Discussion
Overall, we found that long-term survivors of childhood cancer have decreased gut bacterial microbiome diversity and differently abundant bacterial taxa, even years after the completion of therapy. Further, these changes are apparent despite the finding of similar present diet practices between survivors and controls. Additionally, radiation therapy, particularly to the abdomen and pelvis, but also cranial radiotherapy, appears to be associated with reductions in gut microbial diversity and differently abundant taxa. Interestingly, exposures such as chemotherapy and antibiotic intensity were not directly associated with alpha diversity in survivors, though they were associated with differentially abundant taxa. In this study, alpha diversity itself did not appear to be associated with components of the metabolic syndrome or chronic inflammation among childhood cancer survivors, but low prevalence of metabolic derangements in the study population may have precluded this finding. Notably, however, among survivors with increased adiposity, elevated body mass index, and increased inflammation, differently abundant taxa were present, suggesting a potential interaction with the microbiome.
Two previous studies in similar populations also demonstrated reduced microbial diversity in childhood cancer survivors. Among 13 long-term Hodgkin lymphoma survivors and twin controls (8 dizygotic, 5 monozygotic), survivors had fewer operational taxonomic units in fecal samples compared to their co-twin control [39]. A recent study of adult survivors of childhood acute lymphoblastic leukemia from Malaysia compared survivors to healthy controls [18]. Similar to the present study, Chua, et al. recruited controls among healthcare works, siblings, and other volunteers and were matched for sex and ethnicity, though neither subjects nor controls were excluded for recent antibiotic use. Chua, et al. also found that survivors had altered composition of taxa and decreased alpha diversity of bacteria compared to healthy controls, but used anal swabs for sample acquisition.
Survivors of childhood acute lymphoblastic leukemia have been previously demonstrated to have increased levels of inflammation compared to controls [17]. Indeed, Chua, et al. demonstrated modest associations between bacterial taxa and Interleukin-6 and C-Reactive protein, though the role of cancer history, chemotherapy and antibiotics in these associations was unclear [18]. In the present study we were not able to detect an association between alpha diversity and markers of chronic inflammation in survivors, but did see differences in taxa abundance based on levels of C-Reactive protein. As opposed to the work previously performed, this is the first study to clearly demonstrate alterations in microbiota in a heterogeneous cohort of long-term survivors based on multiple different underlying malignancies. Additionally, although radiation has been shown to lead to short term changes in the microbiome in oncology patients [40,41], ours is the first study to demonstrate a relationship between radiation and microbiome diversity many years after treatment.
Our study has some limitations. First, our cohort did not have high levels of phenotypic abnormalities. Hyperlipidemia and insulin resistance were uncommon, and most patients did not possess components of the metabolic syndrome. This limited our ability to detect an association of the microbiome with these late effects, and perhaps a larger or older cohort of patients may have improved that power. Further, the associations between differently abundant taxa and phenotypes demonstrated in this study do not necessarily imply causation, and mechanistic studies would be required to further characterize these relationships. Additionally, we did not perform internal transcribed spacer sequencing in this study to detect a potential interplay between the bacterial microbiome and the gut fungal microbiome, which has been implicated in obesity and other diseases [42,43]. Finally, this was a single institution exploratory study, and microbiome findings may be unique to the patients seen in our clinic or geography. Regardless, this study still defines important relationships ripe for further exploration.
Overall, this study demonstrates that long-term survivors of childhood cancer have decreased gut bacterial microbiome diversity and differently abundant bacterial taxa, even years after the completion of therapy. Additionally, radiation may be particularly important in microbial diversity among this cohort. In future experiments, we aim to further categorize the metabalome in a similar cohort of patients to better understand if the difference found on 16S sequencing results in alterations of metabolic products in the blood-stream. We also aim to explore potential relationships with the fungal microbiome and gain a better understanding of mechanisms via gnotobiotic mouse experiments. Further, in order to detect associations with clinically relevant late effects, we aim to perform a larger multicenter study using older subjects with greater prevalence of comorbidities.