Negative association between antibiotics and clinical activity in patients with advanced gastric cancer

Antibiotics (ATBs) induce dysbiosis of the gut microbiota by altering the diversity and composition of the microbiota, which mediates the ecacy and toxicity of cancer therapy. The inuence of dysbiosis induced by ATB administration on primary resistance to chemotherapy in patients with advanced gastric cancer (GC) has rarely studied. We evaluated the effect of ATB administration on chemotherapy ecacy in patients with advanced GC.


Abstract
Background Antibiotics (ATBs) induce dysbiosis of the gut microbiota by altering the diversity and composition of the microbiota, which mediates the e cacy and toxicity of cancer therapy. The in uence of dysbiosis induced by ATB administration on primary resistance to chemotherapy in patients with advanced gastric cancer (GC) has rarely studied. We evaluated the effect of ATB administration on chemotherapy e cacy in patients with advanced GC.

Methods
Patients with GC were divided into two groups according to the statues of ATB administration: ATBtreated and control groups. Tumor responses, progression-free survival (PFS), and overall survival (OS) were assessed.

Conclusions
ATB administration is associated with reduced chemotherapy e cacy and poor prognosis in patients with advanced GC. Modulations in ATB-related dysbiosis and gut microbiota composition improve the clinical outcomes of chemotherapy.

Background
Gastric cancer (GC) is the fourth most common cancer and second leading cause of cancer mortality worldwide. In China, nearly one million individuals are diagnosed annually (1). Most patients present with inoperable advanced or metastatic disease with a 5-year survival rate of 5-20% and median overall survival (OS) of less than 1 year (2). The general treatment for advanced GC is a uoropyrimidine-based and platinum-based combination with or without a third drug, such as docetaxel or epirubicin.
The microbiota participates in host metabolism, immunomodulation, neuronal development, and maintenance of the gut mucosal barrier (3). Gut bacteria modulate the pharmacological effects of chemotherapeutic agents (5-uorouracil, cyclophosphamide, irinotecan, oxaliplatin, gemcitabine, and methotrexate) through a mechanistic framework as follows: translocation, metabolism, immunomodulation, enzymatic degradation, reduced diversity, and ecological variation (4,5). Antibiotics (ATBs) induce microbiota dysbiosis characterized by the loss of distinct species and expansion of pathogens by initiating changes in bacterial metabolites, disruption of bacterial signals, dysregulation of gut immune cells, and dysfunction of systemic immunity (6). In the present study, we explored the possibility that dysbiosis induced by ATB administration in uences primary resistance to chemotherapy in patients with advanced GC.

Patients
Patients with advanced GC, including 80 male and 19 female patients who received palliative chemotherapy at the Fourth Hospital of Hebei Medical University between March 1, 2011 and July 15, 2019, were enrolled in this study ( Table 1). All patients received chemotherapy alone or in combination with other treatments, including radiation therapy, intervention, or targeted therapy. All procedures were supervised and approved by the Ethics Committee of the Fourth Hospital of Hebei Medical University (2020KS001). Written informed consent was obtained from the participants.
Medical records of all patients were reviewed to collect data regarding the presence, speci c time, class, indications, route, and duration of ATB administration (Table S1). Patients who were administered ATBs within 30 days after initiating chemotherapy were de ned as the ATB-treated group, and the others were de ned as the control group. Clinical characteristics including age, sex, tumor location, histological type, performance status of the Eastern Cooperative Oncology Group, detailed treatment regimen, and metastatic status were also collected.
Computed tomography and gastroscopy (if necessary) data were reviewed regularly to evaluate the tumor response according to the Response Evaluation Criteria in Solid Tumors version 1.1 (7). All patients were followed up every three months until death or until the database was closed (July 15, 2019).

Statistical analysis.
Clinicopathological features and tumor response were compared using Fisher's exact test or the Chisquared test. Progression-free survival (PFS) was de ned as the time interval from the initiation of chemotherapy to the date of disease progression (PD). OS was de ned as the time interval from the initiation of chemotherapy to the date of death. The PFS and OS curves were estimated using the Kaplan -Meier method and compared using the log-rank test. The Cox proportional hazard model was used to calculate the hazard ratios (HRs) and 95% con dence intervals (CIs) in univariate and multivariate analyses. All statistical analyses were performed using SPSS statistical software, version 19.0 (IBM Corporation, Armonk, NY). p<0.05 was considered to indicate statistically signi cant differences.

Results
A total of 99 patients with metastatic GC with the main histological subtype of adenocarcinoma (93.94%) were enrolled in this study (Table 1). Of these, 68 (68.69%) patients received a uoropyrimidine-based combination with platinum therapy, whereas 31 (31.31%) received paclitaxel alone or combined with other drugs, including uoropyrimidine and platinum (Table 1). There was no major statistical difference in the baseline clinicopathological characteristics between the ATB group and control group with the most frequently administered ATB of β-lactam ± inhibitors ( Table 1, Table S1).
In patients with GC, recent ATB administration was associated with an increased PD rate (72.73% vs. 29.55%, p=0.007) compared to that in the control group (Figure 1c). In addition, ATB administration was associated with both shorter PFS (  Figure 1a and 1b).
We next investigated the impact of ATBs on PFS and OS within individual subgroups of patients based on strati ed analysis. As noted in Figure 2, the ATB group displayed a trend of reduced PFS and OS within nearly every subgroup.

Discussion
We investigated the potential clinical impact of ATB use on chemotherapy e cacy in patients with advanced GC, and we found that ATB administration was associated with decreased PFS from 1.47 to 4.97 months and OS from 9.97 to 13.3 months. These decreases are explained at least partly by the higher PD rate in the ATB group. Almost 10% of patients with cancer undergoing chemotherapy require ATB treatment due to the immunosuppressive nature of malignancies and lymphodepletion induced by anticancer therapies (8,9). Recent studies support the viewpoint that the microbiota regulates the e cacy of anticancer therapy and that the corresponding microbiota targeting may change the drug e cacy (10).
Our data demonstrated this viewpoint in patients with advanced GC that ATB modulates the chemotherapy e cacy by changing the equilibrium of the gut microbiota to modify the GC outcomes.
The mechanism by which ATB -induced loss of diversity of the microbiota modulates the chemotherapy e cacy remains unclear. The chemotherapy e cacy of oxaliplatin or cisplatin dramatically decreases in germ-free mice due to reduced oxaliplatin-induced DNA damage (11). The absence of microbiota decreases reactive oxygen species production and attenuates oxaliplatin-induced DNA damage by tumor-in ltrating myeloid cells via reduced nicotinamide adenine dinucleotide phosphate oxidase 2 (11). In addition, the microbiota modulates the adaptive immune response for immunogenic cancer cell death induced by oxaliplatin (12). Furthermore, ATBs downregulate BAX and cyclin-dependent kinase inhibitor 1B, while upregulating vascular endothelial growth factor, thereby attenuating the e cacy of cisplatin in a lung cancer mouse model (13). Regarding the chemotherapeutic agent 5-uorouracil, ATB administrationrelated microbiota dysbiosis with increased pathogenic bacteria Escherichia shigella and Enterobacter impair its e cacy by downregulating the expression of genes involved in amino acid metabolism (14). The fact that the chemotherapeutic agent itself exacerbates the dysbiotic state renders the ATB-related dysbiosis di cult to treat.
The gut microbiota exert their effect on chemotherapy e cacy through medicating chemical signaling transduction (4). Some probiotic bacterial species may promote antitumor effects, as well as prevent the toxic side effects of chemotherapeutic drugs (13). The ATB-induced chemotherapy ine cacy of cisplatin may be rescued by concomitant Lactobacillus bacteria (13,15). These results imply that ATB-related dysbiosis may be neutralized with speci c probiotic bacterial species. Longitudinal chemotherapy studies focusing on host microbiota interactions using metagenomic, metabolomic, and metatranscriptomic approaches are needed to identify probiotic bacterial species for treatment of ATB-induced dysbiosis.
Further studies including clinical trials with larger sample sizes are warranted to con rm the deleterious effects of ATBs on chemotherapy. The identi cation of the key microbiota species capable of neutralizing unfavorable ATB-associated dysbiosis will help build future therapeutic concepts, whereby probiotic bacterial species-related modulation of the gut microbiota may increase chemotherapy e cacy Conclusions ATB administration is associated with reduced chemotherapy e cacy and poor prognosis in patients with advanced GC. Modulations in ATB-related dysbiosis and gut microbiota composition improve the clinical outcomes of chemotherapy.

Declarations
Ethics approval and consent to participate All procedures were supervised and approved by the Ethics Committee of the Fourth Hospital of Hebei Medical University (2020KS001). Written informed consent was obtained from the participants.

Consent for publication
Not applicable.

Availability of data and materials
Research data supporting the results of this paper will be provided by corresponding author at reasonable request.

Competing interests
The authors declare that they have no competing interests.

Funding
The Natural Science Foundation of China of Hebei Province (Grant No. H2019206428) kindly provided research grants for our study.