Efficacy and safety of Chinese patent medicine (Jinlong capsule) in the treatment of advanced hepatocellular carcinoma: a meta-analysis

Hepatocellular carcinoma (HC) is the seventh most common cancer and the third leading cause of cancer-related deaths worldwide [1]. It was estimated that 840000 new cases of HC and 781631 HC-related deaths occurred worldwide in 2018 [1]. Despite advances in diagnostic methods, early detection of HC remains difficult [2]. In most patients, HC progresses to the intermediate and advanced stages, for which the 5-year survival rate is <17% [2,3]. Surgery and liver transplantation are considered to be the optimal treatment options for advanced HC; however, only a small proportion of HC patients are able to undergo potentially curative resection [4]. In addition, the therapeutic effects of current conventional treatments for advanced HC, such as radiotherapy and chemotherapy, remain unsatisfactory [2,5,6]. Therefore, more effective, comprehensive treatment strategies are required.

Jinlong capsule (JLC) is a traditional Chinese medicine with the function of tonifying qi and blood, dredging collaterals and detoxification, and has been widely used as an effective adjuvant drug in cancer treatment [7,8]. It is traditionally prepared from three animals with medicinal properties: Bungarus, Agkistrodon and Gecko [7,8]. The active ingredients of JLC comprise at least 17 amino acids, including histidine, serine, arginine, glycine and aspartic acid, among others, which are extracted using modern cryogenic and biochemical separation techniques from freshly prepared animal drugs [8,9].

Research has shown that JLC can effectively reverse multiple-drug resistance in cancer cells and improve the efficacy of chemotherapy [10,11]. Many in vitro studies have demonstrated that JLC can suppress cellular mitotic division and inhibit proliferation principally by arresting the cell division cycle at S and G₂/M [8,9]. It can also promote cancer cell apoptosis through activation of the pro-apoptotic proteins BNIP3, Bax and caspase-3, and down-regulating the expression of Bcl-2 for survival [8,9]. Several studies have indicated that conventional treatment combined with JLC is more effective for the treatment of advanced HC than conventional treatment alone [12,13]. Despite intensive studies, the clinical efficacy and safety of the combination of conventional treatment and JLC has not been systematically evaluated. In the present study, we conducted a meta-analysis to investigate the efficacy and safety of conventional treatment combined with JLC compared with conventional treatment alone for advanced HC, to provide a scientific reference for the design of future clinical trials.

The present meta-analysis was performed in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, and the Cochrane Handbook. Ethics approval was not necessary due to the nature of the study (i.e. meta-analysis).

A literature search was conducted using eight electronic databases, including the Web of Science, PubMed, Cochrane Library, Embase, Chinese Biological Medicine Database (CBM), China National Knowledge Infrastructure (CNKI), Chinese Scientific Journal Database (VIP) and the Wanfang database, for original articles published before February 2019. The search terms included: ‘Jinlong capsule’ combined with ‘liver carcinoma’ or ‘hepatocellular carcinoma’ or ‘liver cancer’ or ‘hepatocellular cancer’.

Studies fulfilling the following criteria were included in the meta-analysis: controlled trials investigating patients with advanced HC; those with >30 HC patients; studies comparing the clinical outcomes of conventional treatment plus JLC adjuvant therapy (experimental group) with conventional treatment alone (control group); and conventional treatments including transcatheter arterial chemoembolization, chemotherapy, radiotherapy, radiofrequency ablation, percutaneous ethanol injection, support and symptomatic treatment and ultrasound therapy.

Studies involving patients with mixed malignancies, non-controlled trials, non-clinical studies, literature reviews, meta-analyses, meeting abstracts, case reports, duplicate studies, experimental models and those with insufficient available data were excluded.

Data were independently extracted by two investigators (H.X. and W.W.) according to the same inclusion criteria; disagreements were adjudicated by a third reviewer (C.D.). The extracted data were as follows: name of the first author; year of publication; tumor stage or Karnofsky Performance score (KPS); number of cases; intervention methods; control group regimens; dose of JLC and study parameters. To ensure the quality of the meta-analysis, the quality of the included randomized and non-randomized controlled trials was evaluated according to the Cochrane Handbook tool [14] and Methodological Index for Nonrandomized Studies (MINORS, Supplementary Table S2), respectively [15].

Clinical responses included treatment efficacy, quality of life (QoL), immune function, and adverse events. Treatment efficacy was assessed in terms of overall survival (OS), overall response rate (ORR), and disease control rate (DCR). Patient QoL was evaluated using QoL improved rate (QIR) and KPS. Immune function indicators (CD3⁺, CD4⁺, CD8⁺ and natural killer [NK] cell percentage and CD4⁺/CD8⁺ ratio) and adverse events, including leukopenia, adverse gastrointestinal effects, nausea and vomiting, anorexia, thrombocytopenia, hepatotoxicity, myelosuppression and anemia, were assessed and compared between the two groups.

Review Mananger (RevMan) version 5.3 (Nordic Cochran Centre, Copenhagen, Denmark) and Stata version 13.0 (StataCorp., College Station, TX, U.S.A.) were used for statistical analyses. Data were mainly expressed as odds ratio (OR) with corresponding 95% confidence interval (CI), and a two-tailed P<0.05 was considered to be statistically significant. Cochrane’s Q test and I² statistics were used to assess heterogeneity among the studies: if P> 0.1 or I²  <  50%, fixed-effects model was used for the meta-analysis; otherwise, random-effects model was used [16].

The presence of publication bias was investigated using the Begg’s and Egger’s tests, and funnel plots. A pooled analysis of publication bias determined that the trim-and-fill method should be applied to coordinate the estimates from unpublished studies, and the adjusted results were compared with the original pooled OR [17]. Sensitivity analysis was performed to evaluate the impact of different therapeutic regimens, sample size, and type of research on the clinical efficacy of the combination of conventional treatment and JLC.

The initial search retrieved a total of 328 articles, of which 225 were excluded due to duplication. After title and abstract review, 47 articles were further excluded because they did not include clinical trials (n=23), were unrelated studies (n=17), were reviews and meta-analyses (n=3), or were case reports (n=4), leaving 56 studies as potentially eligible. After detailed assessment of full texts, studies without a control group (n=9), those with <30 HC patients (n=6), inappropriate criteria for the experimental or control groups (n=5), and trials with insufficient data (n=7) were excluded. Ultimately, 29 trials [7,18–45], involving 2488 patients with advanced HC, were included in the final analysis (Figure 1).

All included studies were performed in different medical centers in China. In total, 1274 patients with advanced HC were treated using conventional methods in combination with JLC adjuvant therapy, while 1214 patients were treated using conventional methods alone. JLC was manufactured by Beijing Jiansheng Pharmaceutical Co., Ltd. The quality standards of the JLC in the present study were approved by the Chinese State Food and Drug Administration (SFDA), and granted a manufacturing approval number issued by the Chinese SFDA (Z10980041). All pharmaceutical companies involved followed the quality processing procedure outlined in the pharmacopeia. Study and patient characteristics are summarized in Table 1.

Quality assessment of the risk of bias is shown in Supplementary Figure S1 and Table S1. The results revealed that the literature retrieved for the present study was of good quality.

Twenty-five clinical trials [7,18–21,23–30,32–36,38,39,41–45], involving 2161 patients, compared ORR and DCR between the two groups. As shown in Figures 2 and 3, the pooled results revealed that patients who underwent combination therapy experienced significantly improved ORR (OR 2.06 [95% CI 1.71–2.49]; P<0.00001) and DCR (OR 2.17 [95% CI 1.74–2.71]; P<0.00001) compared with those who received conventional treatment(s) alone. The fixed-effect model was selected due to slight heterogeneity among the studies.

Eight clinical trials [18,22,23,26,28,31,35,42] with 1014 HC patients reported OS (Figure 4). Meta-analysis revealed that the 6- (OR 1.63 [95% CI 1.11–2.39]; P=0.01), 12- (OR 2.05 [95% CI 1.54–2.73]; P<0.00001), 24- (OR 2.45 [95% CI 1.44–4.19]; P=0.001) and 36-month OS (OR 2.41 [95% CI 1.56–3.74]; P<0.0001) of patients in the combined treatment group were significantly prolonged compared with the control group. There was statistical heterogeneity in 24-month OS (P=0.03, I² = 58%) according to the heterogeneity test. Therefore, a random-effects model was used to pool this meta-analysis. Otherwise, the fixed-effect model was used.

Ten trials [18,21,24,26,29,31,32,42,43,45] with 1040 participants evaluated QIR, and three trials [7,27,34], including 208 patients, reported KPS data (Figure 5). Results demonstrated that the QoL of HC patients in the combined group was significantly better than that of the control group, indicated by significantly increased QIR (OR 2.71 [95% CI 2.05–3.58]; P<0.00001) and KPS (OR 9.33 [95% CI 4.26–14.40]; P=0.0003). QIR (P=0.86, I² = 0%) was not heterogeneous among the studies; therefore, a fixed-effect model was used to analyze OR. Otherwise, a random-effect model was used.

Immune status of the patients was examined between the two groups in nine controlled studies [19,25,31,32,37,38,40,43,44]. The percentages of CD3⁺, CD4⁺ and NK cells, and CD4⁺/CD8⁺ ratio in the combined treatment group were significantly increased compared with those in the conventional treatment alone group, (CD3⁺, OR 13.69 [95% CI 7.40–19.98], P<0.0001; CD4⁺, OR 8.04 [95% CI 4.94–11.13], P<0.00001; NK, OR 6.29 [95% CI 2.88–9.69], P=0.0003; and CD4⁺/CD8⁺, OR 0.39 [95% CI 0.27–0.51], P<0.00001), whereas the proportions of CD8⁺ were clearly decreased (OR −5.62 [95% CI −7.68 to −3.56]; P<0.00001) (Figure 6). A random-effects model was used to pool this meta-analysis due to significant heterogeneity.

As shown in Table 2 and Supplementary Figure S2, patients treated with JLC and conventional methods exhibited lower incidences of leukopenia, hepatotoxicity and myelosuppression (leukopenia, OR 0.49 [95% CI 0.36–0.67], P<0.00001; hepatotoxicity, OR 0.29 [95% CI 0.12–0.69, P=0.005; and myelosuppression, OR 0.26 [95% CI 0.12–0.56], P=0.0007), whereas analysis of gastrointestinal side effects, nausea and vomiting, anorexia, thrombocytopenia and anemia (gastrointestinal side effects, OR 0.69 [95% CI 0.44–1.07], P=0.10; nausea and vomiting, OR 0.81 [95% CI 0.45–1.48], P=0.49; anorexia, OR 0.89 [95% CI 0.35–2.29], P=0.81; thrombocytopenia, OR 0.76 [95% CI 0.28–2.06], P=0.59; and anemia, OR 0.68 [95% CI 0.29–1.60, P=0.38) did not differ significantly between the two groups. A fixed-effect model was used to analyze OR rate due to low heterogeneity.

Publication bias was visually assessed using funnel plots (Supplementary Figure S3) and quantified using Egger’s test and Begg’s regression test (Supplementary Table S2). The funnel plots were symmetrical for 12- and 24-month OS, and CD4⁺, CD8⁺, CD4⁺/CD8⁺ and leukopenia, but were asymmetrical for ORR, DCR and QIR. To determine whether bias affected the pooled risk of ORR, DCR and QIR, a trim-and-fill analysis was performed. The adjusted OR indicated a trend similar to the results of the primary analysis (Supplementary Table S2), reflecting the reliability of the primary conclusions.

A subgroup analysis was conducted to explore the source of heterogeneity for ORR and DCR. As shown in Supplementary Table S3, results revealed that no significant difference was found between the therapeutic regimens, sample size, and types of studies.

JLC, a type of traditional Chinese medicine, has been clinically applied as an adjuvant therapy for decades. Several studies have reported that addition of JLC could be beneficial to patients with advanced HC [12,13]. Although there was a statistical analysis of published clinical trials, the exact therapeutic effects are yet to be systematically evaluated because of small sample sizes and different protocols among various studies. Therefore, in the present analysis, we performed a wide-ranging online search according to strict inclusion and exclusion criteria, to draw a clear and systematic conclusion.

Data from 29 trials [7,18–45] including 2488 patients with advanced HC were included in our meta-analysis. JLC in all the included studies was manufactured by Beijing Jiansheng Pharmaceutical Co., Ltd. The dosages of JLC were 2.0–3.0 g per day via oral administration. The pooled results revealed that the combination of JLC and conventional treatment for HC achieved more beneficial effects compared with those treated solely with conventional therapy. Compared with conventional treatment alone, JLC could significantly improve ORR, DCR and QoL in patients with HC (P<0.05). The study also assessed whether JLC could prolong the long-term survival rates of HC patients, and the results showed that the 6-, 12-, 24- and 36-month OS of patients were significantly prolonged compared with the control group. These results indicated that using JLC could improve the short- and long-term curative effects of conventional treatment for advanced HC. However, the analysis of therapeutic effects may have been influenced by several factors. Therefore, we performed a subgroup analysis to determine the influence of different therapeutic regimens, sample sizes, study type and evaluation criteria on ORR and DCR. Subgroup analysis revealed that the therapeutic efficacy of JLC did not appear to be affected by these factors. However, there were limited studies and insufficient sample sizes for these investigations, which may have resulted in insufficient assessment. Therefore, these results need to be confirmed by new evidence.

The immunosuppressed status of cancer patients has been reported previously [2]. Therefore, immune system reconstruction is one of several critical aspects in effectively treating malignancies. Many studies have reported that JLC can enhance the ability of the body’s immunity and resistance to tumors [12]. Our analysis demonstrated that the percentages of CD3⁺, CD4⁺ and NK cells, and CD4⁺/CD8⁺ ratio, were significantly increased in HC patients treated with JLC, indicating that immune function in HC patients was improved after JLC adjuvant therapy.

Safety is the top priority of clinical treatment. Although JLC is cytotoxic to tumor cells, it may also be toxic to normal cells. As such, we analyzed 16 trials [18,20,26,28–30,32,33,36,38–44], involving 1192 patients, to evaluate adverse events according to World Health Organization standards. Meta-analysis revealed that patients who underwent JLC plus conventional treatment demonstrated a lower risk for leukopenia, hepatotoxicity and myelosuppression compared with conventional treatment alone, whereas analysis of other toxic side effects did not differ significantly. Therefore, JLC appears to be a safe auxiliary anti-tumor medicine for individuals with HC.

There were some limitations to our analysis. First, as an important Chinese patent medicine, JLC is mainly applied in China, which may lead to unavoidable regional bias. Second, several results demonstrated significant heterogeneity among the included trials, which may be due to the different tumor stages and ages of the HC patients and duration of treatment. However, based on the currently available literature, there are insufficient data to perform more statistical analysis to evaluate correlations. Third, our results may have inherent bias due to unclear randomization methods, allocation concealment and blinding in some of the included trials. Finally, there were limited studies and sample sizes for immune function and safety evaluation; therefore, analytical bias is possible. All these limitations may have resulted in insufficient evaluations of the outcome indicators.

In summary, findings of this meta-analysis indicate that the combination of JLC and conventional treatment is effective in treating patients with advanced HC. The clinical application of JLC not only clearly enhanced the therapeutic effects of conventional treatment, but also effectively improved QoL and immune function in patients with HC. Thus, we anticipate that our study will provide valuable evidence for further evaluation of JLC. On the other hand, the low quality of some of the included publications increased the risk of bias, which, to some extent, affects the reliability of this research. Therefore, additional studies with high-quality evidence to verify the effectiveness of JLC-mediated therapy for HC are warranted.

C.W.D. and H.X. conceived and designed the methods, extracted the original data and drafted the manuscript. H.X., W.W. and M.Y. performed statistical analysis. H.X. and C.W.D. interpreted the results. C.W.D. revised the manuscript. All authors had full access to all data in the study and take responsibility for the integrity of the data and the accuracy of data analysis.

The authors declare that there are no competing interests associated with the manuscript.

This work was supported by the Medical Science Foundation of Shandong, China [grant number 2014WS0300].

Bcl-2

B-cell lymphoma-2

BNIP3

B-cell lymphoma-2/adenovirus E1B nineteen kilodalton interacting protein-3

CI

confidence interval

DCR

disease control rate

HC

hepatocellular carcinoma

JLC

Jinlong Capsule

KPS

Karnofsky Performance Score

NK

natural killer

OR

odds ratio

ORR

overall response rate

OS

overall survival

QIR

quality of life improved rate

QoL

quality of life

SFDA

State Food and Drug Administration