Targeting microtubule stabilization to reduce breast tumor metastasis

靶向稳定微管以减少乳腺肿瘤转移。 乳腺肿瘤细胞通过非黏附的微环境转移到远处的器官，例如 血液或淋巴管。然而，对其动态行为和药物反应知之甚少。 非粘附性肿瘤细胞，这是由于成像非粘附性细胞而不会因细胞漂移而模糊的挑战。 PI的实验室在非粘附性肿瘤细胞表面发现了独特的微触手(McTns)， 促进活体小鼠肺毛细血管内循环肿瘤细胞(CTCs)的聚集和滞留。 这项修订后的研究将检验2微管稳定的治疗靶点的假设 机制(去酪氨酸化、乙酰化)可以提供一种抑制肿瘤转移的新策略 并超越目前针对肿瘤细胞生长的非选择性MT靶向药物。 这一假说的预测将在以下特定的目标上用共焦和电子进行检验 显微镜、CTC转移的全动物成像，以及在患者肿瘤细胞和CTC上测试药物。 具体目的1：抑制微管蛋白降解以减少McTns、干细胞特性和转移。 A)通过靶向TTL或TCP改变微管蛋白的酪氨酸降解，并测量对转移潜能/EMT的影响。 B)测试姜黄素和单烯内酯类似物对EMT、干细胞特性和转移的疗效。 C)确定微管稳定疗法(紫杉醇、伊克沙比隆)是否促进转移潜能。 具体目标2：确定改变微管蛋白乙酰化对转移表型的影响。 A)基因改变微管蛋白乙酰化(ATAT1、K40R、HDAC6)并测量对小鼠转移的影响。 B)分析微管蛋白乙酰化改变细胞中的肿瘤细胞力学和McTN结构。 C)研究通过抑制HDAC使微管蛋白乙酰化水平升高是否会增加转移潜能。 具体目标3：靶向小鼠和人类患者活体CTCs的微管稳定机制。 A)在异种移植、PDX和活体患者CTC上测试2种优先药物，以减少McTns和CTC集群。 B)将CTC转移与MT稳定剂和MT干扰剂在分离和组合方面进行比较。 一种创新的微流控细胞连接系统将用于快速确定活体患者的药物反应 CTCS在不到一小时内完成。纳入许多FDA批准的疗法将增加潜在的 快速转化该项目的成果，以影响转移性乳腺癌的临床治疗。

Targeting microtubule stabilization to reduce breast tumor metastasis. Breast tumor cells metastasize to distant organs through non-adherent microenvironments, such as the bloodstream or lymphatics. However, very little is known about the dynamic behavior and drug responses of non-adherent tumor cells, due to the challenges of imaging non-adherent cells without blurring from cell drift. The PI's lab discovered unique microtentacles (McTNs) on the surface of non-adherent tumor cells that promote the aggregation and retention of circulating tumor cells (CTCs) in the lung capillaries of living mice. This revised study will test the hypothesis that therapeutic targeting of 2 microtubule stabilization mechanisms (detyrosination, acetylation) can provide a novel strategy to suppress tumor metastasis and move beyond the current non-selective MT-targeting drugs that are aimed at tumor cell growth. Predictions of this hypothesis will be tested in the following specific aims with confocal and electron microscopy, whole-animal imaging of CTC metastasis and by testing drugs on patient tumor cells and CTCs. Specific Aim 1: Inhibit tubulin detyrosination to reduce McTNs, stem cell characteristics and metastasis. A) Alter tubulin detyrosination by targeting TTL or TCP and gauge effects on metastatic potential/EMT. B) Test efficacy of Curcumin and Parthenolide analog on EMT, stem cell characteristics and metastasis. C) Define whether microtubule-stabilizing therapies (Paclitaxel, Ixabepilone) promote metastatic potential. Specific Aim 2: Determine the effects of altering tubulin acetylation on metastatic phenotypes. A) Genetically alter tubulin acetylation (ATAT1, K40R, HDAC6) and gauge effects on metastasis in mice. B) Analyze tumor cell mechanics and McTN structure in cells with altered tubulin acetylation. C) Examine whether elevation of tubulin acetylation by HDAC inhibition increases metastatic potential. Specific Aim 3: Target microtubule stabilization mechanisms in live CTCs from mice and human patients. A) Test 2 prioritized drugs on xenografts, PDX and live patient CTCs to reduce McTNs and CTC clusters. B) Compare CTC metastasis with MT-stabilizing and MT-disrupting agents in isolation and combination. An innovative microfluidic cell tethering system will be used to rapidly determine drug responses in live patient CTCs in less than one hour. Inclusion of numerous FDA-approved therapies will increase the potential to rapidly translate the outcomes of this project to impact the clinical treatment of metastatic breast cancer.