Defining pro-tumourigenic neutrophil phenotypes and tumour-neutrophil interactions to identify targets for therapy in metastatic colorectal cancer

死于结直肠癌的患者是由于转移性疾病。结肠直肠癌最常见的转移部位是肝脏。如果能早期发现肝转移，那么患者在手术后会得到最好的结果。不幸的是，有一些病人在肝切除术后表现很差。这项研究旨在识别手术后表现不佳的患者，更好地了解他们的疾病生物学，并为他们未来的癌症提供个性化靶向治疗的希望。为了开发新的治疗方法，我对结直肠癌患者进行了评估，这些患者在切除结直肠癌肝转移灶后早期复发，癌症相关生存率较差。人们普遍认为，癌细胞周围高水平的支持性结缔组织转移患者预后较差，我们对患者队列的分析进一步支持了这一点。我的研究表明，这些患者的转移瘤中有大量的中性粒细胞，这是最常见的白细胞。我使用模拟人类胰腺癌和结直肠癌的小鼠模型表明，在这些转移性癌症中，中性粒细胞在支持疾病进展中发挥重要作用，这表明这种现象可能并非晚期结直肠癌所独有。引人注目的是，干扰到肝脏的中性粒细胞减少了肝脏肿瘤的数量，当给予激活免疫系统的治疗时，我可以提高癌症小鼠模型的存活率。直到最近的研究才表明，中性粒细胞可以以不同的方式发挥作用，在健康和疾病中扮演不同的角色。这些细胞具有很强的可塑性，很容易根据来自癌细胞的信号改变它们的表达，因此可以根据不同的环境显示出促癌和抗癌的特性。正是中性粒细胞的这种可塑性，可能有助于调节免疫系统行为的治疗。仍然存在的问题是：中性粒细胞如何支持转移性癌症的进展？结直肠癌转移灶中存在的中性粒细胞表达哪些标记物？与健康人相比，这些标记物如何支持它们在癌症进展中的功能？一个重要的考虑是中性粒细胞对正常免疫抵抗疾病很重要，通过特异性靶向肿瘤促进中性粒细胞，未来的治疗可能会与先天免疫解决困难。在本提案中，我将通过使用肿瘤产生的分子因子测序，根据患者的疾病生物学对患者进行分型，详细讨论哪些患者具有与转移相关的大量中性粒细胞。利用由行业合作伙伴Nanostring开发的令人兴奋的新技术，我将通过对肝脏肿瘤中的中性粒细胞种群进行深入的空间定向评估，来解决存在哪种类型的中性粒细胞的问题。一旦我确定了与侵袭性疾病生物学相关的中性粒细胞群体，我将使用我在实验室开发的最先进的结肠直肠癌转移小鼠模型，在与人类疾病最相关的生命系统中去除不同群体的中性粒细胞，并在转移发生时使用复杂的小鼠显微镜评估其对转移过程的影响。这些评估将证明这些有针对性的方法对中性粒细胞的行为、与免疫细胞和肿瘤细胞的相互作用以及最终对结直肠癌扩散机制的影响。在高通量的小鼠模型中，我测试了转移性肿瘤系统中中性粒细胞上敲除的许多分子因子，我将能够识别中性粒细胞上的潜在靶标，这些靶标可用于未来的临床试验治疗。

Patients that die from colorectal cancer do so as a result of metastatic disease. The most common site for colorectal metastases is the liver. If liver metastases can be detected early then patients receive the best outcomes following surgery. Sadly there are groups of patients that do very poorly following liver resection. This research looks to identify the patients that do poorly following surgery, understand their disease biology better and provide them with the hope of individually targeted therapy for their cancer in the future.To develop new therapies I have performed assessment of patients with colorectal cancer that have early recurrence and poor cancer related survival following resection of their colorectal liver metastases. It is accepted that patients with high levels of supportive connective tissue surrounding cancer cells, within their metastases have poorer outcomes and this is further supported by analysis in our cohort of patients. My work has shown that these patients have very high numbers of neutrophils, the most common white blood cell, within their metastases. Using mouse models that mimic human pancreatic and colorectal cancer I have shown that in these metastatic cancers neutrophils play an important role in supporting progression of disease, suggesting this phenomenon may not be unique to advanced colorectal cancer. Strikingly, interfering with neutrophils travelling to the liver reduces the number of tumours in the liver and when therapies that activate the immune system are given I can improve survival of the cancer mouse models. Only recently studies have suggested that neutrophils can behave in different fashions, adopting different roles in health and disease. These cells are very plastic and readily change what they express in response to signals from cancer cells and therefore can demonstrate both pro and anti-cancer properties depending on context. It is this plasticity of neutrophils that may lend itself to therapies that modulate the behaviour of the immune system.The questions that remain are: how do neutrophils support progression of metastatic cancer?; What markers do neutrophils that exist in colorectal cancer metastases express and how do these support their function in cancer progression compared with in health? One important consideration is that neutrophils are important for normal immunity to fight off disease and by specifically targeting tumour promoting neutrophils future therapies may negotiate difficulties with innate immunity.In this proposal I will address in detail which patients have high numbers of neutrophils associated with metastases by subtyping patients according to their disease biology using sequencing of molecular factors produced by the tumours. Using exciting new technologies that have been developed by an industry partner, Nanostring, I will then resolve the question of which types of neutrophils are present by performing in-depth spatially orientated assessment of neutrophil populations in liver tumours. Once I have identified neutrophil populations associated with aggressive disease biology I will use a state of the art mouse model of colorectal cancer metastasis I have developed in the laboratory to remove different populations of neutrophils within a living system most relevant to the human disease and assess the impact on the metastatic process using sophisticated microscopy of the mouse as metastases develop. These assessments will demonstrate the impact of these targeted approaches on the behaviour of neutrophils, interactions with immune and tumour cells and ultimately on mechanisms by which colorectal cancer spreads. Using the mouse model in a high throughput manner, where I test a number of molecular factors knocked out on neutrophils within the metastatic tumour system, I will be able to identify potential targets on neutrophils that could be used for therapy in future clinical trial.