Abstract
Background: The DNA methylation dioxygenase TET2 is frequently mutated in hematological malignancies[1,2]; however, TET2 mutations are also frequently detected in individuals with clonal hematopoiesis[3], suggesting additional factors might be required to promote the malignant transformation of TET2-defective hematopoietic cells. Indeed, microbia-triggered inflammation has been recently shown to promote the myeloproliferation of Tet2 deficient hematopoietic cells[4]. This finding points to a strong association between intestinal microbiota and epigenetic alterations in the hematopoietic system during disease progression. Gut microbiota can be affected by antibiotics treatment and antibiotics could alter murine hematopoiesis through the depletion of intestinal microbiota[5]. Antibiotic treatment is commonly used in hematological malignancy patients with infection[6], which is one of the major complications in these patients. Nonetheless, it is not clear how antibiotics might impinge on malignant blood cells with epigenetic defects (e.g., Tet2 deficiency). In this study, we aim to characterize the impact of antibiotics treatment on Tet2 deficient malignant cells in the hematological system.
Methods: We established two sets of Tet2 knockout (KO) malignant cell lines derived from aged Tet2KO mice with T cell lymphoma (n = 6) or MDS/CMML (n =3). Flow cytometry analysis and molecular analysis confirmed the tumor phenotypes. Furthermore, these cells were transplantable in immune competent CD45.1 mice and underwent expansion without losing their tumor properties. To evaluate the impact of antibiotics treatment on tumor growth, we transplanted Tet2KO T cell lymphoma cells or CMML/MDS cells into CD45.1 recipient mice followed by antibiotic treatment (ampicillin, vancomycin, neomycin, and metronidazole) for 10 days. Then we monitored tumor cell growth in peripheral blood, spleens and bone marrows collected from CD45.1 mice with and without antibiotics cocktails treatment. We also collected the Tet2KO tumor cells 20 days after antibiotics treatment and will perform transcriptomic and 5hmC profiling to further investigate how antibiotics alters the molecular signatures in Tet2KO tumor cells.
Results: Consistent with earlier reports, we observed a moderate bone marrow suppression during the antibiotics treatment in recipient mice but not after cessation of treatment[5]. Interestingly, we also observed that antibiotics treatment significantly suppressed Tet2KO tumor growth in the recipient mice. Mice treated with antibiotics cocktails showed a pronounced decrease of microbiota in collected feces. Furthermore, the antibiotics treated group displayed significantly longer survival compared with the untreated group. Flow cytometry analysis showed a significant decrease of CD45.2+ Tet2KO tumor cells in peripheral blood, spleens and bone marrow 20 days after antibiotics treatment. Detailed analysis in bone marrow collected from recipient mice transferred with CMML/MDS-like Tet2KO revealed up to 80% reduction of c-Kit+ cancer stem-like cells in the antibiotics treated group without significantly affecting endogenous hematopoietic cells in recipient mice 20 days after cessation of antibiotic treatment.
Conclusion: In summary, we conclude that antibiotics treatment could partially suppress/delay Tet2KO tumor cells growth. Further RNA-seq and 5hmC mapping will be performed to gain detailed mechanistic insights. Furthermore, we will determine whether the suppressive effect is due to direct action on tumor cells or because of indirect effects from the removal of intestinal microbiota. Nonetheless, our data demonstrated the potential benefits of antibiotics treatment on hematological malignancies associated with Tet2 loss of function.
References
1. Couronne, L., Bastard, C. & Bernard, O. A. (2012). N Engl J Med 366, 95-96.
2. Delhommeau, F., Dupont, S., Della Valle, V. et al. (2009). N Engl J Med 360, 2289-2301.
3. Busque, L., Patel, J. P., Figueroa, M. E. et al. (2012). Nat Genet 44, 1179-1181.
4. Meisel, M., Hinterleitner, R., Pacis, A. et al. (2018). Nature 557, 580-584.
5. Josefsdottir, K. S., Baldridge, M. T., Kadmon, C. S. et al. (2017). Blood 129, 729-739.
6. Safdar, A. & Armstrong, D. (2011). Clin Infect Dis 53, 798-806.
Disclosures
No relevant conflicts of interest to declare.
Antibiotic Intervention Affects Maternal Immunity During Gestation in Mice
