Introduction
Colorectal cancer (CRC) is the third most common cancer in the world and it is one of the major causes of death and morbidity [
1]. Among the causes of colorectal cancer development, the so-called driver mutations such as that of
APC,
KRAS,
BRAF,
PIK3CA,
SMAD4, and
TP53 play significant roles. Huang et al. found that mutations of
KRAS,
TP53,
SMAD4, and
BRAF were associated with CRC metastasis and may be both potential biomarkers of metastasis as well as a therapeutic target in CRC [
2].
The gut microbiota plays an important role in the development and progression of colorectal cancer. Several plausible mechanisms have been proposed for intestinal microbiota to bind to colorectal cancer cells such as inflammation, DNA damage effects, and non-DNA damage effects, all of which may be mechanically important [
3]. There is increasing evidence that the gut microbiota and its products are linked to CRC. For example, it was reported that the existence of
Bacteroides and
Bifidobacterium species correlates positively with a high risk of CRC whereas the existence of
Lactobacillus species and
Eubacterium aerofaciens correlate negatively [
4,
5]. According to some reports in the literature, the existence of
Clostridium nuclei,
Streptococcus haemolyticus,
Bacteroidetes fragilis enterotoxin,
Enterococcus faecalis, and
Escherichia coli have been identified to be associated with the development of CRC [
6,
7]. Also, the bacterial driver-passenger model could explain the microbial involvement in CRC development. In this model, a driver bacterium initiates CRC development by a transient colonization upon which it is replaced by a passenger bacterium displaying a competitive growth advantage in the tumor niche [
8]. The candidate driver bacteria showed pre-carcinogenic characteristics such as the production of DNA damaging compounds, the disruption of tumor suppressor protein function, and induction of a host inflammatory response [
9]. The identification of driver and passenger bacteria can therefore be used as classifiable biomarkers to detect high-risk groups or patients with CRC, respectively [
10].
In recent years, the development of high-throughput sequencing technologies was a major step in cataloging the intestinal microbiome. More than 1,000 microbial species have been identified in the human gastrointestinal tract by analysis of the small subunit ribosomal RNA gene sequence. Most studies have focused on fecal samples to understand the composition of the gut microbiome during the development and progression of CRC. However, limited studies have assessed the association of intestinal microbial richness and biodiversity with driver gene mutations. In this study, we evaluated changes in the microbiome of CRC patients and healthy controls and explored driver gene mutations in CRC patients by comparing them to the wild-type.
Discussion
CRC is the third most common malignant tumor in the world and the second most common cause of malignant tumor-related death (about 9.2% of the total number of malignant tumor deaths), ranking first in all gastrointestinal malignancies [
13]. Diet, lifestyle, and host genotype may be involved in the occurrence and development of colorectal cancer through metabolic and inflammatory mechanisms [
14]. Genes are known to regulate the pathogenesis of CRC and are associated with the survival outcomes of patients. Among these genes are
KRAS,
TP53,
APC,
SMAD4,
BRAF, and
PIK3CA [
2]. Gurjao et al. found that overeating of red meat leads to CRC by altering
KRAS and
PIK3CA and the alkylation state of these genes [
15]. In our study, the driver gene mutations were mainly distributed among
APC,
TP53,
KRAS, and
PIK3CA. APC inhibits Wnt signaling by promoting phosphorylation and degradation of β-catenin. In intestinal stem cells (ISCs), loss of
APC function mutations drives intestinal adenomas by enhancing intracellular Wnt signaling [
16]. Brandt et al. found that oncogenic
KRAS, together with β-Catenin, favoured the expansion of crypt cells with high ERK activity [
17].
At present, molecular targets of colon cancer comprise EGFR, VEGF, ERBB2, BRAF, KRAS, PD-1, CTLA-4, NTRK etc. The targeted therapy of CRC patients displaying EGFR and EGFR-related pathway gene mutations can be divided into those using monoclonal antibodies and those using small molecule tyrosine kinase inhibitors (TKIs). The used monoclonal antibodies comprise anti-EGFR monoclonal antibodies such as cetuximab and panizumab, anti-HER2 monoclonal antibodies such as pertuzumab and trastuzumab, and the Insulin like growth factor 1 receptor (IGF1R) inhibitor such as darlozumab. TKIs include BRAF inhibitors such as vimofinib, Darafenib, and encofanil, MEK inhibitors comprise trametinib, cobitinib, bemitinib, and Selumetinib, and eventually, the HER2 inhibitor lapatinib (dual EGFR and HER2 targeting), etc. Cetuximab can bind to EGFR on the surface of tumor cells, competitively blocks the EGFR signaling pathway and inhibits the proliferation of tumor cells. Cetuximab is widely used in the treatment of CRC patients with KRAS/NRAS/BRAF wild-type genome. Currently, the treatment of patients with KRAS mutation is the focus of targeted therapy. Sotorasib has been approved for the treatment of NSCLC with KRAS mutations by the FDA. The CodeBreak 101 study showed that the objective response rate (ORR) of Sotorasib combined with panitumumab achieved 27% in patients with advanced/metastatic CRC with a KRAS G12C mutation and the disease control rate (DCR) reached 81%.
Similar to key metabolic and immunomodulatory agents, the intestinal flora is believed to play an important role in the development of colorectal cancer [
18]. There is a rich microbiota in the colon lumen. Previous studies have shown that dysbiosis and imbalance in the gut microbiome can mediate or alter the impact of environmental factors on CRC risk [
19,
20]. In this study, we depicted the overall composition of the gut microbiota by 16S rRNA sequencing, demonstrating that microbial dysbiosis is characterized by a distinct microbial composition and altered relative abundances of species with specific functions. Compared to healthy controls, the CRC patient group displayed a decreased microbial diversity and an increased microbial richness which is consistent with arguments outlined in previous reports [
21‐
23]. Additionally, our study could show that at the phylum level,
Bacteroidetes and
Proteobacteria were enriched and at the genera level,
Bifidobacterium,
Shigella Escherichia coli and
Bacteroides were more abundant in the CRC group. This is in accordance to previous studies that showed a higher abundance of Bacteroidetes and Proteobacteria in the CRC patients [
24]. The relationship between
Fusobacteria and CRC was studied on a large mass cohort comprising 3,157 individuals, including CRC patients and healthy controls, and revealed that
Fusobacterium varium and
Fusobacterium ulcerans were related to a homologue of FadA adhesin [
25]. Ma et al. reported that colorectal tumor apoptosis is induced by sitosterol through promoting gut microbiota to produce SCFAs which display antiproliferative effects on human colorectal cancer cells via gene expression inhibition [
26]. Also, the intestinal flora may promote serrated lesions through EGFR signaling, the induction of cellular proliferation, the activation of a tumor immunosuppressive microenvironment, and the induction of an inflammatory response [
27]. ROC analysis suggested that
Faecalibacterium,
Collinsella, and
Bacteroides may be potential biomarkers for CRC.
Host genes can also regulate the growth of microbiota and influence the composition of the intestinal microbial community. Thus, to achieve a comprehensive analysis, we investigated the association between host gene mutations and microbial composition in CRC patients. The most significant associations of host gene mutations with gut bacterial composition were a higher abundance of Faecalibacterium and lower abundance of Bifidobacterium in patients with KRAS mutations, a higher abundance of Eubacterium coprostanoligenes in TP53 mutated patients, and a higher abundance of Lactobacillus in PIK3CA mutated patients compared to patients without these gene mutations. This suggests that host driver mutations affect the gut microbiota composition in CRC patients. We will explore the relationship between the effect of targeted therapy and the intestinal flora in these patients with gene mutations.
A previous CRC study reported that
Faecalibacterium was enriched in the survival group whereas
Fusobacterium nucleatum and
Bacteroides fragilis were more abundant in the worse prognosis groups which is consistent with our findings, at least to some extent [
28]. Specifically,
Faecalibacterium displays an anti-inflammatory effect and its metabolites block the activation of NF-kB and the secretion of IL-8 [
29]. Interestingly,
KRAS mutations happen to be more common in colorectal serrated adenocarcinoma that involves the NF-kB pathway and the secretion of IL-8 which may account for the association of the higher abundance of
Faecalibacterium in
KRAS mutated CRC patients [
30,
31]. Moreover,
Lactobacillus was more abundant in CRC patients with
PIK3CA mutations in our study.
Lactobacillus inhibits the growth of colorectal cancer by secreting short-chain fatty acids (SCFAs) that enhance the intestinal barrier function [
32]. Taken together, our findings are supported by the above-mentioned reports, which highlight the clinical relevance of
Faecalibacterium and
Lactobacillus in the development of CRC and suggest their potential to become a therapeutic agent by producing bioactive compounds that may benefit the host. We hypothesized that this relationship between host driver mutations and intestinal flora might play a pivotal role in the pathogenesis of CRC and provided a new idea for the treatment of CRC in the future. However, further detailed studies are needed to confirm our findings and to investigate the molecular mechanisms of host driver mutations and their effects on gut microbiota in CRC patients.
Although our study indicated an association of the intestinal flora composition with gene mutations such as KRAS and TP53, our results have some limitations. In this analysis, the number of patients included was not large enough. Besides, only a small panel of 18 genes was tested in this study. We expect that a larger panel of NGS parameters as well as a larger sample size will hopefully lead to more discoveries in the future. Combined analysis of multi-omics data, such as transcriptome and methylome, must be conducted for further experimental studies and clinical trials to validate and reinforce our findings.
In conclusion, we revealed a potential relationship between the host genome and the gut microbe composition in CRC patients. The results of the gene and intestinal flora analyses provided a clearer understanding of the pathogenesis of colorectal cancer. The correlation between relative bacterial abundance and host gene mutations suggests that intestinal microbiota may influence the growth of mutated cells and that their dysregulated cellular pathways influence the abundance of specific bacteria. Thus, our study might provide a new perspective therapeutic approach for the treatment of CRC.
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