Developmental and Stem Cell Biology Program, The Hospital for Sick Children, Toronto, ON, Canada.
Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, ON, Canada.
BMC Cancer. 2019 Aug 30;19(1):864. doi: 10.1186/s12885-019-6018-1.
Bronchial carcinoids are neuroendocrine tumors that present as typical (TC) and atypical (AC) variants, the latter being more aggressive, invasive and metastatic. Studies of tumor initiating cell (TIC) biology in bronchial carcinoids has been hindered by the lack of appropriate in-vitro and xenograft models representing the bronchial carcinoid phenotype and behavior.
Bronchial carcinoid cell lines (H727, TC and H720, AC) were cultured in serum-free growth factor supplemented medium to form 3D spheroids and serially passaged up to the 3rd generation permitting expansion of the TIC population as verified by expression of stemness markers, clonogenicity in-vitro and tumorigenicity in both subcutaneous and orthotopic (lung) models. Acetazolamide (AZ), sulforaphane (SFN) and the AZ + SFN combination were evaluated for targeting TIC in bronchial carcinoids.
Data demonstrate that bronchial carcinoid cell line 3rd generation spheroid cells show increased drug resistance, clonogenicity, and tumorigenic potential compared with the parental cells, suggesting selection and expansion of a TIC fraction. Gene expression and immunolabeling studies demonstrated that the TIC expressed stemness factors Oct-4, Sox-2 and Nanog. In a lung orthotopic model bronchial carcinoid, cell line derived spheroids, and patient tumor derived 3rd generation spheroids when supported by a stroma, showed robust tumor formation. SFN and especially the AZ + SFN combination were effective in inhibiting tumor cell growth, spheroid formation and in reducing tumor formation in immunocompromised mice.
Human bronchial carcinoid tumor cells serially passaged as spheroids contain a higher fraction of TIC exhibiting a stemness phenotype. This TIC population can be effectively targeted by the combination of AZ + SFN. Our work portends clinical relevance and supports the therapeutic use of the novel AZ+ SFN combination that may target the TIC population of bronchial carcinoids.
支气管类癌是神经内分泌肿瘤，有典型（TC）和非典型（AC）两种变体，后者侵袭性更强，更具转移性。由于缺乏能够代表支气管类癌表型和行为的合适的体外和异种移植模型，对肿瘤起始细胞（TIC）生物学的研究受到了阻碍。
将支气管类癌细胞系（H727、TC 和 H720、AC）在无血清生长因子补充培养基中培养形成 3D 球体，并进行连续传代，最多传代至第 3 代，通过表达干性标志物、体外克隆形成能力和皮下及原位（肺）模型中的致瘤性来验证 TIC 群体的扩增。评估乙酰唑胺（AZ）、萝卜硫素（SFN）和 AZ+SFN 联合用药对支气管类癌 TIC 的靶向作用。
数据表明，与亲本细胞相比，支气管类癌细胞系第 3 代球体细胞显示出更高的药物耐药性、克隆形成能力和致瘤潜能，提示选择和扩增了 TIC 分数。基因表达和免疫标记研究表明，TIC 表达干性因子 Oct-4、Sox-2 和 Nanog。在肺原位模型中，支气管类癌细胞系衍生的球体以及由基质支持的患者肿瘤衍生的第 3 代球体显示出强大的肿瘤形成能力。SFN，特别是 AZ+SFN 联合用药，能有效抑制肿瘤细胞生长、球体形成，并减少免疫缺陷小鼠的肿瘤形成。
作为球体传代的人支气管类癌肿瘤细胞含有更高比例的表现出干性表型的 TIC。该 TIC 群体可被 AZ+SFN 联合靶向。我们的工作预示着临床相关性，并支持使用新型 AZ+SFN 联合用药治疗支气管类癌，该药物可能靶向支气管类癌的 TIC 群体。
