensitivity analysis model to analyze the robustness of the results before and after eliminating the trials with poor quality, underestimation, or overestimation.

Subgroup Analysis
Following the guideline (45) and our previous experiences (38,43,44), we established a subgroup analysis model to analyze the clinical heterogeneity and the effects of variables on CR, treatment failure, and PD and to reveal their treatment thresholds and optimal usage for achieving an ideal response. The variables included patient features, drainage methods, and combinations of Rh-endostatin plus chemical irritants and their dose, treatment frequency, and times. Finally, a univariable random-effects meta-regression was conducted to reveal the relevance between each variable and CR, treatment failure, or PD and a post-hoc multiple regression analysis adjusting for their OR under all variables.

Summary of Evidence Quality
Following the Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach and integrating the results of the sensitivity analysis, we developed a quality summary model to summarize the evidence quality and classify them as "high," "moderate," "low," or "very low" (38,43,44) (Supplementary Material S3). The quality was downgraded according to five domains as follows: (1) methodological bias risk; (2) statistical heterogeneity; (3) indirectness; (4) imprecision; and (5) publication bias. Two evaluators (X-FC and C-QW) used the GRADE profiler to summarize the quality and generate the absolute estimates for the CR, treatment failure, PD, OS rate, PFS rate, QOL, ADRs, and TRAEs (46).

Methodological Quality Assessment
Thirty-four trials reported the random sequence generation using a random number table (a low risk of selection bias) (30, 31, 54, 56-58, 62, 65, 67, 69, 75, 81, 82, 84, 85, 89, 90, 93, 97, 101, 102, 105, 106, 109-114, 116, 117, 120, 126, 127), and two trials reported the odd or even random (a high risk of selection bias) (60,123). Two trials reported the allocation concealment using an envelope (a low risk of selection bias) (73,76), and two trials reported the allocation exposure (a high risk of selection bias) (60,123). With the exception of one open RCT (119), the remaining trials failed to clearly report the blindings (an unclear risk of performance bias). All trials reported the complete outcome data (a low risk of attrition bias). Forty-four trials selectively reported the ADRs, and one trial selectively reported the CR (a high risk of reporting bias) (66). The comparability between groups (an unclear risk of other biases) was unclear in 12 trials (Figure 2).

Overall Survival
Nine trials reported the survivals (29-31, 63, 71, 75, 98, 105, 116). Only five trials reported the OS time and PFS of Rh-endostatin with DDP (30,71,75,98) or LBP (29), but without the available data. Five trials reported the OS rates, and three reported the 1year OS rate of Rh-endostatin with DDP (31,75,116). The statistical heterogeneity between trials was not found using Cochran's c 2 test and I 2 statistic (I 2 = 0%). So, we pooled the data using an FEM. The 1-year OS rate of Rh-endostatin with DDP was significantly higher than that of DDP alone. The OR was 3.32 (95% CI 1.63-6.75, p = 0.0009) (Figure 4). The remaining OS rates were reported in only one trial, and the data were analyzed descriptively using forest plots. Statistical analysis showed that the 0.5-year OS rate of Rh-endostatin with DDP (116), 1-year OS rate of DDP/BLM (63), 2-year OS rate of DDP (75), and 3-year OS rate of LBP (105) were significantly higher than that of irritants alone. The ORs were 5.36 (95% CI    Figure 5). The statistical heterogeneity between trials was not found using Cochran's c 2 test and I 2 statistic (I 2 = 0%). So, the data were pooled using an FEM. The OR was 3.01 (95% CI 2.49-3.63, p < 0.00001), which indicated that the QOL was significantly higher than that of DDP alone.

Subgroup Analysis of Clinical Responses
The patient feature was defined as primary tumor, pleural fluid volume, treatment history, KPS score, and AST. First, the primary tumor was classified as lung cancer, breast cancer, or malignant tumors. In patients with lung cancer/malignant tumors, Rh-endostatin with DDP obtained a significant increase of CR and a reduction of failure and PD. In breast cancer, it only obtained a reduction of failure and PD (Table 3A and Figures S12, S14, S16). The pleural fluid was classified as small to large, moderate to large, or large (Table 3B and Figures S18, S20, S22); treatment history was initial treatment, retreatment, or both (Table 3C and Figures S24, S26, S28); KPS score was <50, ≥50, or ≥60 (Table 3D and Figures S30, S32, S34); and the AST was ≥2 months or ≥3 months (Table 3E and Figures S36, S38, S40). In patients with moderate to massive effusion, initial treatment, KPS score (≥60), or AST (≥3 months), the Rh-endostatin with DDP groups obtained a significant increase of CR and a reduction of failure and PD. The majority of patients mainly received the IPCs (Table 3F and Figures S42, S44, S46). Subgroup analyses found that whether IPC is used or not had no effect on the clinical responses. Rh-endostatin was used with 30-90 mg each time, once or twice a week 1-12 times (Tables 3G-I and Figures S48-S64). DDP was used with 30-40 mg/m 2 or 50-60 mg/m 2 each time (Table 3J and Figures S66, S68, S70). Rh-endostatin (30-35 mg or 40-45 mg each time, once or twice a week 3-4 times) However, univariate regression analysis did not discover a positive or negative correlation between CR, treatment failure, and PD and each variable (Table 3 and Figures S13-S71).
Multiple meta-regression analysis also did not discover a positive or negative correlation (Table 3).

Publication Bias Analysis
In Rh-endostatin with DDP, more than 10 trials were included for the CR, treatment failure, PD, QOL, and ADRs. So, funnel plot and Egger/Begg's tests were used to analyze their potential bias of publication. The analysis found a publication bias in CR (p < 0.001, 95% CI 0. 74 (Figures 6E-L). The trials objectively reported the ADRs.

Sensitivity Analysis
In Rh-endostatin with DDP, the poor trials involved clinical response, 1-year OS rate, QOL, and ADRs. Some trials overestimated the CR, 1-year OS rate, and QOL and underestimated the treatment failure and PD. According to the underestimating efficacy and overestimating risk, we evaluated the robustness through removing the poor trials, overestimation/ underestimation, and both. Before and after removing the poor trials, the results demonstrated a good robustness of all outcomes. Before and after removing the overestimation and both, the OR of 1-year OS rate was poor robust, and other indicators were robust. In addition, the OR of CR was robust in Rh-endostatin with NDP, and the OR of CR and PD was robust in Rh-endostatin with BLM ( Table 4).

Quality of Evidence
In methodology, 46 poor trials were included for this analysis. Sensitivity analysis demonstrated that the OR of 1-year OS rate was poor robustness in Rh-endostatin with DDP, the CR and PD were poor in Rh-endostatin with NDP, and the CR was poor in Rh-endostatin with BLM. Therefore, we downgraded their quality by two grades. Other results had good robustness, and we downgraded their quality by one grade. No heterogeneity was found in all of the indicators; all indicators were not downgraded. In Rh-endostatin with DDP, the sample size of 1-year OS rate, alopecia, hypertension, and hemorrhage was lower than 300 subjects. In Rh-endostatin with NDP, CBP, LBP, or BLM, the CR, treatment failure, and PD were lower than 300. So, we downgraded their quality by one grade. In addition, the funnel plot and Egger's test showed a publication bias of CR, treatment failure, PD, and QOL in Rh-endostatin with DDP. The sensitivity analysis results were good robust, and we did not downgrade their quality. So, we summarized a low quality for 1-year OS rate, alopecia, hypertension, and hemorrhage and a moderate quality for other results of Rh-endostatin with DDP; a low quality for CR and treatment failure in Rh-endostatin with NDP or BLM; and a very low quality for the remaining indicators ( Table 5).

DISCUSSION
Intrapleural administration of Rh-endostatin alone or plus chemical irritants is recommended for the control of MPE by expert consensus from China (28). To demonstrate the optimal combinations of Rh-endostatin with chemical irritants and their clinical efficacy and safety, we further included 75 trials for analysis (29-31, 54-123, 126, 127). In this study, we found six combinations such as Rh-endostatin with DDP, CBP, NDP, LBP, BLM, or paclitaxel. The results of meta-analysis demonstrated that the Rh-endostatin with DDP might improve the response and reduce the failure and PD, with "moderate" quality. We further found that this combination might also improve the QOL, without increasing the risk of hematotoxicity, cardiotoxicity, hepatotoxicity, nephrotoxicity, gastrointestinal reaction, chest pain, and fever, with "moderate" quality. In addition, there were limited reports on the combinations of Rh-endostatin with CBP, NDP, LBP, BLM, or paclitaxel. Only the combinations Rh-endostatin with NDP and LBP might increase the response and reduce the failure and PD, but with "low to very low" quality. A few trials reported the survival; only Rh-endostatin with DDP or LBP might improve 1-to 2-year OS rate, with "low to very low" quality. And most trials failed to report the TRAEs and TRM. Evidently, these outcomes are not fully evaluated and need to be further confirmed. Eight previous evaluations had reported that the intrathoracic infusion with Rh-endostatin combined with platinum (17,18), DDP (19)(20)(21)(22)(23), or chemotherapeutic agents (24) might improve the objective response rate, disease control rate, and QOL without an increase in the incidence of ADRs in MPEs from malignant tumors. Rh-endostatin with DDP might also obtain the same effects in MPEs from lung cancer (25)(26)(27). In this evaluation, we redefined the clinical efficacy as CR, treatment failure, PD, and survival and added the TRAEs and TRM as security indexes, further integrated previous studies (17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27), and     1 Most trials had an unclear risk, and some trials had a high risk. If good robustness, we downgraded it by one grade. 2 Publication bias was found in them, and the result was overestimated. The result showed good robustness and was not downgraded. 3 Publication bias was found in them, and the result was underestimated. The result showed good robustness and not downgraded. 4 The number of patients in each result was less than 300, and we downgraded it by one grade. 5 Most trials had an unclear risk and no high risk, and we downgraded it by one grade. 6 Most trials had an unclear risk, and some trials had a high risk. If sensitivity analysis results had poor robustness, we downgraded it by two grades. 7 All trials had a high risk, and we downgraded it by two grades.
added 36 trials with 2,209 patients for analysis. This evaluation found that all six combinations, especially Rh-endostatin with DDP, might show an improvement of clinical response and a reduction of failure and PD, without an increase of the ADRs. The result indicates that a significant synergistic effect exists between Rh-endostatin and DDP. In clinical practice, the BRMs (38,128) and TCMIs (129)(130)(131) were also used in the control of MPEs through intrathoracic infusion. Previous studies (129)(130)(131) had reported that chemical irritants plus TCMIs might increase the clinical benefit rate and decrease the ADRs. Chemical irritants plus BRMs (38,128) also obtain the same benefit. But compared with TCMIs and BRMs, Rh-endostatin did not reduce the risk of ADRs, which may limit its clinical application. All in all, the results indicate that intrapleural administration of TCMIs, BRMs, or Rh-endostatin might be an important pathway to perform pleurodesis and control the hydrothorax (Figure 7). In a previous analysis (38), we found that moderate to large pleural fluid, KPS scores ≥50, or AST ≥3 months might be the treatment thresholds for lentinan with DDP. So, we performed a series of subgroup analyses to reveal the therapeutic thresholds and optimal usage of Rh-endostatin with DDP for achieving a desired response and security. Our analyses found that MPE patients with lung cancer, moderate to massive effusion, initial treatment, KPS score ≥60, or AST ≥3 months might be more suitable for Rh-endostatin with DDP infusion than patients with other conditions. The infusion conditions, the volume of pleural effusion, treatment history, and AST are the same as that of lentinan with DDP infusion. But Rh-endostatin infusion requires a higher KPS (≥60) than lentinan infusion, which suggests that Rh-endostatin infusion seems to have a higher threshold than lentinan. Yoon et al. (132) had reported that poor performance status [Eastern Cooperative Oncology Group (ECOG) 3 or 4] was an independent risk factor of poor survival after video-assisted thoracic surgery (VATS) talc pleurodesis. Compared with VATS talc pleurodesis, endostatin infusion seems to have a lower threshold. In all, the results indicate that endostatin seems to have a special threshold for infusion. The moderate to massive effusion, KPS score ≥60, or AST ≥3 months may be a treatment threshold for Rh-endostatin with DDP, which may be more suitable for MPE with initial treatment or for lung cancer. So, the objective assessment of patients' baseline should be considered when choosing Rh-endostatin with DDP. In expert consensus (28) (Figure 7). The dose of Rh-endostatin and DDP may be lower than the recommended dose (28). All these findings demonstrate a possible treatment threshold and optimum strategy of intrapleural administration of Rh-endostatin with DDP for MPEs, which is of important clinical significance for further improving scientific decision-making of drug rational application. But the meta-regressions did not further confirm the positive or negative correlation. In addition, whether endostatin with DDP infusion is suitable for drug-resistant, refractory, retreatment, or recurrent MPEs and MPEs from other tumors remains unclear. For Rh-endostatin with CBP, NDP, or LBP/BLM, the treatment threshold and optimal strategy remain unclear. So, these questions need to be further answered.
All kinds of potential limitations should be taken into consideration. First, in this study, only Chinese and English databases were searched, which might result in potential retrieval biases. Second, a considerable number of trials did not clearly describe the baseline features such as the volume of hydrothorax, KPS score, AST, initial treatment, retreatment, drug-resistant, refractory, or recurrent. Third, only 34 studies described the generation of random sequence, and 44 studies selectively reported the CR, ADRs, or TRAEs. Fourth, there was lack of a unified standard for clinical efficacy of chemical pleurodesis in MPEs, and the majority of trials did not clearly report the survivals, TRAEs, and TRM. Fifth, due to limited trials for Rhendostatin with CBP, NDP, LBP/BLM, the treatment thresholds and optimal strategy remain unclear. Sixth, the univariate or multivariate regression analysis did not find any positive or negative correlation between clinical responses and all variables.

CONCLUSION
The evidence indicates that among all six combinations, only Rhendostatin with DDP may be an optimal combination, which may improve the clinical response and QOL and reduce the failure and PD without increasing the ADRs in MPEs. For Rh-endostatin with DDP infusion, the treatment threshold may be moderate to massive effusion, KPS score ≥ 60, or AST ≥3 months. The combination may be more suitable for MPE with initial treatment or for lung cancer. Rh-endostatin (30-40 mg each time, once or twice a week 3-4 times) with DDP (30-40 mg/m 2 ) may be a possible strategy for achieving an ideal response. The pooled results from limited trials reveal that Rh-endostatin with DDP/LBP might increase the 0.5-2-year OS rate. But the evidence fails to support that Rh-endostatin plus chemical irritants also does for MPE what it does for non-lung cancer, refractory/recurrent, or drug-resistant patients. Their ADRs and potential TRAEs remain unclear. In addition, whether Rh-endostatin with CBP, NDP, or LBP/BLM improves the clinical response and their treatment thresholds and optimal strategy also remains unclear. All of these questions need further new trials to demonstrate. Finally, these findings provide valuable references for an optimal control strategy based on Rhendostatin in MPE.

DATA AVAILABILITY STATEMENT
The original contributions presented in the study are included in the article/Supplementary Material further inquiries can be directed to the corresponding authors.

AUTHOR CONTRIBUTIONS
Conception and design by ZX, XX, and X-FC. Development of methodology by ZX, C-QW, and X-FC. Literature search by C-QW and HJ. Article selection by C-QW and MH. Assessment of methodological bias risk by X-RH and QC. Data extraction by X-TZ and T-yF. Statistical analysis by C-QW and X-RH. GRADE assessment by X-FC and C-QW. Preparing the manuscript draft by ZX, XX, and X-FC. Review and revision of the manuscript by XX, X-FC, LZ, JL, and J-HF. Study supervision by ZX. All authors contributed to the article and approved the submitted version.