Abstract
Microbiome and radiotherapy represent bidirectionally interacting entities. The human microbiome has emerged as a pivotal modulator of the efficacy and toxicity of radiotherapy; however, a reciprocal effect of radiotherapy on microbiome composition alterations has also been observed. This review explores the relationship between the microbiome and extracranial solid tumors, particularly focusing on the bidirectional impact of radiotherapy on organ-specific microbiome. This article aims to provide a systematic review on the radiotherapy-induced microbial alteration in-field as well as in distant microbiomes. In this review, particular focus is directed to the oral and gut microbiome, its role in the development and progression of cancer, and how it is altered throughout radiotherapy. This review concludes with recommendations for future research, such as exploring microbiome modification to optimize radiotherapy-induced toxicities or enhance its anti-cancer effects.
The association between microbial dysbiosis and cancer has gained extensive attention and is being widely explored. Since the discovery of Helicobacter pylori in the 1980s, many other cancerogenic bacterial, viral, protozoal, and parasitic entities have been successfully identified. The microbiome has also been closely linked to the treatment response to chemotherapy and immunotherapy; however, its association with radiotherapy has been ambiguous for a long time.
Microbiome and radiotherapy represent bidirectionally interacting entities that can, directly and indirectly, affect each other. Dysbiosis was observed to affect development, characteristics, progression, and even radiotherapy response and toxicity in cancer. Radiotherapy, on the other hand, has a major impact on the diversity and incidence of microbial taxa in the irradiated and even non-irradiated field. The exact mechanisms are, however, still unknown. In this review, we aimed to provide a complex state-of-the-art insight into the problematics of radiotherapy and microbiome across various cancer types.
Methodology
Eligibility criteria. For the use of this review, only extracranial cancers routinely treated with radiotherapy were included. By consensus of the authors (MP, ML, RS), head and neck cancer (HNC), lung cancer, esophageal cancer, gastric cancer, pancreatic cancer, prostate cancer, gynecological cancers, colorectal cancer, and urinary bladder cancer were chosen for further investigation.
Search strategy. In this systematic review, PubMed, Web of Science and Cochrane Central were used to search for published literature on the effect of cancer radiotherapy on humane microbiome. Two independent investigators (MP and ML) conducted the comprehensive literature review. The principal search terms included “radiotherapy”, “chemoradiotherapy”, and “microbiome”. These terms were subsequently paired with terms “head and neck cancer”, “lung cancer”, “esophageal cancer”, “lung cancer”, “gastric cancer”, “pancreatic cancer”, “skin cancer”, “breast cancer”, “prostate cancer”, “gynecological cancer”, and “pelvic radiotherapy” for the database searches.
1,452 publications were identified and automatically screened for duplicates; 974 repetitive sources were excluded. Abstracts of 478 publications were read, subsequently, 286 publications were excluded. One hundred and ninety-two full-text articles were read, out of which 38 were excluded (mostly research papers investigating cell lines and/or animal models with limited translational possibilities). After filtration, 152 publications emerged. References were supplemented by manual searching in manuscripts references. Search strategy is demonstrated in Figure 1.
Search strategy for the systematic review.
Site-specific Microbiome and its Association With Anticancer Radiotherapy
Head and neck cancer. Head and neck cancer (HNC) represents a heterogeneous group of malignant diseases. It currently accounts for approximately 4% of all malignancies, making it the sixth most common cancer worldwide. A total of 40% of HNCs are located in the oral cavity (1, 2). The most common histological subtype in head and neck cancers is squamous cell carcinoma (represented in ca. 95%) (3).
There was found a direct correlation between oral dysbiosis, in which chronic alcohol intake, smoking, malhygiene, and microbial infections are involved, and tumors of the oral cavity, as well as tumors of distant sites such as the oropharynx, esophagus, pancreas, colorectum, breast, prostate, and lungs, has been reported (4-8). Meta-analyses have shown that dysbiosis significantly increases the risk of oral cancer (9, 10). Among the identified microbial agents whose abundance is significantly higher in the oral cavity of oral squamous cell carcinoma (OSCC) patients are e.g., Fusobacterium spp. such as F. nucleatum, Pseudomonas aeruginosa or Porphyromonas gingivalis (11-13).
Kumpitsch et al. investigated the representation of specific taxa in the saliva of patients with HNC and healthy controls. Congruent to the previous studies, no significant difference in the diversity of microbial taxa in HNC patients and controls was observed; however, significant differences in the abundance of some microbes were reported. Firmicutes was identified as a dominant phylum in HNC patients, especially genera Veillonella, Rothia, Haemophilus, and Neisseria. Surprisingly, the presence of Fusobacterium spp. was significantly correlated with host healthy status, although it is usually linked to cancer development (14). Mäkinen et al. performed an investigation on a larger group of ca. 100 HNC patients and 100 healthy controls, concordant with the Kumpitsch study, they observed a direct association of OSCC on the salivary microbiome, which prevailed significantly, even after removing the cofactors of smoking, alcohol drinking, and being edentate. OSCC did not significantly affect the microbial diversity, but the relative abundance of specific bacteria such as Streptococcus anginosus, Abiotrophia defectiva, and F. nucleatum which was significantly higher in the OSCC cohort, whereas a higher abundance of Prevotella histolitica, Haemophilus parainfluenzae, and F. periodonticum was reported in the healthy cohort (15).
Radiotherapy (RT) plays a key role in therapeutic management in HNC and is used either as a definitive treatment often with concurrent chemotherapy or as postoperative adjuvant treatment (16). RT of HNC is associated with a high incidence of post-RT toxicity, acute as well as late, significantly affecting patients’ quality of life (QoL), e.g., radiation-induced oral mucositis (RIOM), dermatitis, dysphagia, etc. (17-19). The severity of such toxicities is enhanced with concurrent chemotherapy (CRT) (20). High radiation doses on salivary glands can cause a significant decrease in salivary flow and cause xerostomia, which is the most common post-RT complication in HNC patients (21). Xerostomia may lead to impaired QoL of HNC patients, alterations in speech and taste, malnutrition, and because of more acidic intraoral pH, it predisposes HNC patients to develop oral mucositis, dental caries, mucosal ulcerations, and oral infections (22). It has been demonstrated in several studies that interrupted salivary flow leads to shifts in oral microbiome resulting in higher abundances of S. mutans, Lactobacillus spp., Candida spp. and Staphylococcus spp., in contrast, the number of S. sanguinis, Neisseria spp., and Fusobacterium spp. decreases (23). These findings were later supported by Mojdami et al., who reported an increase in the relative abundance of genera Streptococcus, Lactobacillus, Treponema, and Prevotella. Interestingly, these alterations did not recover to baseline within 6 months after RT (24). Then, Kumpitsch et al. described a certain decrease in the diversity of salivary microbiome, which, however, was not statistically significant. In contrast, a significant reduction in the relative abundance of Haemophilus spp., Veillonella spp., and Granulicatella spp. was observed after CRT. Surprisingly, also other genera’s abundance such as Lactobacillus, Scardovia, Acinetobacter, and Enterococcus was increased after the treatment (14).
Moreover, exposure to ionizing radiation leads to changes in the microbiome that can contribute to more severe side effects. Hu et al. demonstrated a dose-dependent shift in oral microbiome after RT. Higher doses were positively associated with increased abundances of Pseudomonas spp., Treponema spp., and Granulicatella spp., on the contrary, an abundance of other microbes such as Prevotella spp., Fusobacterium spp., Leptotrichia spp., Campylobacter spp., Peptostreptococcus spp., and Atopobium spp. were inversely associated (25). Another study demonstrated a direct and significant association between oral dysbiosis and risk of developing radiation induced oral mucositis (RIOM). An increased abundance of several Gram-negative bacteria (Fusobacterium spp., Haemophilus spp., Tannerella spp., Porphyromonas spp., and Eikenella spp.) was observed in pre-treatment buccal mucosa in patients who later developed RIOM of Grade >2 (26). The effect of HNC RT on the salivary and oral microbiome is still to be fully understood; however, based on recent studies, we should stress the impact RT has on carcinogenesis (13, 25, 27, 28). Post-RT dental issues are together with xerostomia the dominant factors which impair the QoL of HNC patients and lead to osteoradionecrosis of the jaw. Regular dental follow-ups are necessary, and specialized dental care should be provided to all HNC patients who underwent both radical CRT or adjuvant RT (17).
Shifts in microbial representation have been shown to affect treatment response to radiotherapy as well as chemotherapy and immunotherapy. Some studies already proposed that the microbiome can be modified to maximize treatment response and minimize adverse effects through the use of probiotics in HNC; however, the oral microbiome in terms of predictive and prognostic biomarkers is yet to be investigated in detail (29-31).
Table I summarizes the currently published articles on the oral microbiome and its association with RT in patients with HNSCC.
Articles investigating oral microbiome and RT of HNSCC patients.
Lung cancer. Lung cancer is the most common malignancy worldwide, sex specifically, it is second only to prostate cancer and breast cancer in men and women, respectively, hand in hand with the leading incidence it is also the leading cause of cancer-related death (32). Although the lung tissue of healthy individuals was long considered sterile, recently, paramount evidence of an existing unique bacterial environment has been reported (33). Thanks to 16S rRNA sequencing, the most common bacteria were described in lung tissue, these include phyla Bacteroidetes, Firmicutes, and Proteobacteria and genera like Streptococcus, Pseudomonas, Veillonella, and Prevotella (34). Researchers investigated possible bacterial dysbiosis in patients with lung cancer. Significantly compared to healthy controls, higher abundance of Granulicatella, Thermus, Legionella, and Streptococcus genera has been reported in the lower respiratory airways of lung cancer patients (35-37). It was proposed that bacteria directly induced carcinogenesis. This postulate has been supported by recent trials demonstrating upregulation of extracellular signal-related kinase (ERK) and phosphoinositol-3-kinase (PI3K) pathways once bacterial oral taxa enriched the lung microbiome, correlation of tumor protein 53 (TP53) mutation and presence of genus Acidovorax in lung niche, the same genus is also significantly higher abundant in smoking patients with squamous cell carcinoma, etc. (38-40). Currently, Pseudomonas spp., Streptococcus spp., Staphylococcus spp., Veillonella spp., and Moraxella spp. are considered the most relevant lung cancer-related microbes (34, 40-56).
A pilot study on a limited number of lung cancer patients who underwent surgical treatment reported specific microbial signatures in healthy lung tissue; however, not in the tumor tissue. Patients who relapsed during the follow-up had greater bacterial richness as well as diversity represented in the healthy lung tissue. Longer disease-free survival (DFS) has been reported in patients whose healthy lung microbiome was abundant in bacteria of the Corinebacteriaceae family, whereas the high abundance of family Lachnospiraceae was associated with shorter DFS. The lung tumor microbiome has been found less diverse compared to the healthy microbiome but constitutionally similar to the healthy lung tissue microbiome, and no significant association between either bacterial diversity or its richness and DFS has been found (49).
For inoperable locally advanced disease the standard of care is radiotherapy with concurrent chemotherapy, up to date no studies investigated shifts in lung or tumor microenvironment after CRT for obvious ethical reasons of obtaining samples either by invasive techniques or BAL (32). Currently, among promising objects of interest belong the identification of radiosensitizing microbiota as promising biomarkers of better treatment effects or possible therapeutics to decrease radiation-induced toxicity (30, 48, 53). Bo et al. investigated the gut microbiome of lung cancer patients undergoing CRT, two distinctive microenvironments were represented and correlated with progression free survival (PFS). Phylum Firmicutes/class Clostridia, especially Dorea spp., was the harbinger of a favorable prognosis (DFS >11 months) (57). These results are concordant with other studies (58). However, the relationship between gut microbiome and radiotherapy of lung cancer remains unclear.
Esophageal cancer. Esophageal cancer is the eighth most diagnosed cancer and the sixth most common cause of cancer-related death worldwide (59). Esophageal cancer represents a group of histologically and etiologically varying entities, the two most common ones are adenocarcinoma and squamous cell carcinoma.
The first attempts to study esophageal microenvironments date back to 1980s; however, these studies focused on perioperative infectious complications rather than on the characterization of the whole microenvironment (60-62). Narikiyo et al. characterized in the early 2000s’ the microbiome of patients with esophageal cancer and the healthy controls, describing the increased abundance of periodontopathic spirochete T. denticola, and bacteria S. mitis, and S. anginosus (63). Almost ten years later, Blackett et al. compared the microbiome of patients with gastroesophageal reflux disease (GERD), Barett’s dysplasia (BD), and esophageal carcinoma with those having asymptomatic GER. Interestingly, Campylobacter spp. has been found significantly more abundant in GERD and BD than in healthy controls (64). Moreover, procarcinogenic cytokines, e.g., IL-18, overexpression has been observed in esophageal tissues colonized by Campylobacter spp. These findings suggested that the role of Campylobacter spp. in esophageal cancer could mimic the role of H. pylori in the development and progression of gastric cancer (65). A comparison of the microbiome of BD and esophageal adenocarcinoma revealed decreased abundances of Veillonella spp. and S. granulosa, while Lactobacillus spp. emerged as the dominant taxa in cancer environment (66, 67).
Currently, no conclusive information about the impact of microbial dysbiosis on the development or progression of esophageal adenocarcinoma is available; however, it is assumed that microbial alterations are involved in GERD/BD progression into adenocarcinoma (68). The relationship between microbiota and squamous cell carcinoma of esophagus is far less characterized compared to adenocarcinoma. Gao et al. observed that P. gingivalis selectively inhabits cancerous and adjacent esophageal mucosa, whereas not the mucosa of healthy controls. The presence of P. gingivalis, usually occurring as a part of the salivary microbiome, has been reported in association with the progression and aggressiveness of esophageal SCC and generally with poor prognosis (69). Oral malhygiene and poor health have been proven as a risk factor for developing esophageal cancer. The abundance of the genera Lautropia, Bulleidia, Catonella, Corynebacterium, Moryella, Peptococcus, and Cardiobacterium has been observed significantly decreased compared to the healthy controls (70). Up to date, only limited evidence is available to prove that esophageal cancer is linked with gut microbiome (71). Sasaki et al. recently investigated gut microbiome as a novel biomarker of CRT efficiency in esophageal SCC. They reported that the abundance of Fusobacteriaceae family was enriched in the group of poor prognosis (stage II-IVb), also there was a significant association between increased relative abundance of Lactobacillaceae family and patients who achieved partial or complete response, thus suggesting that enriched relative abundance of Lactobacillaceae taxa can predict the effect of CRT (72). Moreover, F. nucleatum present in esophageal cancer tissue has been significantly associated with higher aggressiveness through activation of chemokines as CCL20, and consequently with shorter survival (73).
Gastric cancer. Gastric cancer remains one of the most common and most deadly cancers worldwide, currently being the fifth most common malignancy and the third most common cause of cancer-related death (74). Nowadays, according to the stage of gastric cancer, surgical resection, neoadjuvant CHT, or palliative chemotherapy, targeted therapy and/or immunotherapy is a standard of care. Primary endoscopic or surgical resection is reserved for small muscle non-invading tumors (cT1) or patients not suitable for CHT, the rest of the patients undergo neoadjuvant or palliative CHT, definitive CRT is indicated individually in patients internally not suitable for surgical treatment (75).
H. pylori was the first bacterial taxum causally associated with carcinogenesis (76, 77). Since this discovery by Marshall and Warren, many other representatives of gastric microbiota have been identified. Recent studies imply that the gastric environment itself affects the microbiome (78-80). Patients treated with PPI and absented from H. pylori infection tend to have a more acidic gastric environment which leads the phyla Firmicutes, Bacteroidetes, Actinobacteria, and Proteobacteria to be dominantly abundant, whereas in H. pylori-infected patients abundance of Streptococcus spp., Neisseria spp., and Staphylococcus spp. was enriched (81).
Ferreira et al. recently investigated the shift between the microbiome of patients with chronic gastritis and patients with gastric cancer, showing a significant decrease in microbial diversity in cancer patients. In terms of characteristics of the specific bacterial taxa, the abundance of Helicobacter spp. and Neisseria spp. and the growth of Citrobacter spp., Lactobacillus spp., and Clostridium spp. were both significantly lower in cancer patients (82). Coker et al. investigated the microbiome of patients with superficial gastritis, atrophic gastritis, intestinal metaplasia, and gastric cancer identifying that P. stomatis, S. anginosus, Parvimonas micra, Slackia exigua spp., and Dialister pneumosintes were enriched only in the gastric cancer patients’ microbiome. Based on receiver operating characteristic analysis (ROC) they were able to distinguish gastric cancer from gastritis based solely on their microbiome (83).
Although gastric cancer and gut microbiome have been quite profoundly investigated in terms of how the gut microbiome affects treatment response to chemotherapy and immunotherapy, up to date, no studies have investigated how either gastric or gut microbiota affect the treatment response to RT (84-92). The main possible reason for this is that RT is used primarily as a palliative hemostatic treatment and is not routinely performed with curative intent.
Pancreatic cancer. Pancreatic cancer has been consistently ranked among the last in terms of long-term survival of patients and as the seventh most common malignancy worldwide, with almost the same incidence as mortality (93). Nowadays, according to the stage of pancreatic cancer, surgical resection, neoadjuvant CHT and/or CRT (PREOPANC or other RT regimens), or palliative chemotherapy is a standard of care (94).
With regards to the poor prognosis of pancreatic cancer patients, extensive efforts were made to identify certain biomarkers, in terms of the microbiome (mainly oral and gut), Porphyromonas spp., Actinomycetes spp., Neisseria spp., Streptococcus spp., Bacteroides spp., Bifidobacterium spp., and Fusobacterium spp. have been linked to the occurrence and development of pancreatic cancer (95). Riquelme et al. investigated the pancreatic microbiome of patients with pancreatic ductal adenocarcinoma identifying the presence of the Pseudoxanthomonas-Streptomyces-Saccharopolyspora-Bacillus clausii as a predictor of good prognosis and long-term survivorship (96). Another potential bacterial risk factor for developing pancreatic cancer seems to include a higher representation of P. gingivalis in the oral microbiome, which has been supported by many studies (97-101). For the last 30 years, H. pylori has been the object of interest as a possible biological risk factor for pancreatic cancer, which was later supported by many meta-analyses (102-105). Historically, it was accepted that the pancreas has no specific microbiota as cultivation techniques were unyielding. However, new bacterial taxa were identified in pancreatic tissue with modern molecular techniques such as 16S rRNA sequencing, which are routinely used now. In addition to H. pylori and P. gingivalis, a high abundance of phyla Proteobacteria, Bacteroidetes, and Firmicutes was identified in pancreatic cancer tissue; however, these alterations have not been sufficiently explained yet (87, 106-108).
Although pancreatic cancer and gut microbiome have been quite profoundly investigated in terms of how the gut microbiome affects treatment response to chemotherapy and immunotherapy, the impact of RT on the microbiome and vice versa has been in the background of research interest (95, 109-111). Just recently, the pancreatic microbiome and its crosstalk with the pancreatic tumor microenvironment (TME) have started to be more and more investigated in hopes of finding a way to overcome chemoresistance and immunoresistance provided by unique pancreatic TME. Other researchers suggested that RT can disrupt TME, cause dysbiosis and impact anticancer treatment effectiveness; however, no significant findings have been so far reported (50, 52, 112-114).
Breast cancer. Breast cancer (BC) globally accounts for ca. quarter of all cancers and is the leading cause of cancer-related death among women (115). Although risk factors of BC have been extensively investigated, up to 70% of BC occur in women of average risk, and current prediction models fail to explain why (115-117). Historically, breast tissue was for long considered sterile; however, it has been recently hypothesized that microbiome could represent another risk factor, which had been previously omitted. Mounting evidence has supported this hypothesis as different microbial representations have been identified in the breast tissue of BC patients and the healthy controls. Moreover, specific bacterial assemblage was linked to cancer development and its aggressiveness (117).
Hieken et al. compared surgically obtained breast tissues of BC and benign tumor patients and observed a significantly higher abundance of Fusobacterium spp., Atopobium spp., Gluconacterobacter spp., Hydrogenophaga spp., and Lactobacillus spp. in the BC tissue (115). Although no studies have yet investigated the role of Fusobacterium spp. on the development and progression of BC, it has been hypothesized that fusobacteria could promote carcinogenesis in a similar matter as in HNSCC or CRC (118). Specific microbial signatures have been linked to the histological subtypes of BC (119).
Origin of BC microbiome includes three possible ways of access: a) passage from the skin via nipple-areolar orifice; b) nipple-oral contact during lactation or sexual intercourse; and c) bacterial translocation from the gut. Data supporting the first two theories come mostly from studies on the bacterial composition of breast milk and breastfed children (120). The indirect proof of possible gut microbiome translocation comes from the observation that orally administered probiotics are used to effectively treat lactational mastitis and can be detected in breast milk (121). This gut-breast microbial translocation led to the hypothesis that gut dysbiosis can lead to carcinogenesis in distant non-intestinal locations. Lakritz et al. demonstrated in a mouse model that gastric gavage of H. hepaticus leads to carcinogenesis in mammal glands (118).
Currently, almost no data supporting the influence of the microbiome on RT-response in breast cancer exists. Shiao et al. observed that the mouse BC model pretreated with antibiotics (ampicillin, imipenem, cilastatin, and vancomycin) had worse treatment response, i.e., shrinkage of tumor and/or time to progression or death, compared to the non-pretreated mice. Interestingly, they also observed an association between fungi and RT-response, which was enhanced after fungal depletion by fluconazole or 5-fluorocytosine (122).
Skin cancer. Compared to other cancers, skin cancer has been relatively understudied in terms of the microbiome and its role in carcinogenesis, progression of the disease and treatment response (123). RT-induced dermatitis is the most common adverse effect of skin cancer treatment affecting up to 90% of the patients, which can dramatically affect patients’ QoL (124-131). Ramadan et al. reported a significant reduction of bacterial diversity in the patients developing RT-induced dermatitis compared to healthy controls. They also observed delayed recovery in patients with an enriched abundance of Pseudomonas spp., Staphylococcus spp., and Stenotrophomonas spp. and an increased ratio of phyla Proteobacteria/Firmicutes, therefore, the shift in the structure and prevalence of the skin microbiome could support the role in the initiation and lasting of RT-induced dermatitis in cancer patients (131).
Colorectal cancer. Colorectal cancer (CRC) is second only to lung cancer in incidence and third in cancer-related deaths worldwide, and its incidence is gradually rising especially in developing countries that are adopting the “Western” lifestyle and diet (132). CRC is closely related to chronic inflammation and gut dysbiosis (133-136). The phylum Firmicutes and Bacteroidetes are the most common taxa in the gut microbiome, their relative abundance is significantly reduced in CRC patients’ gut microbiome (137-140). In contrast, the Fusobacterium spp. is significantly enriched in CRC patients and has been found as a risk factor for carcinogenesis as it produces butyric acid cytotoxic to intestinal mucosa leading to ulcerative colitis and chronic inflammation in whose terrain CRC can arise (141).
In terms of oncologic treatment, for locally advanced rectal cancer, surgery alone is not recommended, and neoadjuvant chemoradiotherapy (nCRT) or total neoadjuvant treatment (TNT) consisting of neoadjuvant CHT and nCRT should be implemented prior to the surgery (142). Recently, the gut microbiome of CRC patients has been investigated not only as a biomarker of RT-induced toxicity but also concerning the treatment response to CHT and/or RT (140, 143).
The high Fusobacterium spp. abundance in the gut microbiome is strongly associated with microsomal instability (MSI) of CRC, higher aggressiveness, and chemoresistance to 5FU, which may directly affect treatment response to nCRT, although no clinical data to support this hypothesis have been published (50, 82, 90, 99, 138, 139, 144-146). Mann et al. did not observe a significant association between levels of F. nucleatum abundance in the pre-treatment microbiome samples and chance of achieving CR after nCRT; however, they hypothesized that persistence of F. nucleatum after treatment could be associated with relapse and/or worse treatment results in rectal cancer patients undergoing nCRT (141).
Sun et al. investigated microbial taxa among other potential biomarkers of good response treatment to nCRT for locally advanced rectal cancer describing a complex model of associations between microbiome, the host immunomodulatory proteins, and immune cells. They observed a baseline abundance of Clostridium sensu stricto 1, and a fold increase of Intestinimonas spp. significantly correlated with the treatment response (143). Complex data regarding treatment response and RT-induced toxicities in rectal cancer were reported by Shi et al. Concordant to other studies, they observed significant changes in microbiome during and after nCRT. Increased abundance of Shuttleworthia spp. was observed in responders to nCRT. Several taxa from the genus Clostridium were in contrast enriched in the poor response group of patients. Moreover, the abundance of Bifidobacterium, Clostridium, and Bacteroides genera etc. were increased in patients having no or mild RT-induced diarrhea (146). Jang et al. reported Bacteroides spp. to be relatively more abundant in non-CR patients after CRT for rectal cancer compared to the CR group, where Duodenibacillus massiliensis was enriched (147).
The gut microbiome and pelvic radiotherapy. The gut microbiome is the largest and most complex microbiome of the human body and it is believed to influence the development, progression of disease and treatment response of gastrointestinal tumors, e.g., it has been proven that F. nucleatum, which usually inhabits the oral cavity and causes periodontal disease, contributes to the aggressive behavior of esophageal and colorectal cancer through activation of CCL20 cytokines in the tumor tissues (148, 149). Vice versa, it has been proven in animal models that the human gut microbiome has an impact on tumor development and progression in distant sites by a complex cascade of actions, including DNA damage, activating oncogenic signaling pathways, producing cancerogenic metabolites, and immunomodulating anti-tumor response (150-156). Although the gut microbiome is known to affect the treatment result of many anticancer therapies including chemotherapy, immunotherapy or even androgen deprivation therapy, relatively little is known about how gut microbiome impacts the treatment response of RT (157-163).
RT and/or CRT have bidirectional function, metanalysis by Touchefeu et al. described significant changes in gut microbiome after RT and/or CHT. They observed an increase of relative abundance in Bacteroides spp. and Entero-bacteriaceae family and a decrease in Bifidobacterium spp., Faecalibacterium prausnitzii, and Clostridium cluster XIVa (164). Based upon the observation of several recent studies investigating the impact of pelvic RT on gut microbiome, it seems that the relative abundance of phylum Fusobacteriota increases after radiotherapy, while phyla Firmicutes and Bacteroidetes significantly decreases; specifically, families Lachnospiraceae and Erysipelotrichaceae, genera Phasco-larctobacterium, Veillonella, Roseburia, Clostridium, and Ruminococcus were observed to increase in their relative abundance, while lactobacilli and genera Faecalibacterium, Peptococcus, Peptostreptococcus, Roseburia, and other anaerobes, to decrease (165).
Currently, no clinical studies provide more detailed insight into the possible role of the gut microbiome as a factor of radioresistance of radiosensitivity; however, in some preclinical animal trials, a similar pattern has been observed. Uribe-Herranz et al. reported interesting findings that oral vancomycin treatment of melanoma and lung/cervical model in C57/BI6 mice enhances treatment response to hypofractionated RT (either 21 Gy or 3´8 Gy) directly on the tumor, while the rest of the mice body was shielded. Treatment response was significantly higher in the orally vancomycin-pretreated mice both in the irradiated tumor as well as in the shielded tumor (166).
Radiotherapy-induced diarrhea (RID) can affect up to 80% of patients undergoing pelvic RT, depending on its severity, it can have a debilitating impact on the QoL of the patients (165). Manichanh et al. described that pre-RT gut microbiome is significantly different in patients who develop RID and in patients who do not, suggesting a possibility of decreasing the risk of developing RID via modulating the gut microbiome through identification of biomarkers and personalized medicine (167). Several studies concordantly demonstrated that a healthy gut microbiome of moderate or high diversity is a protective factor against developing RID, whereas dysbiosis and low microbial diversity before RT are strongly associated with developing RID (167). Numerous studies have shown radiotherapy of gynecological, prostate and lower gastrointestinal tract cancers majorly affects the gut microbiome (165, 167-171). These alterations include a reduction in the variety of bacterial taxa, which was associated with the manifestation of RID (138). The microbiome of the patients who developed RID showed an increased abundance of Bacteroides spp., Dialister spp., and Veillonella spp., and a reduced abundance of Clostridium XI and XVIII, Faecalibacterium spp., Oscillibacter spp., Parabacteroides spp., and Prevotella spp. (167, 172-174).
The impact of pelvic RT on the gut microbiome is outlined in Table II.
Focused literature review – Impact of pelvic RT on gut microbiome.
The effect of probiotics on reduction of incidence or severity of RID has been demonstrated in animal models, however, the evidence from clinical trials is still lacking (175-178). Urbancsek et al. described a significant benefit of Lactobacillus-based probiotics in management of RID in double-blinded randomized prospective trial undergoing abdominal RT (179). Salminel et al. observed a significant reduction of severe RID in the experimental group of women treated by pelvic RT for gynecological malignancies, the women supplied with fermented milk products containing L. acidophilus developed severe RID only in 30% of cases compared to the 90% of women in the control group (180). Delia et al. later observed a significant reduction of diarrhea in the experimental group using a probiotic VSL#3 compared to placebo (incidence of all-grades RID 38% vs. 58%, and Grade 3-4 6% vs. 29%) (181). Similar results were also reported with other probiotics, such as B. breve and L. acidophilus, significantly reducing the incidence of Grade 2-3 RID (9% vs. 45% in the group with placebo) (182).
Table III comprehensively outlines the impact of probiotics on RID.
Focused literature review – Efficacy of probiotics on radiotherapy-induced diarrhea.
Conclusion
We are still far from fully comprehending the complex interconnectedness of site-specific microbiomes. Mounting evidence support the theory of microbial communication and translocation between organ specific microbiomes, e.g. translocation of ingested microbes from digestive tract into mammal glands and breast milk etc. To characterize human microbiome more accurately, we should not consider each anatomical site as an isolated entity, but we should rather focus on description of microbial representation shifts in other microbiomes such as the oral and/or the gut microbiome as well.
In terms of cancer, some distant microbiome alterations, existing prior to as well as being induced by the treatment, have been described as significant harbingers of the poor prognosis, and/or treatment-resistance of certain cancers. Higher abundance of Fusobacterium spp. in oral microbiome significantly correlates with incidence of several distant cancers, and its intratumoral presence correlates with early progression and higher aggressiveness in esophageal, pancreatic, and colorectal cancer. Similarly, the higher abundance of P. gingivalis in saliva correlates with worse prognosis of esophageal cancer patients.
From the RT perspective, it appears that the microbiome is an important player in local control as well as systemic therapy response. So far, the most studied is the gut microbiome. Based on clinical studies, the gut microbiome can directly affect the response to CRT, incidence, and severity of acute toxicity. This has been observed not only in the case of nCRT of locally advanced rectal cancer. Distinctive microbial profiles were observed in advanced rectal cancer patients gut microbiome who reached CR or not after nCRT, moreover, it was observed that certain microbial profiles also significantly correlated with the incidence and severity of RID. Similar results were also observed in patients undergoing pelvic RT for prostate or gynecological cancers. First attempts to decrease the incidence and severity of RID by the administration of oral probiotics were reporting significant several fold decrease. Interestingly, lung cancer patients undergoing definitive CRT having higher abundance of phylum Firmicutes/class Clostridia in their gut microbiome were associated with a favorable prognosis and longer PFS; however, it remains unclear whether it is caused by bacterial metabolites affecting CHT efficacy and/or transposition of radiosensitizing bacteria into lung cancer tissue.
Considering the oral microbiome and RT, the scope of scientific knowledge is eminently narrower. The oral microbiome is altered throughout the RT, and its constitution is associated with the severity of RIOM; however, no convincing evidence that the oral microbiome has a significant impact on RT efficiency has been published. Certain probiotics have already been designed to relieve the severity of RIOM and thus enhance the QoL of HNC patients, which hypothetically opens the door for further investigation if the RT efficiency can be enhanced by concurrent use of selected probiotics. The feasibility and efficacy of microbiome-targeted interventions to enhance HNC RT outcomes require rigorous evaluation.
What appears to be an intriguing approach in future cancer research is combined microbial research focusing on the changes in the oral and gut microbiome, and potentially even in the intratumor microbiome. These alterations have not been properly investigated, although the interconnections between gut microbiota and distant niches were described. Focusing on the changes in the oral and gut microbiome could answer questions of (chemo)-radioresistance of certain tumors, including the HNSCC. Another very interesting aspect is the possibility to modify the incidence of severity of radiation-induced adverse effects, such as RIOM and/or RID. Based on the recent reports, probiotics can effectively decrease the incidence and severity of RID and/or RIOM; however, this requires further investigation. However, already published results are promising.
In conclusion, the human microbiome plays a pivotal role in modulating RT and CRT outcomes, influencing both treatment response and toxicity profiles. For potential future research in cancer radiotherapy field, we propose these directions:
1. Investigation of microbial alteration in distant, i.e., out-of-field, microbiomes in cancer patients treated with radiotherapy;
2. Characterization of intratumor microbiome as possible marker of radioresistance;
3. Clinical trials investigating the effect of pro-/pre-/ symbiotics in prevention and/or alleviation of acute radiotherapy-induced toxicities.
- Received July 29, 2024.
- Revision received October 2, 2024.
- Accepted October 14, 2024.
- Copyright © 2025 The Author(s). Published by the International Institute of Anticancer Research.
This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY-NC-ND) 4.0 international license (https://creativecommons.org/licenses/by-nc-nd/4.0).
