Background: Cervical cancer is a global health problem. Despite scale up of prevention programs, there will be an ongoing need to improve the effectiveness of radiotherapy/cisplatin (RTCT) for women with established, locally advanced disease. We have shown that RTCT with the CXCR4 inhibitor plerixafor improves tumor control, reduces metastases and reduces RT-related side effects compared to RTCT alone. X4-136 is an oral CXCR4 inhibitor that is better suited to long-term, once daily use. In this study we examined: 1. The efficacy of RTCT and X4-136 in cervical cancer; 2. Biomarkers of response to RTCT and X4-136; and 3. Effects of RTCT and X4-136 on intestinal toxicity, an important dose-limiting RT side effect in these patients. Methods: Orthotopic cervical cancer xenografts derived from our patients were treated with RT (30 Gy; 2 Gy/day) and cisplatin (4 mg/kg/week ip) with or without concurrent X4-136 (100 mg/kg/day orally) for 3 weeks. Mice were sacrificed immediately after treatment for biomarker assessment. In separate experiments, mice had CT imaging weekly after treatment to evaluate tumor response. Acute and late intestinal toxicity was assessed histologically 3.5 and 90 days after treatment respectively. Results: RTCT alone increased CXCL12/CXCR4 signaling, intratumoral accumulation of myeloid cells and PD-L1 expression. The addition of X4-136 during RTCT abrogated these effects and improved tumor response in 2 cervical cancer models. Furthermore, the addition of X4-136 increased the proportion of surviving intestinal crypt cells in keeping with a reduction in acute RT toxicity, and reduced late histologic changes of late RT toxicity. Conclusions: The addition of X4-136 blunts RTCT-induced upregulation of the CXCL12/CXCR4 pathway, reduces intratumoral myeloid cells, improves tumor control and reduces side effects in cervical cancer. Few if any pharmacologic strategies have been identified previously that expand the therapeutic window with RT in this way. The combination of RTCT and CXCR4 inhibition warrants testing in clinical trials to validate these findings. These benefits may apply to other tumors where RTCT plays a curative role.