Abstract Acute lymphoblastic leukemia (ALL) is the most common pediatric cancer and the second most common acute leukemia in adults. Despite best existing therapy, older children and elderly ALL patients have poor prognoses. Thus, there is a need to identify alternative therapeutics for these patients. There has been controversy about the function of the gut microbiota in cancer, with some reports showing that it supports tumor growth and others showing tumor growth inhibition. Recently, Dr. Melvyn Greaves proposed that ALL development may have a microbial trigger; however, gut microbiota studies in ALL are scarce. Little is known about the role of the gut microbiota in ALL progression and in reshaping cancer cell-intrinsic pathways that are as important as immune responses in the outcome of ALL. Recently, we found that gut microbiota depletion in immunocompromised mice reduces ALL growth by inducing apoptosis and inhibiting proliferation. We show that in immunocompromised microbiome-depleted mice, the rate of ALL tumor growth is reduced compared to microbiome-competent mice. Molecularly, we found that microbiome ablation results in changes in cancer cell-intrinsic pathways of apoptosis and proliferation in these tumors. To further understand how the gut microbiota affects these cancer cell-intrinsic pathways, we have performed RNA sequencing on tumors from microbiome-competent and microbiome-depleted mice accompanied by 16s rRNA sequencing from stool samples and mass spectrometry of blood serum to identify metabolites secreted by the gut microbiota that reach the tumor site, affecting cancer cell growth. Our goal is to define bacterial species and metabolites involved in ALL progression and to mechanistically determine how these bacterial species and metabolites regulate ALL growth via regulation of cancer cell-intrinsic pathways. These data suggest that the gut microbiota may regulate ALL progression in an immune-independent manner via transcriptional changes of cancer cell-autonomous pathways. We envision that the gut microbiota can be reshaped in ALL patients who are elderly, chemo-refractive, and/or immunocompromised to alter cancer cell-intrinsic pathways to treat ALL. The clinical impact of these findings is significant given that ALL patients who are immunocompromised could still benefit from therapeutics aiming at modulating the gut microbiota. Citation Format: Wadie Mahauad-Fernandez, Soumaya Zlitni, Ami Bhatt, Dean Felsher. The gastrointestinal microbiota controls cancer cell intrinsic mechanisms to promote the progression of acute lymphoblastic leukemia [abstract]. In: Proceedings of the AACR Special Conference on the Microbiome, Viruses, and Cancer; 2020 Feb 21-24; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2020;80(8 Suppl):Abstract nr B21.