Combined inhibition of CXCR4 and FLT3-ITD signaling in acute myeloid leukemia

描述（由申请人提供）：携带FLT3突变的复发性急性髓性白血病（AML）患者预后极差。我们已经证明微环境/白血病相互作用在白血病干细胞的化疗耐药中起主要作用，并且SDF-1a/CXCR4轴是这种相互作用的关键调节因子。我们最近发现索拉非尼是一种极好的FLT3-ITD抑制剂，具有很高的临床单药活性，在I期研究中包括完全缓解1,2。高水平的CXCR4与不良预后相关，FLT3突变上调CXCR45。我们最近报道了用FDA批准的CXCR4抑制剂AMD3100（Plerixafar）的类似物抑制CXCR4导致白血病细胞的动员和对Sorafenib6促凋亡作用的致敏。G- CSF切割SDF-1，下调CXCR4，与AMD3100在造血干细胞动员中具有高度协同作用，最近获得FDA批准。我们曾报道，接受AMD3100/G-CSF治疗的缓解期AML患者有大量AML细胞进入血液循环，这为白血病细胞动员原理提供了第一个证据。此外，我们观察到急性髓性白血病患者在CR和活动性疾病患者中优先动员正常细胞，这些患者接受AMD3100/G-CSF作为预备方案的一部分，然后进行SCT，进一步支持了这种治疗概念的有效性。值得注意的是，索拉非尼对正常的造血细胞没有毒性。基于这些发现，我们提出验证AMD3100/G-CSF通过破坏SDF-1a/CXCR4来动员白血病干细胞的假设，从而提高Sorafenib在FLT3突变的AML患者中的抗白血病活性。此外，我们将研究这种靶向治疗对非动员性AML原细胞的影响。我们将开展一项临床试验，以确定AMD3100、G-CSF和递增剂量的FLT3-ITD抑制剂索拉非尼对携带FLT3-ITD突变的AML患者的安全性和有效性，并研究AMD3100/G-CSF破坏SDF-1a/CXCR4相互作用的体内生物学效应。FLT3- itd突变患者可以有不同数量的细胞具有三种不同的FLT3基因型：杂合子FLT3- itd，纯合子FLT3- itd和野生型FLT3。因此，我们还将通过单细胞PCR确定该疗法在AMD3100/G-CSF动员期间和索拉非尼治疗后对携带这些基因型的细胞的影响。综上所述，这些研究将建立AMD3100/G- CSF/sorafenib的安全性和抗白血病活性，并提供白血病细胞动员和细胞信号抑制机制的综合评估。

DESCRIPTION (provided by applicant): The prognosis of patients with relapsed acute myeloid leukemia (AML) harboring FLT3 mutations is extremely poor. We have demonstrated that microenvironment/leukemia interactions play a major role in the chemoresistance of leukemic stem cells and that the SDF-1a/CXCR4 axis is a key regulator of this interaction. We recently discovered that Sorafenib is a superb FLT3-ITD inhibitor with high clinical single agent activity, including complete remissions in Phase I studies 1,2. High CXCR4 levels are associated with poor prognosis4, and FLT3 mutations upregulate CXCR45. We recently reported that inhibition of CXCR4 with an analogue of the FDA approved CXCR4 inhibitor AMD3100( Plerixafar) resulted in mobilization of leukemic cells and sensitization to the pro-apoptotic effects of Sorafenib6. G- CSF cleaves SDF-1, down-regulates CXCR4, was found to be highly synergistic with AMD3100 in mobilization hematopoietic stem cells9 and was recently approved by FDA. We have reported that AML patients in remission, who were treated with AMD3100/G-CSF, had massive egress of AML cells into the circulation, providing first proof of principle for leukemia cell mobilization10. In addition, we observed preferential mobilization of AML over normal cells, both in AML patients in CR and in patients with active disease who received AMD3100/G-CSF as part of a preparative regimen followed by SCT, further supporting the validity of this therapeutic concept. Of note, Sorafenib has no toxicity against normal hematopoietic cells. Based on these findings, we propose to test the hypothesis that mobilization of leukemic stem cells by disruption of SDF-1a/CXCR4 by AMD3100/G-CSF results in improved anti-leukemia activity of Sorafenib in AML patients with mutant FLT3. In addition, we will study the effects of this targeted therapy on non-mobilized AML blasts. We will conduct a clinical trial to determine the safety and efficacy of AMD3100, G-CSF and escalating doses of the FLT3-ITD inhibitor Sorafenib in patients with AML harboring FLT3-ITD mutations and study the in vivo biological effects of disrupting SDF-1a/CXCR4 interactions by AMD3100/G-CSF. Patients with FLT3-ITD mutations can have variable numbers of cells with three distinct FLT3 genotypes : heterozygous FLT3-ITD, homozygous FLT3-ITD and wild type FLT3. Therefore, we will also determine, by single-cell PCR, the effects of this therapy against cells carrying these genotypes, during mobilization with AMD3100/G-CSF and following treatment with sorafenib. Taken together, these studies will establish the safety and anti-leukemia activity of AMD3100/G- CSF/sorafenib and provide a comprehensive assessment of mechanisms involved in leukemia cell mobilization and inhibition of cell signaling. PUBLIC HEALTH RELEVANCE: Acute myelogenous leukemia is a mostly fatal disease, in particular in patients whose leukemia cells are characterized by a mutation in the FLT3 gene. We have recently found, that the drug Sorafenib, which is FDA approved for certain other cancers, is a superb inhibitor of cells carrying this mutation, resulting in complete clearance of leukemia cells from the blood and a reduction in bone marrow leukemia cells. We are proposing to combine sorafenib with a blocker of the chemokine receptor CXCR4, AMD3100(Plerixafor), in a clinical trial in relapsed/refractory patients with AML carrying the FLT3-ITD mutation with the aim of determining the safety and efficacy of this approach.