Precision Targeting of Myeloid Src-family Kinases in Acute Myelogenous Leukemia

急性髓系白血病中髓系 Src 家族激酶的精确靶向

• 批准号: 10434077
• 负责人: Thomas E. Smithgall
• 金额: $ 42.69万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-15 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10434077
• 关键词: Acute Myelocytic Leukemia; Adult; Aftercare; Apoptosis; Architecture; Binding; Biological Assay; Bone Marrow; Bone Marrow Stem Cell; Cell Cycle Arrest; Cell Line; Cell Survival; Cells; Chemicals; Clinical; Clinical Trials; Codon Nucleotides; Combined Modality Therapy; Complex; Crystallization; Cytogenetics; Data; Development; Dose; Engineering; Engraftment; Evolution; FLT3 gene; Family member; Fibroblasts; Future; Hematologic Neoplasms; Hematopoietic Neoplasms; Human; In Vitro; Lead; Length; Leukemic Cell; Libraries; Maps; Modeling; Mus; Mutagenesis; Mutation; Myelogenous; Myeloid Cells; Oral Administration; Outcome; Pathogenesis; Pathway interactions; Patients; Pharmaceutical Preparations; Phosphotransferases; Population; Probability; Protein Tyrosine Kinase; Proteins; Reading Frames; Receptor Protein-Tyrosine Kinases; Recombinants; Relapse; Resistance; Rodent; Roentgen Rays; Sampling; Signal Pathway; Signaling Molecule; Signaling Protein; Specificity; Spleen; Structure; Testing; Therapeutic; Up-Regulation; Validation; X-Ray Crystallography; Xenograft Model; acquired drug resistance; acute myeloid leukemia cell; analog; base; cancer cell; cell growth; cell transformation; clinical development; cytotoxicity; drug action; efficacy evaluation; exome; in vivo; in vivo Model; inhibitor; kinase inhibitor; leukemic stem cell; mouse model; mutant; mutational status; nanomolar; next generation sequencing; novel therapeutic intervention; overexpression; patient derived xenograft model; peripheral blood; purge; relapse risk; resistance mechanism; resistance mutation; response; small molecule; small molecule inhibitor; src-Family Kinases; targeted treatment; transcriptome sequencing

Abstract. Acute myelogenous leukemia (AML) is a devastating hematologic cancer with limited treatment options. While diverse genetic changes are associated with AML, upregulation of tyrosine kinase signaling pathways is a common feature that offers opportunities for targeted therapy. These pathways involve the non- receptor tyrosine kinases Fes, Syk and the three Src-family kinases expressed in myeloid cells (Hck, Fgr, and Lyn). The focus of this application is a unique small molecule kinase inhibitor (TL02-59) with remarkable anti- AML efficacy. This compound potently suppressed the proliferation of bone marrow samples from twenty of twenty-six AML patients, with a striking correlation between inhibitor sensitivity and expression levels of the myeloid Src family kinases Fgr, Hck, and Lyn. No correlation was observed with Flt3 expression or mutational status, with the four most sensitive patient samples wild-type for Flt3. Kinome-wide target profiling and kinase assays demonstrated a narrow specificity profile for TL02-59, with picomolar potency against the myeloid Src- family member Fgr both in vitro and in cells. In a mouse xenograft model of AML, oral administration of TL02- 59 for just three weeks at 10 mg/kg completely eliminated leukemic cells from the spleen and peripheral blood while significantly reducing bone marrow engraftment. In this application, we propose to explore the mechanism of TL02-59 action, propensity for acquired resistance, and in vivo efficacy using mouse models of AML with the following Specific Aims: Aim 1. Determine the structural basis for TL02-59 potency and selectivity for the AML-associated Src-family kinase, Fgr. Preliminary data show that Fgr is inhibited by TL02-59 in vitro with an IC50 value of 30 picomolar, while other AML-associated kinases tested (including Flt3) are inhibited in the 100 nanomolar range. X-ray crystallography of Fgr in complex with TL02-59 and PCR-based codon mutagenesis of Fgr will be combined to explore the structural and functional basis of inhibitor specificity. Aim 2. Test the hypothesis that Fgr is the primary target for TL02-59 in AML. Engineered AML cell lines expressing TL02- 59-resistant Fgr mutants will be tested for loss of inhibitor sensitivity. In addition, AML cell populations will be evolved with acquired resistance to TL02-59, and resistance mechanisms explored by next-generation sequencing (whole exome and RNAseq). This unbiased approach may uncover unanticipated allosteric mechanisms of inhibition as well as additional targets for TL02-59 action in AML cells. Aim 3. Evaluate the efficacy of TL02-59 in mouse models of AML. Preliminary data demonstrate remarkable TL02-59 efficacy following short-term treatment in a mouse xenograft model using an AML cell line. This Aim will study inhibitor effects on long-term survival using AML patient-derived xenograft mice, including its ability to purge human leukemic stem cells from the bone marrow, and relate expression of the presumptive primary target for TL02-59 (Fgr) to the in vivo response. Successful outcomes with these in vivo models will make a compelling case for future clinical trials with TL02-59 or related analogs targeting Fgr for AML therapy.

摘要。急性髓性白血病（AML）是一种毁灭性的血液系统癌症，治疗方法有限