Search

Relapsed/refractory T cell acute lymphoblastic leukemia (T-ALL) is difficult to salvage especially in heavily pretreated patients, thus novel targeted agents are sorely needed. Hyperactivated JAK/STAT and BCL2 overexpression promote increased T-ALL proliferation and survival, and targeting these pathways with ruxolitinib and venetoclax may provide an alternative approach to achieve clinical remissions. Ruxolitinib and venetoclax show a dose-dependent effect individually, but combination treatment synergistically reduces survival and proliferation of Jurkat and Loucy cells in vitro. Using a xenograft CXCR4+ Jurkat model, the combination treatment fails to improve survival, with death from hind limb paralysis. Despite on-target inhibition by the drugs, histopathology demonstrates increased leukemic infiltration into the central nervous system (CNS), which expresses CXCL12, as compared to liver or bone marrow. Liquid chromatography-tandem mass spectroscopy shows that neither ruxolitinib nor venetoclax can effectively cross the blood-brain barrier, limiting efficacy against CNS T-ALL. Deletion of CXCR4 on Jurkat cells by CRISPR/Cas9 results in prolonged survival and a reduction in overall and neurologic clinical scores. While combination therapy with ruxolitinib and venetoclax shows promise for treating T-ALL, additional inhibition of the CXCR4-CXCL12 axis will be needed to eliminate both systemic and CNS T-ALL burden and maximize the possibility of complete remission.

Background Relapsed pediatric sarcomas have a poor prognosis with no available curative options. Alpha-Tocopheryloxyacetic acid (a-TEA) is a redox-silent analog of alpha-tocopherol that induces apoptotic and immunogenic cell death of tumor cells at doses that are not harmful to healthy normal cells. In a first-in-human clinical trial, a-TEA was well tolerated in adults with advanced solid tumors (NCT02192346), but has not yet been studied in pediatric sarcoma. We used a murine model of rhabdomyosarcoma (M3-9-M RMS) to assess the in vitro and in vivo anti-tumor effects of a-TEA. Methods In vitro studies were performed on the M3-9-M RMS cell line to measure a-TEA-mediated apoptosis using flow cytometry (Annexin V+/7AAD+ cells) and live cell imaging (Annexin V+ cells). In vivo studies involved orthotopic implantation of luciferase+ M3-9-M tumor cells into syngeneic C57BL/6 recipients. Once tumors were palpable, mice were randomized to a control diet or a-TEA-supplemented chow for 21 days and evaluated for bioluminescence, tumor growth and overall survival. Gene expression of tumor-infiltrating and splenic T cells were analyzed by bulk RNA-Seq and flow cytometry respectively. Results M3-9-M RMS treatment with 2.5–100 uM a-TEA induced apoptosis in a dose-dependent manner within 24 hours (p 4 weeks) compared to recipients of matched control chow (p Conclusions These data indicate that a-TEA mediates apoptosis of RMS in vitro and suppresses in vivo tumor growth, leading to prolonged survival likely via enhanced activation of adaptive immunity through CD4+ T cells and suppression of innate immunity through regulation of myeloid cell subsets. Furthermore, a-TEA may have direct effects on tumor cell proliferation through EP300 and c-Jun as well as indirect effects on tumor growth by regulation of immune cell recruitment through CD24 and CXCR4 gene expression. Administration of a-TEA as a potential salvage treatment for RMS is warranted. Acknowledgements The study was supported by NIH TL1 TR002375 (FS), St. Baldrick’s Stand up to Cancer (SU2C) Pediatric Dream Team Translational Research Grant SU2C-AACR-DT-27-17, NIH/NCI R01 CA215461, American Cancer Society Research Scholar Grant RSG- 18-104-01-LIB, and the Midwest Athletes Against Childhood Cancer (MACC) Fund (CMC). SU2C is a division of the Entertainment Industry Foundation. Research grants are administered by the American Association for Cancer Research, the scientific partner of SU2C. The contents of this article do not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the US government. Ethics Approval The University of Wisconsin-Madison Animal Care and Use Committee approved all protocols (M005915).

Treatment of relapsed/refractory B cell acute lymphoblastic leukemia (B-ALL) remains a challenge particularly in patients who do not respond to chemotherapy or immunotherapy. While a subset of these patients can be cured with allogeneic bone marrow transplant, success of the transplant is contingent upon achieving a minimal residual disease negative complete remission. The objective of this study was to assess the efficacy of fedratinib, a semi selective JAK2 inhibitor and venetoclax, a selective BCL-2 inhibitor, on the B-ALL cell lines RS4;11, NALM-6, and SUP-B15. While select studies have examined the efficacy of these drugs on B-ALL as single agents, data is lacking on whether these drugs work optimally as single agents or in combination. Cell lines were treated with either vehicle (DMSO), single agent fedratinib or venetoclax, or the combination of both inhibitors for 48 and 72 hours. Fedratinib doses were tested from 150 nM to 1 uM and venetoclax doses were tested from 2.5 nM to 500 nM. Live/dead discrimination was performed using flow cytometry and a 72-hour MTS assay to assess proliferation. Drug synergy was calculated using CompuSyn software to generate combination index (CI) scores, with a CI score less than 1 reflecting synergy. RNA was isolated from treated leukemia cells and processed on a Nanostring nCounter platform to assess potential changes in gene expression. Results were then analyzed using Rosalind data processing software. In RS4;11, the combination of 5 nM venetoclax and 525 nM fedratinib significantly increased cell death compared to single agent venetoclax (p=0.0451) and fedratinib (p=0.0077). Furthermore, the combination significantly decreased leukemia cell proliferation compared to single agent fedratinib (p Disclosures Capitini: Novartis: Honoraria; Nektar Therapeutics: Honoraria.