Thesis

Heterogeneity in Small Cell Lung Cancer

ABSTRACT Heterogeneity in Small Cell Lung Cancer By La France Daniels Jr Master of Science in Biology Small Cell Cancers of the Lung (SCLC) are poorly understood, rare, aggressive cancers that arise in many organs. These cancers represent a class of tumors that have undergone a reversal in differentiation losing specialized characteristics. In addition, theses tumors almost always recur from cells that behave like cancer stem cells. Histologically, SCLC are characterized by uniformly small-sized cells with a high nuclear to cytoplasmic ratio. Chemotherapy remains the standard form of therapy and patients usually exhibit an initial response to treatment. However, almost all patients relapse and the overall 5-year survival rate is only around 5%. For this project, I used biological tools to better understand the role of heterogeneity of cells in driving small cell cancers of the lung therapeutic resistance. First, I used a 3D lung organoid model of small cell lung cancer with an alginate based inverse opal scaffold and populations of cells from SCLC cell lines. Second, I used a mouse model of SCLC (p53fl/fl and Rbfl/fl) to examine the contribution of epithelial cells versus mesenchymal cells in SCLC tumor formation. I found that the 3D lung organoid model morphologically recapitulated the tumors seen in patients with sheets of small cells and neural rosette structures. I compared the expression patterns of neuroendocrine-specific-protein (NSP) marker Synaptophysin, serum marker Chromogranin A, and epithelial marker EpCAM, with non-neuroendocrine protein stem cell marker CD44, by immunofluorescence in the SCLC cell lines grown in 2D culture against those grown in the 3D lung organoid model. I found these markers were expressed in the same pattern in the 3D lung organoid model as in the patient tumor tissue but not in 2D cultures. This 3D lung organoid model of SCLC holds promise as a tool for elucidating the behavior of different SCLC cell populations in tumor initiation and chemotherapeutic resistance. To assess the contribution of epithelial cellular subpopulations in SCLC tumor formation, I isolated basal stem cells from the mouse tracheae epithelium and seed these cells into a matrigel culture for proliferation into tracheae spheres. Tracheae spheres in the next phase of this experiment are passaged, cultured and proliferated into differentiated epithelial cell populations. These newly differentiated cell populations are then transduced with Lentiviral-Cre to remove tumor suppressor genes Rb1 and Trp53. This will propagate the formation of SCLC tumors for thorough examination and study to better understand the role these heterogeneous subpopulations of cells play in small cell cancers of the lung therapeutic resistance.

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