Development of a Mouse Model to Study Targeted Therapy to Prevent CNS Invasion by Pediatric T-ALL

标题：建立一种小鼠模型来测试靶向治疗以防止儿童T-T侵袭中枢神经系统 全 摘要 白血病是儿科最常见的癌症，其中急性淋巴细胞性白血病(ALL)最为常见 儿童白血病类型。在15%-20%的儿童T细胞ALL(T-ALL)患者中，癌症侵犯中枢神经系统 中枢神经系统(CNS)，这个部位受到血脑屏障的保护，不受全身化疗的影响。侵袭性白血病 细胞是储存库，可以在治疗后出现并重新进入循环，导致T-ALL复发。至 预防复发，儿科患者接受头颅放射治疗和毒性化疗，并注射 直接进入中枢神经系统。不足为奇的是，这些治疗方法会导致病态的终身副作用，如减少 智力、发育迟缓和中枢神经系统的继发性癌症。一般情况下，T-ALL在进入中枢神经系统之前 诊断，目前尚不清楚是否在T-ALL进入中枢神经系统后阻止他们进入， 提供任何治疗益处。该方案的主要三个目标是：1)建立T-T小鼠模型。 所有这些都概括了人类T-ALL，在其中我们可以看到活着的动物的细胞2)以确定是否阻断T-ALL 细胞进入CNS，防止T-ALL进一步侵袭CNS，以及3)确定T-ALL是否已经入侵CNS 中枢神经系统最终会清除中枢神经系统，并恢复流通。我们的长期目标是减少或消除 用于破坏T-ALL患者的鞘内或颅脑治疗。我们的研究是基于人体异种移植 T-ALL小鼠模型，发现在白血病发生过程中，转录因子NOTCH1的表达 在80%以上的儿童T-ALL中表达，诱导CC趋化因子受体7(CCR7)的表达。这项研究 发现CCR7在T-ALL中被其一个配体激活，CCL19促进T-ALL的趋化迁移到 中枢神经系统。我们的建议是基于从莫妮卡·贾斯蒂斯博士那里获得的小鼠模型，该模型使用了可诱导的CCR7(+)rosa26 FLOXED-STOP-PRDM14(R26PR)基因座。当与MX1-Cre杂交时，这些R26PR小鼠会患上白血病 NOTCH1过表达，侵袭中枢神经系统。随后繁殖这些带有rosa26的小鼠-停止 荧光素酶小鼠将导致Cre重组酶在T-ALL细胞中诱导表达生物发光荧光素酶。要确认 CCR7在中枢神经系统入侵中的作用，这些R26PR小鼠将与我们自己的CCR7条件基因敲除杂交 小鼠(CCR7F/f)，产生R26PRDM14/CCR7f/f/MX1-Cre小鼠。这一代鼠标的目标是1。 确定哪些免疫细胞在进入CNS时与T-ALL细胞共定位是AIM2，并确定是否阻断 CCR7使用CCL19拮抗剂CCL198-83防止或显著减少T-ALL中枢神经系统的侵袭，清除T-ALL中枢神经系统 并提高小鼠存活率是目标3。最终，如果成功，这种小鼠模型将为开发新的 可阻断T-ALL中枢神经系统侵袭，而不会对患者造成长期中枢神经系统损害的多肽疗法及其研究 中枢神经系统的侵袭机制。

TITLE: Development of a Mouse model to test targeted therapy to prevent CNS Invasion by Pediatric T- ALL Abstract Leukemia is the most frequent pediatric cancer with acute lymphoblastic leukemia (ALL) being the most common leukemia type in children. In 15-20% of pediatric T-cell ALL (T-ALL) patients, the cancer invades the Central Nervous System (CNS), a site which is protected from systemic chemotherapies by the blood brain barrier. The invading leukemic cells are reservoirs, which can emerge and re-enter the circulation following treatment and cause T-ALL relapse. To prevent relapse, pediatric patients are treated with cranial irradiation and toxic chemotherapies, which are injected directly into the CNS. It is not surprising that these treatments can lead to morbid life-long side effects such as reduced intelligence, stunted growth, and secondary cancers in the CNS. Generally, the T-ALL has entered the CNS prior to diagnosis, and it is unclear if blocking the entry of T-ALL into the CNS at that point, after they have already entered, provides any therapeutic benefit. The overarching three goals of this proposal are: 1) to develop a mouse model of T- ALL that recapitulates human T-ALL, in which we can see the cells in living animals 2) to determine if blocking T-ALL cells from entering the CNS, prevents further T-ALL CNS invasion and 3) to determine if the T-ALL that has invaded the CNS will eventually clear the CNS and return to the circulation. Our long-term goal is to reduce or eliminate the need for damaging intrathecal or cranial treatments in patients with T-ALL. Our studies are based on a human xenograft mouse model of T-ALL, which revealed that during leukemogenesis, expression of NOTCH1, a transcription factor which is expressed in more than 80% of pediatric T-ALLs, induces expression of CC chemokine receptor 7 (CCR7). The study revealed that activation of CCR7 in T-ALL by one of its ligands, CCL19 promotes chemotactic migration of T-ALL into the CNS. Our proposal is based on a mouse model obtained from Dr. Monica Justice which uses an inducible CCR7(+)ROSA26 floxed-stop-PRDM14 (R26PR) locus. When crossed to an MX1-Cre these R26PR mice develop leukemia in response to overexpression of NOTCH1, which invades the CNS. Subsequent breeding of these mice with ROSA26 floxed-stop luciferase mice will result in Cre-recombinase induced expression of bioluminescent luciferase in T-ALL cells. To confirm a role for CCR7 in the invasion of the CNS, these R26PR mice will be crossed with our own CCR7 conditional knockout mice (CCR7floxed/floxed (CCR7f/f)) to produce R26PRDM14/CCR7f/f/MX1-CRE mice. Generation of this mouse is aim 1. Determining which immune cells co-localize with T-ALL cells upon entry into the CNS is aim2 and determining if blocking CCR7 using the CCL19 antagonist, CCL198-83 prevents or significantly reduces T-ALL CNS invasion, clears the CNS of T-ALL and increases mouse survival is aim 3. Ultimately, if successful, this mouse model will provide a tool for developing novel peptide therapies that can block T-ALL CNS invasion, without causing long-term CNS damage in patients and to study mechanisms of CNS invasion.