P319: The Testicular Niche of Acute Lymphoblastic Leukemia – Molecular and Cellular Factors for the Preferential Migration and Survival

Abstract

Background:Background: The testis is the second most frequent (30%) non-hematological extramedullary site of relapse in pediatric acute lymphoblastic leukemia (ALL) treated according to European protocols. Testicular relapses usually occur late (> 6 months after completion of frontline treatment), and boys with a ETV6::RUNX1 fusion gene positive ALL have a significantly higher risk of a testicular relapse. In adult ALL, only about 1% of patients have a testicular relapse. Event-free survival is between 40%-80% depending on timing of relapse, involvement of the contralateral testis and of the bone marrow. The surgical removal of the clinically-involved testis or high-dose irradiation are the only current treatment options for testicular relapses to achieve a long-term event-free survival, but it impacts longterm quality of life. The molecular mechanisms regulating leukemic cell migration, growth, and survival in the testis have not been sufficiently assessed. Aims:Aims: We aimed at investigating our hypothesis that the testis is a frequent source of extramedullary relapse in pediatric ALL because of the physiologically strong CXCL12 gradient. This allows circulating leukemia cells to migrate into the testis during pre-puberty and to survive for potentially longer periods because of the specific immune suppressive microenvironment at this unusual anatomical site. Methods:Methods: To dissect the pre-pubertal cellular requirements and molecular pathways contributing to testicular leukemic cell dissemination and survival, we combined analysis of primary human leukemias with a patient derived xenograft (PDX) mouse model. We analyzed chemokine receptor expression profiles of pediatric B-ALL samples from patients with different relapse sites, studied the crosstalk of leukemia cell-stroma in co-cultures, and established a pediatric B-ALL PDX mouse model with testicular involvement. Results:Results: The CXCL12-CXCR4 was identified as the driving force for B-ALL cell migration and survival in the testicular leukemic niche. Analysis of primary pediatric patient samples revealed that CXCR4 was the only chemokine receptor being robustly expressed on B-ALL cells both at the time of diagnosis and relapse. One prerequisite for leukemic cell infiltration in the testis mice was high surface expression of CXCR4 on PDX-ALL cells, and CXCL12 secretion from testicular stroma. In affected patient testes, leukemic cells localized within the interstitial space in close proximity to testicular macrophages. Another requirement for migration and survival of leukemia cells in the testis was pre-pubertal age of the recipient mice. Leukemic cell interactions with specific testis macrophage subpopulations, isolated from affected testes, altered their phenotype towards a pro-tumorigenic M2-like phenotype. The blockage of CXCR4-mediated functions by anti-CXCR4 antibody treatment reduced testicular infiltration of PDX-ALL cells.Summary/Conclusion: Summary/Conclusion: Collectively, a pre-pubertal condition together with high CXCR4 expression are fac...