Profiling the microbiome in breast cancer patients before and after treatment with cannabis oil

With long-term cancer treatment, female breast cancer patients encounter debilitating side effects from a current range of cancer therapies. Cannabinoids (CBs) have been widely used due to their efficacy on chemotherapy-associated pain. Although there is some evidence that cannabis oil may enhance quality of life and lessen symptoms in people with breast cancer, this is not very substantial. No randomised trials evaluating the effects of cannabis in breast cancer patients has been conducted before. The microbiome is critical to human health and disease, and accumulating data suggests that it is influenced by a variety of external variables, including marijuana, cannabis, cannabinoids and endocannabinoid mimics. We, therefore, conducted a series of comprehensive systematic reviews of the literature, 1) to analyse the microbiome in breast cancer, 2) to assess effects of cannabis and cannabinoids on the human microbiota, 3) to evaluate the impact of other interventions in breast cancer such as probiotics, 4) to establish a microbiome pipeline using three DNA lysis methods, optimised using bacterial and fungal community standards, 5) to evaluate optimised extraction pipeline for three newly available NIBSC, WHO recommended, microbial (bacteria) reference standards, and amplicon sequencing performed across Illumina and Nanopore platforms, and 6) to develop and support a comprehensive randomised placebo-controlled clinical trial (RCT) assessing cannabis oil treatment in breast cancer patients receiving chemotherapy, that afforded opportunity to examine for changes in the gut and oral microbiome, cytokines, bacteria-derived faecal metabolites, and markers of bacterial translocation to the circulation, before and after intervention. In the RCT, new or current breast cancer patients receiving chemotherapy, aged >18 years old, were recruited and randomised to 3 different intervention arms of the study: Ganja oil, Metta Osot and placebo coconut oil, at which blood, faeces and saliva samples were collected at the baseline, and after 12 weeks intervention. The pilot data, which involved 10 patients, was used for this thesis. Due to a limited sample size, the analysis was conducted between 2 main groups: cannabis oil (Ganja oil and Metta Osot) versus placebo. From the systematic review of the microbiome in breast cancer, it revealed lower bacterial diversity and distinctive patterns of gut microbiota, particularly higher Firmicutes/Bacteroidetes ratio in the patients. It was observed both positive and negative correlations with microbiota (oral, gastrointestinal, faecal and vaginal) influenced by marijuana/cannabis use, and interventions with cannabinoid and endocannabinoid-like mediators, extending different clinical contexts, including cognitive deficit/depression, HIV infection, inflammation/pain, obesity and oral disease. Analysis of a significant number of intervention studies that probiotic use in breast cancer patients and survivors, including combination prebiotic use, showed some benefit for preventive and palliative care. From the optimisation study, we recommended DNA extraction using lysis buffer (ZymoBIOMICSTM) alone as an optimal approach and applied it at the RCT project. Using four microbial standards, two lysis methods, and two sequencing chemistries, the observed microbial profile was compared with the expected abundance, revealing the most similarity with the utilisation of DNA_Mix at our microbiome pipeline. Assessing microbiome profiling using Illumina and Nanopore in two centres in Thailand, microbial pattern variation across centres were reported, probably biases from the library preparation and sequencing. This thesis addresses the need for clinical study to comprehensively examine the action of cannabis oil on the gut microbiome, and metabolome in breast cancer patients as part of a bigger trial that anticipates the beneficial actions of cannabis intervention, cost-effective traditional medicine to help alleviate the debilitating side-effects of chemotherapy in breast cancer patients.