摘要1

食管癌是发病率排第七，死亡率排第六的恶行肿瘤，是全人类主要的健康杀手之一。在我国，食管鳞癌为食管癌的主要类型，约占90%。较早分期食管鳞癌通过手术治疗多能取得较好效果，但由于早期食管鳞癌症状不明显，大多数患者就诊时已处于中晚期。新辅助治疗联合手术治疗是局部晚期食管鳞癌的首选治疗方式。虽然新辅助治疗给这些患者带来新的希望，但不可避免会发生耐药，故探索食管鳞癌发生发展及耐药机制显得尤为重要。

成纤维细胞生长因子 (FGF) 家族可以调节机体内的多种发育过程，并且被证实在多种肿瘤的发生发展中起到关键作用。FGF19作为其中的重要一员，其对肿瘤发生发展作用的研究也日渐增多。最近的研究表明，FGF19对肝癌、前列腺癌、乳腺癌、甲状腺癌、肺癌等多种肿瘤的发生发展均有促进作用，是比较明确的促癌因子，但其在食管鳞癌中的作用研究较少。

本部分论文旨在探索FGF19在食管鳞癌发生发展和新辅助化疗耐药中作用及其相关机制。我们首先进行了泛癌分析，发现FGF19在多种肿瘤中高表达，且与不良预后密切相关。接着我们分析了FGF19在食管鳞癌中的表达情况，基于TCGA和GEO数据库中数据发现FGF19基因在食管鳞癌中异常高表达且与不良预后显著相关。此外，在155例在我院接受外科治疗食管鳞癌患者的测序数据中，相比于癌旁组织，FGF19的mRNA在食管鳞癌癌组织中显著增高。通过分析组织芯片103例及我院72例食管鳞癌患者的信息，结果提示食管鳞癌癌组织中FGF19的表达与淋巴结转移和TNM分期显著相关，且FGF19蛋白高表达可能是食管鳞癌患者预后的独立危险因子。另外，我们发现在接受新辅助化疗的食管鳞癌患者中，疗效PD/SD患者的FGF19表达水平明显高于疗效CR/PR的患者，差异具有统计学意义。并且FGF19高表达与接受新辅助化疗食管鳞癌患者不良预后显著相关。功能上，过表达FGF19可以促进食管鳞癌的的增殖、迁移、侵袭、抗凋亡和成瘤以及化疗抵抗的能力。机制方面，通常FGF19与FGFR4相互作用调控下游信号通路以参与肿瘤的发生发展，我们的实验结果发现FGF19-FGFR4通过调控食管鳞癌生物学功能的机制是通过PI3K/AKT信号通路实现的。

综上所述，FGF19可能是多种肿瘤的新型预后标志物和癌基因。进一步探讨FGF19在食管鳞癌中的作用，发现FGF19在食管鳞癌中高表达且与其不良预后及新辅助化疗抵抗有关，过表达FGF19可促进食管鳞癌的增殖、迁移、侵袭、抗凋亡、成瘤及化疗抵抗的能力，这一过程可能是通过激活PI3K/AKT信号通路实现的。

摘要2

食管鳞癌被认为是一种高度恶性的肿瘤，并且容易发生淋巴结转移。淋巴转移是胸段食管鳞癌最常见的转移途径之一，也是重要的预后影响因子之一。目前，单纯手术治疗和以手术治疗为基础的综合治疗仍为可切除食管鳞状细胞癌的最主要治疗方式。手术治疗被推荐的标准治疗方式包括食管根治手术和区域淋巴结清扫术。左气管支气管旁（4L）淋巴结，位于主动脉顶点与隆突之间，被定义为食管癌的区域淋巴结。近些年，气管旁淋巴结清扫已经成为区域淋巴结清扫的重点，4L淋巴结的清扫也逐渐引起学者的关注。虽然4L淋巴结被认为是食管鳞癌的区域淋巴结，但是，考虑到解剖的局限和可能引起的并发症，对于所有可切除的食管鳞癌进行常规4L淋巴结清扫仍存在争议。目前，很少有研究报道4L淋巴结清扫和4L淋巴结转移对食管鳞癌患者的预后影响。因此，我们开展了这项研究，以评估4L淋巴结清扫和4L淋巴结转移对胸段食管鳞癌患者生存的影响。并且我们进行了多因素分析以识别4L淋巴结转移的独立危险因素。以期为食管鳞癌4L淋巴结清扫决策和筛选出合适的人群提供临床依据。

本研究从回顾性数据库中收集2010年1月至2018年12月在胸外科接受治疗的3522例食管癌患者的临床资料。所有患者均接受了根治性食管癌手术联合系统淋巴结清扫术。通过计算标准差值 (Standardized differences，SD) 来评估两组变量的均衡性，并且将值< 0.10 视为变量平衡的标准。另外，研究的主要终点是总生存期（Overall survival，OS），我们采用 Kaplan-Meier方法估计生存曲线，并采用Log-rank检验进行生存差异比较。将相对危险因素纳入多变量Cox生存分析，以确定患者预后的独立危险因素。此外，为了减少其他混杂变量的干扰，我们进行了倾向评分匹配分析（Propensity score matching，PSM）。最后，我们进行单变量和多变量logistic回归分析以识别4L淋巴结转移的独立危险因素。

在研究队列中，608例（17.3%）患者接受了4L淋巴结清扫，其中45 例（7.4%）患者出现 4L 淋巴结转移。本研究的目的是探究4L淋巴结清扫和4L淋巴结转移对胸段食管鳞癌患者生存的影响。首先，我们对比了接受4L淋巴结清扫和未清扫患者之间的生存差异。分析结果提示，接受4L淋巴结清扫患者的5年OS率为69.3%，未接受淋巴结清扫患者的5年OS率为61.7%。接受4L淋巴结清扫患者的5年OS率高于未接受淋巴结清扫的患者，差异具有统计学意义（P=0.001）。PSM后，接受4L淋巴结清扫患者的5年OS率为68.2%，未进行淋巴结清扫患者的5年OS率为64.6%。接受4L淋巴结清扫患者的长期生存率显著高于未进行淋巴结清扫的患者，差异具有显著性（P=0.012）。同时，我们在所有食管鳞癌患者（3522例）中进行了多变量 Cox 生存分析。分析结果提示4L淋巴结清扫是较好预后的独立影响因素（HR：0.850，95%CI：0.725-0.996，P=0.045）。另外，在接受4L淋巴结清扫的患者人群中，我们对比了4L淋巴结转移和无4L淋巴结转移患者之间的生存差异。分析结果提示，4L淋巴结转移患者的5年OS率为45.5%，无4L淋巴结转移患者的5年OS率为71.1%。4L淋巴结转移患者的5年OS率低于无4L淋巴结转移的患者，差异具有统计学意义（P < 0.001）。为了减少其他混杂变量的干扰，我们进行了倾向评分匹配。PSM后，两组患者的基线特征的无显著性差异（SD<0.10）。4L淋巴结转移患者的5年OS率为40.5%，未发生4L淋巴结转移患者的5年OS率为62.2%。4L淋巴结转移患者的长期生存率显著低于未发生4L淋巴结转移的患者，差异具有显著性（P=0.029）。同时，我们在所有接受4L淋巴结清扫的食管鳞癌患者（608例）中进行了多变量Cox生存分析。结果提示4L淋巴结转移是预后的独立影响因素（HR：1.932，95%CI：1.245-2.999，P=0.003）。最后，为了识别4L淋巴结转移的独立危险因素，我们在接受4L淋巴结清扫的食管鳞癌患者（608例）中，进行了多变量logistic回归分析。结果提示肿瘤分化程度（HR：2.495，95%CI：1.326-4.695，P=0.005）和临床T分期（HR：1.281，95%CI：1.065-2.209，P=0.028）是4L淋巴结转移的独立预测因素。

综上所述，我们的研究首次揭示了4L淋巴结转移与胸段ESCC患者的预后较差显著相关，4L淋巴结清扫可能改善患者的预后。多变量logistic分析显示临床T分期和肿瘤分化程度是4L淋巴结转移的独立预测因素。对于具有高危因素的患者，应推荐进行常规4L淋巴结清扫术。

Abstract 1

Esophageal carcinoma is a malignant disease with the seventh incidence and sixth leading cause of cancer-related mortality worldwide, and is one of the major health killers of all mankind. In China, esophageal squamous cell carcinoma is the main type of esophageal carcinoma, accounting for 90% of all patients. Usually, surgical strategy is chosen to treat early stage tumors which can often achieve satisfactory results. However, due to the insignificant symptoms of early esophageal squamous cell carcinoma, most patients are already in locally advanced stages when they seek help. Neoadjuvant therapy combined with surgery is the preferred treatment for locally advanced esophageal squamous cell carcinoma. Although neoadjuvant therapy brings new hope to these patients, drug resistance will inevitably occur, so it is particularly important to explore the development and drug resistance mechanism of esophageal squamous cell carcinoma.

The fibroblast growth factor (FGF) family can regulate various developmental processes in the body and has been shown to play a key role in the occurrence and development of various tumors. As an important member of FGFs, the role of FGF19 on tumorigenesis and development is also increasing recently. Studies have shown that FGF19 can promote the occurrence and development of various tumors such as liver cancer, prostate cancer, breast cancer, thyroid cancer and lung cancer. FGF19 is a relatively clear tumor-promoting factor, but its role on the mechanism of esophageal squamous cell carcinoma development is still unclear due to limited studies.

This part of our study aims to explore the role of FGF19 on the occurrence, development and neoadjuvant chemotherapy resistance of esophageal squamous cell carcinoma and its related mechanisms. Firstly, we conducted a pan-cancer analysis and found that FGF19 was highly expressed in a variety of tumors and correlated with their poor prognosis. Secondly, we analyzed the expression level of FGF19 in esophageal squamous cell carcinoma, through analyzing the data in TCGA and GEO databases. We found that FGF19 gene was abnormally highly expressed in esophageal squamous cell carcinoma and was associated with its poor prognosis. Thirdly, in the sequencing data of 155 patients with esophageal squamous cell carcinoma who received surgical treatment in our hospital, the mRNA expression of FGF19 was significantly increased in esophageal squamous cell carcinoma tissues compared with adjacent tissues. By analyzing the information of 103 cases of tissue microarray and 72 cases of esophageal squamous cell carcinoma patients in our hospital, the results showed that the expression of FGF19 in esophageal squamous cell carcinoma was significantly correlated with lymph node metastasis and TNM stage, and the high expression of FGF19 protein may be an independent risk factor for the prognosis of patients with esophageal squamous cell carcinoma. In addition, we found that in all patients with esophageal squamous cell carcinoma who received neoadjuvant chemotherapy, the expression level of FGF19 in patients with response PD/SD was significantly higher than that with response CR/PR, and the difference was statistically significant. And high expression of FGF19 is significantly associated with poor prognosis in patients with esophageal squamous cell carcinoma receiving neoadjuvant chemotherapy. Fourthly, the biological functions of FGF19 in ESCC cells was studied, the results showed that overexpression of FGF19 can promote the proliferation, migration, invasion, anti-apoptosis, tumorigenesis and chemoresistance ability of esophageal squamous cell carcinoma. Finally, we investigated the role of FGF19 on the mechanism of esophageal squamous cell carcinoma development. Usually, FGF19 interacts with FGFR4 to regulate downstream signaling pathways to participate in the occurrence and development of tumors. Our results showed that the mechanism that PI3K/AKT signaling pathway participate in the process of FGF19-FGFR4 regulating the biological function of esophageal squamous cell carcinoma.

Taken together, FGF19 may be a novel prognostic marker and oncogene for multiple tumors. We further explored the role of FGF19 in esophageal squamous cell carcinoma, and found that FGF19 is highly expressed in esophageal squamous cell carcinoma and is related to chemotherapy resistance and poor prognosis. In vivo and in vitro experiments showed that overexpression of FGF19 can promote the proliferation, migration, invasion, anti-apoptosis, tumorigenesis and chemoresistance ability of esophageal squamous cell carcinoma. And the regulation process may be achieved by activating the PI3K/AKT signaling pathway.

Abstract 2

Esophageal squamous cell carcinoma (ESCC) is considered a highly malignant tumor and prone to lymph node metastasis (LNM). The presence of LNM is associated with the poor prognosis of patients with thoracic ESCC. Currently, the standard treatment strategy for resectable ESCC involves esophagectomy combined with systematic lymphadenectomy. The left tracheobronchial lymph node (4L) is considered regional lymph node (LN) for thoracic ESCC, but routine prophylactic 4L LN dissection performed for all resectable ESCC is still under debate. Station 4L LN dissection is more technically challenging and time-consuming than other lymph node stations because of anatomic limitations. They are located deeply within the subaortic region close to the left recurrent laryngeal nerve, aortic arch, left pulmonary artery and left main bronchial membrane. Currently, few studies reported the clinical significance of 4L LN dissection and station 4L LNM in patients with ESCC. Thus, we carried out this study to evaluate the effect of 4L LN dissection and station 4L LNM on the survival of patient with ESCC, and analyze the risk factors of station 4L LNM.

From January 2010 to December 2018, 3522 patients with EC who underwent radical esophagectomy combined with lymphadenectomy were collected from the retrospective institutional database. Clinical characteristics, including age, sex, smoking index, alcohol abuse, degree of differentiation, tumor length, tumor location, clinical T stage and N stage, were analyzed. Categorical variables were presented as absolute number and percentage, and Chi-square or Fisher’s exact tests were conducted to compare patient characteristics. The balance of variables between two groups were evaluated by calculating the standardized differences (SD), and a value <0.10 was considered as criteria of sufficient balance. The primary end point was overall survival (OS). Survival curves were estimated by a Kaplan-Meier method and log-rank test was conducted to compare differences. Multivariate Cox survival analysis was used to identify independent risk factors for prognosis. Moreover, the propensity score matching (PSM) method was then performed to balance patient characteristics between two groups. Finally, univariable and multivariable logistic regression analysis were performed to evaluate the relation between risk factors and station 4L LNM.

In the whole cohort, 608 (17.3%) patients underwent station 4L LN dissection. Of patients underwent 4L LN dissection, 45 patients had 4L LNM (7.4%). We carried out this study to evaluate the effect of 4L LN dissection and station 4L LNM on the survival of patient with ESCC. The 5-year OS rates were 69.3% in patients with station 4L LN dissection and 61.7% in patients without, indicating a significant difference (P =0.001). After PSM, patients who received the 4L LN dissection had a superior long-term survival compared with those did not (5-year OS rate: 68.2% vs. 64.6%, P =0.012). Simultaneously, multivariate Cox survival analysis was performed in the original cohort (3522 patients), and the results showed that station 4L LN dissection was an independent influencing factors for better prognosis (HR: 0.850, 95%CI: 0.725-0.996, P =0.045). In addition, in the patient population who received 4L LN dissection, the OS of patients with station 4L LNM was significantly poorer than those without station 4L LNM (5-year OS rate: 45.5% vs. 71.1%, P <0.001). After balance in baseline characteristics, the 5-year OS rates were 40.5% in patients with station 4L LNM and 62.2% in patients without, indicating a significant poorer long-term survival in patients with station 4L LNM (P =0.029). Simultaneously, multivariate Cox survival analysis was conducted in all patients who received 4L LN dissection (608 patients), and the results showed that station 4L LNM was an independent risk factors for poorer prognosis (HR: 1.932, 95%CI: 1.245-2.999, P =0.003). Finally, all patients (608 patients) who underwent station 4L LN dissection were included in subgroup analysis to identify risk factors for 4L LNM. Multivariate logistic analysis revealed that clinical T staging (OR, 1.281; 95% CI, 1.065-2.209; P =0.028) and tumor differentiation (OR, 2.495; 95% CI, 1.326-4.695; P =0.005) were independent risk factors for station 4L LNM.

In this study, station 4L LNM was associated with a poorer prognosis of patients with thoracic ESCC, but station 4L LN dissection seems to be associated with a more favorable prognosis. Multivariable analysis showed that clinical T stage and tumor differentiation were independent risk factors for station 4L LNM. For patients with high risk, routine prophylactic 4L LN dissection should be recommend.