Analysis of the Signaling and Mechanical Cues Promoting Invasion in Melanoma

描述（由申请人提供）：转移性黑色素瘤是皮肤癌中最致命的一种，也是最具侵袭性的癌症类型之一，能够迅速从原发部位扩散。本提案旨在分析黑色素瘤细胞的遗传和信号状态变化以及细胞微环境对细胞侵袭程度和推定转移潜力的综合影响。我们分析的主要假设是，黑色素瘤细胞的侵袭性是由不同因素的组合决定的，主要来自细胞微环境的状态，包括ECM的结构和可溶性化学因素的存在。细胞微环境的改变，如ECM纤维的排列，可以使非转移性细胞变得更具侵袭性，而表面转移细胞的侵袭性可以通过环境变化和信号级联调控的靶向干预来调节。我们还假设细胞不仅可以通过分泌或降解ECM成分，还可以通过主动施加力，从而使ECM纤维在最近的锚点（包括脉管系统）的方向上对齐，从而积极地与微环境相互作用。这反过来可以产生更多的各向异性力，使细胞进一步变形ECM，并最终利用重新排列的ECM结构向血管迁移。我们将与ICBP@MIT密切合作测试这些假设，首先利用shRNA文库的新方法来干扰遗传靶标（目标1）以及广泛的环境扰动，然后使用ICBP@MIT上可用的最先进的统计分析工具来降低结果数据集的复杂性，将多种环境和遗传扰动与进攻性细胞行为联系起来（目标2）。我们将在这个过程中使用几种新的分析方法，增加数据采集和处理的可管理性，以及增强实验的仿生学性质。最后，我们将通过一系列测试来测试Aim 2中开发的模型所产生的预测，从细胞培养到组织构建再到模型组织水平（Aim 3）。我们预计，拟议的研究将揭示新的，以前未预料到的环境和遗传因素之间的相互作用，以控制黑色素瘤的转移事件，并可能提出新的方法来对抗这种致命的疾病。研究结果也可能与其他转移性癌症有关。

DESCRIPTION (provided by applicant): Metastatic melanoma is the deadliest form of skin cancer and one of the most aggressive cancer types, capable of rapid spreading out of the primary site. This proposal is aimed at analyzing the combined effect of changes in the genetic and signaling states of melanoma cells and the cell micro-environment on the degree of cell aggressiveness and putative metastatic potential. The main hypothesis underlying our analysis is that aggressiveness of melanoma cells is determined by a combination of different factors, with a major contribution from the state of cell microenvironment, including the structure of the ECM and the presence of soluble chemical factors. Alteration in the cell micro-environment, such as alignment of ECM fibers, can permit an otherwise non-metastatic cell to become more aggressive, whereas the aggressiveness of ostensibly metastatic cell can be moderated by environmental changes and targeted intervention in regulation of signaling cascades. We also hypothesize that cells can actively interact with the micro-environment not only by secreting or degrading ECM components, but by actively exercising forces on and thus aligning ECM fibers in the direction of a nearest anchor point, including the vasculature. This can in turn generate more anisotropic forces allowing cells to deform ECM even more and ultimately take advantage of the re-aligned ECM structure to migrate towards blood vessels. We will test these hypotheses in close collaboration with ICBP@MIT, first taking advantage of the new methodologies using shRNA libraries to perturb genetic targets (Aim 1) along with a broad range of environmental perturbations and then use the state of the art statistical analysis tools available at ICBP@MIT to reduce complexity of resulting datasets, linking multiple environmental and genetic perturbation to aggressive cell behavior (Aim 2), We will use several novel assays in the process, increasing manageability of data acquisition and processing, as well as enhancing the biomimetic nature of experimentation. Finally, we will test the predictions arising from the models developed in Aim 2 through a series of tests, ranging from cell culture to tissue construct to model tissue levels (Aim 3). We anticipate that the proposed research will reveal new, previously unanticipated interplay between environmental and genetic factors in control of metastatic events in melanoma and potentially suggest new ways of battling this deadly disease. The results might also have relevance to other metastatic cancers. PUBLIC HEALTH RELEVANCE: Early detection is further hampered by the lack of biomarker-based detection tests. One of the critical reasons for this situation is the lack of understanding of the mechanisms underlying the metastatic switch in melanoma and the mechanisms allowing metastatic cells to successfully and quickly navigate all the hurdles associated with the metastatic spread, including effective migration to and intravasation into the blood vessels. This project is aimed at filling this critical gap.