https://openworks.mdanderson.org/sumexp22/1054/thumbnail.jp

Gonzalez, Caleb

Pilla, Pranathi

Piwnica-Worms, David

English

OpenWorks @ MD Anderson

Tumor Microbiome in Murine 4T1 Triple Negative Breast CancerPranathi Pilla, Caleb Gonzalez, David Piwnica-WormsUniversity of Texas MD Anderson Cancer Center Department of Cancer Systems ImagingAbstractQuestion 3Preliminary ConclusionsApplication1. Tumor Microbiome Screen (A)2. Bacteria Culture3. Bacteria Construction4. Ideal use-caseThe presence of certain microorganisms has been known to have a strong association with the prognosis and development of human cancers. Recent findings show that the gut microbiome can have a significant role in activating antitumor innate immune responses. In addition, the microbiome in the tumor and its compartment has been found to impact progression and immunotherapy outcomes in patients.Herein, we utilized 16S deep sequencing platform to profiled 4T1 tumor microbiome of mice treated with immune checkpoint therapy (anti-PD-1 + anti-CTLA4) and found that: 1) 4T1 tumors are mostly colonized by aerobic bacteria, 2) tumor progression may be affected by gut microbiota composition, and 3) immune checkpoint therapy may affects microbiome profile and the biodiversity. Furthermore, preliminary qPCR evidence indicates that 4T1 tumors may feature a low microbial load, compared with pancreatic, liver, and lung tumors. Future applications of probiotics as an anti-tumor drug vectors remain open for future exploration.Key Questions1. Are 4T1 tumors colonized by bacteria?2. Does the microbiome affect tumor progression?3. Does ICT treatmentaffect the tumor microbiome?This is important because it can…• Help understand factors that determine failure or success of treatment.• Open a new drug delivery route using oral intake of probiotics. MethodsResults/ObservationsNot treated ArmVehicle n =2ICT (α-PD-1 + α-CTLA-4) n = 3Baytril Treated ArmVehicle n =2ICT (α-PD-1 + α-CTLA-4) n = 3Does Immune Checkpoint Therapy (ICT) affect the tumor microbiome?Results/Observations1. ICT Treatment may result in an increase in biodiversity of speciesBaytril Treated Mice Samples4T1 FUGW Tumor Microbiome Composition in Untreated MiceFuture Work1. Identify bacterial candidates• Previously known to colonized human tumors• Biosafety Level 1• Commercially available• Culturable• Genome sequenced2. Culture bacteria – for lab repository and experimental purposes3.    Experimental Opportunities• In vivo real time imaging of gut bacteria colonization of tumors – engineer Lux operon into bacteria• Bacteria as drug-delivery vectorInitial Questions This Project’s Findings•Are 4T1 tumors colonized by gut bacteria?•How does tumor microbiome affect tumor progression?•Does ICT treatment affect the tumor microbiome?•Yes•Increased tumor volume in vehicle control mice treated orally with antibiotic points at an antitumor role for the tumor microbiota•May increase diversity of tumor microbiota•Unknown if it increases/decreases number of bacteria•Changes in tumor microbiome affecting treatment is to be determinedSimpson Reciprocal Values in 4T1 FUGW BaytrilAntibiotic Mice TumorsSimpson Reciprocal Values in 4T1 FUGW Untreated Mice TumorsUntreated Mice SamplesLikely engaging in an anti-tumor response Orthotopic tumorimplantationWeek 0 Week 2Start of treatmentsTumor extraction fortumor-microbiomeprofilingWeek 3Mouse (Balb/c by J)Tumor4T1 FUGW-FLTreatmentTumor growthICT i.p. VehicleDay ICT (i) onlyTumor implantationICT onlyPBSTumor extraction0 15 17 19 21 22 23 Microbiome Facility Core16S analysis provides: taxonomy table, level abundance table, microbial composition, alpha, and beta diversity files. 1 2 34681012WeekTotal Photon Flux (log [photons/sec])1 2 34681012WeekTotal Photon Flux (log [photons/sec])2 30.00.20.40.6WeekTumor Volume (cm3)Vehicle Mouse 1Vehicle Mouse 22 30.00.20.40.6WeekTumor Volume (cm3)Vehicle Mouse 1Vehicle Mouse 2BaytriltreatedNot treatedVehicletreated1 2 34681012WeekTotal Photon Flux (log [photons/sec])1 2 34681012WeekTotal Photon Flux (log [photons/sec])2 30.000.050.100.15WeekTumor Volume (cm3)ICT Mouse 2ICT Mouse 1ICT Mouse 32 30.000.050.100.15WeekTumor Volume (cm3)ICT Mouse 2ICT Mouse 1ICT Mouse 3ICT treatedBaytriltreatedNot treated4T1 FUGW Tumor Progression MeasurementsRelative AbundanceQuestion 14T1 FUGW Tumor Microbiome Composition in Baytril MiceRelative AbundanceAre 4T1 Tumors colonized by bacteria?Question 20.00.20.40.6Tumor Volume (cm3)46810Total Photon Flux (log [photons/sec])Not TreatedBaytril Treated0.000.050.100.150.6Tumor Volume (cm3)46810Total Photon Flux (log [photons/sec])Not TreatedBaytril TreatedVehicle Control ICT TreatedDoes the 4T1 tumor microbiome affect tumor progression?Results/Observations1. Bacteria colonizes 4T12. Tumor microbiome affected by oral antibiotic treatmentResults/Observations1. Untreated mice have lower tumor volume and BLI than Baytril group, indicating relationship between microbiome and tumor progression qPCR Standard CurveBacterial load is below detectable levels for qPCR, indicating that 4T1 tumors have lower bacterial load than pancreatic, liver, and lung tumors.The Simpson’s reciprocal index quantifies biodiversity by taking into accountspecies richness and evenness. High species diversity suggests a greater number of successful species and a more stable ecosystem Candidate MicroorganismsPelomonas saccharophila• ~ 60% relative abundance in vehicle control mice 1, genome sequenced• Aerobic, gram negative, Biosafety level 1, commercially availableBrevundimonas nasdae• ~ 80% relative abundance in vehicle control mice 2, genome sequenced• Aerobic, gram negative, Biosafety level 1, commercially availableLachnospiraceae NK4A136• ~ 20% relative abundance in ICT mouse 1, genome sequenced• Immunogenic, indirectly associated with immune therapy response1.• Anaerobic, gram negative, Biosafety level 1, commercially availablel r iliaBrevundimonas nasdaeLachnospiraceae NK4A136Does gut microbiome affect tumor microbiome?AcknowledgementsSpecial thanks to the CPRIT-CURE program and the CPRIT grant for funding this experience and this opportunity. 

Tumor Microbiome in Murine 4T1 Triple Negative Breast Cancer

https://openworks.mdanderson.org/cgi/viewcontent.cgi?article=1054&context=sumexp22

Tumor Microbiome in Murine 4T1 Triple Negative Breast Cancer

Abstract

Similar works

Full text

Available Versions

OpenWorks @ MD Anderson