Gastrointestinal (GI) cancers, including gastric cancer, colon cancer, and colorectal cancer (Soleimanpour et al., 2020), are among the most common cancers (Wang et al., 2020a), accounting for approximately 26% of total cancer incidence and about 36.4% of cancer-related deaths (Arnold et al., 2020). Recently, the incidence and mortality of GI cancers have been increasing (Bray et al., 2018), so exploring the protective factors and risk factors for the occurrence and development of GI cancers will be conducive to effectively preventing and treating these cancers. Clinically, GI cancers are usually treated by radiotherapy, chemotherapy, surgery, drugs, and immunotherapy, while TCM, generally considered as an adjuvant therapy combined with radiotherapy and chemotherapy, plays an effective anti-tumor role by inducing tumor cell apoptosis and inhibiting tumor angiogenesis (Wang S. et al., 2021). At the same time, it decreases the gastrointestinal reactions caused by radiotherapy and chemotherapy (Zhang et al., 2021). However, with the in-depth study of the relationship between TCM and intestinal flora and gastrointestinal cancers, we found that TCM can adjust intestinal flora, promote beneficial bacteria to produce more Short-chain fatty acids (SCFAs) (Martin-Gallausiaux et al., 2021), mainly including acetate (C2), propionate (C3) and butyrate (C4), and improve the microenvironment of the gastrointestinal tumors, thereby having a certain beneficial impact on the occurrence, development, and prognosis of GI cancers (Sivan et al., 2015). In addition, intestinal flora also has a therapeutic effect on radiation enteritis caused by radiotherapy (Jian et al., 2021). Therefore, we believe that TCM can treat patients with gastrointestinal cancers by regulating intestinal flora in multiple ways.

Human intestinal microbes constitute a complex ecosystem, with around 800 species and more than 7,000 bacterial strains (Ley et al., 2006). In the intestine, symbiotic microorganisms are dynamic, which can maintain intestinal stability and inhibit pathogen colonization. When the balance is broken, the intestinal mucosal barrier and immune function will be undermined, leading to additional pathogenic factors, which are risk factors for colorectal cancer as well (Si et al., 2021).Clinical studies have found significant changes in the structure and characteristics of the intestinal flora in gastrointestinal cancer patients (Ferreira et al., 2018). Additionally, intestinal flora affects the absorption of anticancer drugs and correlates with the prognosis of these patients (Wertman et al., 2021). The pathological mechanisms by which intestinal flora affects colorectal cancer are currently thought to be achieved through multiple pathways, such as the induction of inflammation and immunity (Meng et al., 2018). Notably, intestinal pathogenic bacteria can drive tumorigenesis by shaping the tumor microenvironment or forming biofilms, such as Bacteroides, Escherichia coli, and Clostridium difficile, which can secrete a variety of virulence factors that damage intestinal epithelial cells and trigger chronic inflammatory responses, and develop into colorectal cancers (Hayase and Jenq, 2021). Meanwhile, some intestinal probiotics can directly produce tumor suppressive substances or enhance related antigens to achieve anti-tumor effects (Song et al., 2021). The ferritin produced by Lactobacillus casei ATCC334, for instance, can act as a tumor suppressor through the JNK signaling pathway (Konishi et al., 2016). Therefore, we consider how to balance the environment of intestinal microbiota deserves further exploration.

Among many pathogenic bacteria, Escherichia coli and Enterococcus, belonging to neutral bacteria, are not pathogenic when their population is within a certain range, however, an excessive number of these bacteria may produce Enterotoxin that are highly pathogenic (Wassenaar, 2018; Alhinai et al., 2019). Bifidobacteria and Lactic acid bacteria, as probiotics, produce a large amount of SCFAs, which are beneficial to intestinal health (Zaharuddin et al., 2019). Because of their large number, more in-depth basic research, and easy clinical detection, when intestinal pathology changes, the flora changes significantly, so they are commonly used as clinical indicators for evaluation of intestinal flora (Kuugbee et al., 2016).

In fact, the TCM adjuvant therapy for cancer has achieved a remarkable clinical efficacy (Wang S. et al., 2020c). Nowadays, the main TCM therapies commonly used in the clinics are oral therapy, external therapy and injection therapy (Huang et al., 2018). The classification is based on different routes of administration. Oral treatments of TCM are absorbed through the gastrointestinal tract, external treatments of TCM are absorbed through the skin and mucosa by physical therapy or enema, and TCM injections are the components directly enter the bloodstream. To the best of our knowledge, most of the previous studies have focused more on TCM oral administration, and less on external treatments and TCM injections. It has been shown that TCM can inhibit the development of cancer by regulating intestinal microbes (Chen et al., 2021). XiaoYao decoction (a medicinal diet with Ginseng, Atractylodes and Fushen as the main ingredients), for instance, can increase the abundance of Bacteroides, Lactobacillus, and Proteobacteria, and reduce the abundance of Desulfovibrio and Rickerella (Zhang Z. et al., 2020).

Given that the evidence in the current literature not able to determine which one is the most effective Therefore, we tried to select the best treatment by counting and analyzing the changes of 4 indicators in intestinal flora after the application of three TCM treatments in the previous literature. To date, no meta-analysis has been conducted to compare the effects of CT in combination with each of these three TCM methods on intestinal flora in gastrointestinal cancer patients. We present the paper on the basis of the checklist of the extended PRISMA for network meta-analysis.

This meta-analysis was registered in PROSPERO (CRD42022332553).

A total of 20 RCTs with 1774 patients was included in this paper. Through the comparison of SUCRA results, Aidi injection is the most effective in increasing the number of Bifidobacteria and Lactobacillus, and in inhibiting the number of Escherichia coli and Enterococcus. In the pairwise comparison of three TCM treatments, injection of TCM plus CT or oral Chinese medicine plus CT are effective for Bifidobacteria and Lactobacillus. For Escherichia coli, TCM injection plus CT takes effect. For Enterococcus, TCM oral treatment plus CT is practical.

There are three treatments of TCM, including internal treatment (oral absorption), external treatment (physical therapy or skin mucosal absorption), and injection (direct blood injection). Internal treatment mainly uses oral Chinese medicine decoction, the preparation of which is to soak the traditional Chinese medicine in boiling water or hot water to produce an aqueous extract containing a mixture of chemical components (Zhou et al., 2016; Deng et al., 2019). Specifically, after oral administration of TCM into the colon, intestinal microbiota converts carbohydrates, proteins, lipids and small non-nutritive compounds from TCM into chemical metabolites that may have beneficial or adverse effects on human health (Wang et al., 2013). For example, the continuous digestion of polysaccharides and carbohydrates (PS) produces many short-chain oligosaccharides, which can promote the growth of probiotics such as Bifidobacteria and Bacteroides. Shorter PSs are digested to form monosaccharides, which can be continuously catabolized to form short-chain fatty acids (SCFA) (e.g., formate, acetate, propionate, butyrate), lactic acid, hydrogen, carbon dioxide and other metabolites. Valproic acid, a kind of SCFAs, has antitumor activity, and its main mechanism is to inhibit histone deacetylase (Gurvich et al., 2004). These metabolites may directly affect the host intestinal environment and improve the microenvironment of gastrointestinal cancer (Vernocchi et al., 2016; Feng et al., 2019).

Aidi injection is mainly composed of ginseng, Astragalus membranaceus, canthatis, and acanthopanax senticosus. The active components are ginsenoside, astragalus polysaccharide, astragalus saponin, cantharidin and Acanthopanax Senticosus Polysaccharide (Quirke et al., 2007). Astragalus polysaccharides can increase the number of lactic acid bacteria and Bifidobacteria, thereby reducing pro-inflammatory factors, such as interleukin-6 and tumor necrosis factor-α. Therefore, as an inflammatory response inhibitor, it can also reduce the inflammatory response by reducing Salmonella typhi in the intestine (Tang et al., 2021). Ginsenoside-rb3 and ginsenoside Rd can promote the growth of beneficial bacteria, such as Bifidobacterium, Lactobacillus, Acidophilus and Anisoid, and can also reduce a number of cancer-related pathogens and Helicobacter pylori spp to prevent the development of colorectal cancer (CRC) (Huang et al., 2017). There are few studies on other drugs. We attribute the better effect of Aidi injection to its high bioavailability compared with the other two methods. When it comes to the cold and hot nature of the drug, all four drugs in the prescription are warm products. Therefore, it can be inferred that Aidi injection is hot and may be more suitable for the body of cancer patients undergoing radiotherapy and chemotherapy. External treatments include enema, acupoint embedding, skin scratching and ginger moxibustion. These methods are rarely studied in the field of intestinal flora research.

Many studies have shown that there is a causal relationship between changes in the intestinal flora and colorectal cancer. Patients with colorectal cancer have poor nutritional status and low systemic and partial resistance, which inhibit the growth of intestinal dominant bacteria, such as Lactobacillus and Bifidobacterium, resulting in the imbalance of the intestinal microenvironment. Meanwhile, intestinal flora imbalance will decrease the immune function of the body, and the decline of immune function will aggravate the flora imbalance, thus forming a vicious circle. In addition, chemotherapy drugs further reduce the immunity of patients and interfere with the proportion of normal intestinal flora. Moreover, the more obvious the imbalance is before chemotherapy, the more serious the imbalance is after chemotherapy. Therefore, the anti-cancer research of intestinal flora is of great significance. Lactic acid bacteria and Bifidobacterium strains induce dendritic cell (DC) to mature (Hickey et al., 2021) and produce IFN- γ (IFN- γ), enhancing the cytolytic potential of NK cells (Zhou et al., 2019). Probiotics induce apoptosis by inhibiting the expression of COX-2, NF KB, and MAPK, suppressing the inactivation of inflammatory bodies, and activating Caspase-3 (Iyer et al., 2008). It also induces cell death through autophagy (Engevik et al., 2019). Bifidobacterium can increase anti-PD-L1 and inhibit tumor volume by inducing anti-inflammatory activity of macrophages and dendritic cells (Xu et al., 2020). Probiotics such as Lactobacillus and Bifidobacterium inhibit the growth of colorectal cancer by suppressing inflammation and angiogenesis, and enhance the intestinal barrier function by secreting short-chain fatty acids (SCFAs) (Koh et al., 2016). Escherichia coli is more prevalent in colorectal cancer tissues (Buc et al., 2013). Enterococcus faecalis produces enterotoxins (e.g., tartary buckwheat glucoside) and reactive oxygen species, which can lead to DNA oxidative damage and intestinal epithelial cell inflammation (Baldassarri et al., 2005). Enterococcus faecalis is responsible for producing reactive oxygen species and superoxide anions, resulting in DNA damage and genomic instability in colorectal cancer (Geravand et al., 2019). Fecal Escherichia coli induces mucosal macrophages to produce DNA damage inducers (Goodwin et al., 2011), such as 4-hydroxy-2-nonyl, through COX-2 (Yang et al., 2013).

We did a general meta-analysis, separating the patients with gastric cancer and colorectal cancer, to compare the differences between CT only and the three methods combined with CT. For colorectal cancer patients, in Bifidobacteria and Lactobacillus, external treatment plus CT or injection of TCM plus CT, are more effective than CT. For Escherichia coli, the TCM external treatment plus CT and the TCM injection plus CT are more effective than CT only. For Enterococci, We prefer TCM injection plus CT because it exhibits better effect than CT only.

The following limitations should be considered in this study. The methodological quality of the effect of TCM on intestinal flora of gastrointestinal cancers is subject to some risk deviation, such as insufficient sample size and short duration. In addition, gastrointestinal cancers are subdivided into gastric and colorectal cancers. Because the number of articles is too small to perform a heterogeneity testing, we did a general meta-analysis to assist the results. TCM is divided into three categories. The external treatment includes enema, the effect of which overlaps with that of oral TCM. However, considering its direct effect on intestinal flora and its short duration, it is placed in the external treatment.

In this study, Bifidobacteria and Lactobacilli, Escherichia coli and Enterococci were used as the main therapeutic indicators for comprehensive evaluation. Overall, TCM injection may be the best treatment, followed by TCM external treatment. TCM plays a certain role in the intestinal flora of patients with gastrointestinal cancers through multi-targeted comprehensive intervention. Clinically, it can be used in combination with other therapies depending on the actual situation of patients, and is suitable for the whole treatment process for gastrointestinal cancer patients.