| Non-small cell lung cancer (NSCLC), which includes squamous lung cancer, adenocarcinoma and large cell carcinoma, comprises 80% of lung cancers. Bronchioloalveolar carcinoma (BAC) is a subtype of adenocarcinoma with growth along the alveolar septa and without evidence of stromal, vascular, or pleural invasion. Although only ~4% of lung cancers meet this definition, up to 20% of lung cancers comprise a heterogeneous group of tumors with BAC histology mixed with varying populations of invasive cells, ranging from predominant BAC histology with a mall focus of invasion, to invasive adenocarcinoma with an isolated group of cells with BAC features at the periphery. It occurs more frequently among non-smokers, women and Asians, so it is most likely to be unpreventable. The only potentially curative therapy for BAC is surgical resection. Localized BAC is treated like other NSCLC with lobar lung resection and ipsilateral mediastinal lymphadenectomy. Unresectable or metastatic BAC has historically been considered to be resistant to chemotherapy. Therefore, it is critically important to search for novel ways to kill BAC cells or increase its sensitivity to chemotherapy. Based on tissue microarray (TMA) analysis, 76% of 59 NSCLC and 6/7 BAC expressed Msi1, whereas only rare Msi1-positive cells were found in normal lung tissue. Therefore, this proposal will address the hypothesis that BAC stem cell survival is dependent on the stem cell regulator, Musashi1. These studies will focus on three specific aims:
Specific Aim #1: Determine the tumor stem cell population in BAC cell lines. Tumor cells will be grown as non-adherent spheroids in cell culture to determine the cancer stem cell phenotype by determining the percentage of cells that are CD133+ and/or CD44+/CD24lo by fluorescence-activated sort sorting (FACS).
Specific Aim#2: Determine the effect of inhibiting Msi1 expression on BAC stem cells by RNA interference using short hairpin RNA (shRNA). Msi1shRNA will be stably expressed in BAC cells that will then be characterized for tumor stem cell markers by FACS. Proliferation and apoptosis in BAC cell lines after stable expression of Msi1shRNA will be determined. Tumorigenicity of BAC stem cells stably expressing either control shRNA or Msi1shRNA will be determined by transplanting varying dilutions of cells into SCID mice.
Specific Aim #3: Determine the sensitivity of BAC stem cells stably expressing either control shRNA or Msi1shRNA to therapeutic drugs. Sensitivity to gefitinib and carboplatin will be examined initially since these are the primary agents used to treat BAC. Through these Specific Aims, we expect to determine the utility of targeting Msi1 in tumor stem cells as a potential target by which to sensitize BAC to inhibition of cell growth
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