REGULATION OF LUNG CANCER GROWTH

细胞周期进程的调节在肿瘤中被打乱 转化的细胞，通常是因为控制细胞周期的蛋白质 异常表达的异常表达或激活的这项建议的目的是 确定小细胞中控制细胞周期进程的蛋白质 肺癌(SCLC)是一种高度转移性的肿瘤，可导致 在美国，每年有25,000人死亡。一种独特的方法将是 用于识别这些蛋白质，基于我发现激活这些蛋白质 M3胆碱乙酰胆碱受体抑制细胞周期 小细胞肺癌细胞的进展。细胞周期调节蛋白将是 有两个标准来区分。首先，蛋白质必须被修饰(通过 转录、翻译或翻译后处理中的更改) 当小细胞肺癌细胞在细胞周期中进行时。第二，这些 当细胞周期进程被抑制时，不得进行修饰 MAChR激活。这一新的方法也可能确定致癌因素 小细胞肺癌细胞中的蛋白质。MAChR刺激可能独特地抑制小细胞肺癌 增殖，因为它使异常表达正常化或 激活有助于小细胞肺癌转化的蛋白质。为 例如，mAChR刺激可能诱导表达或激活 小细胞肺癌中低表达或失活的肿瘤抑制蛋白 细胞。这些受影响的蛋白质的特征可能会揭示 有助于SCLC转换的流程。 细胞周期依赖、转录后的蛋白质 将检测到由mAChR激活禁止的修改 比较了它们在小细胞肺癌细胞周期中独特的35S或32P标记 至静止或用氨基甲胆碱(MAChR)处理的细胞 激动剂)。这些蛋白质将通过抗体定向鉴定。 针对已知的调控蛋白和蛋白质微序列 分析。Northern印迹和核径流转录分析将 鉴定其细胞周期依赖积累的mRNA转录本 被mAChR激活改变。加快寻找监管机构 蛋白质，Southern印迹分析将用于鉴定蛋白质 无法参与mAChR介导的生长抑制，因为 基因缺失或基因重排。曾经的潜在调控蛋白 抗体和寡核苷酸探针将在 为了比较已鉴定的蛋白质的表达和活性 在小细胞肺癌和其他组织中。它们的独特表达或激活 与其他类型的细胞相比，小细胞肺癌细胞中的蛋白质将提供 令人信服的证据表明，已鉴定的蛋白质有助于 小细胞肺癌的肿瘤性转化。

The regulation of cell cycle progression is disrupted in neoplastically transformed cells, often because proteins controlling the cell cycle are abnormally expressed or activated. The objective of this proposal is to identify proteins which control cell cycle progression in small cell lung carcinoma (SCLC), a highly metastatic tumor which causes over 25,000 deaths per year in the United States. A unique approach will be used to identify these proteins, based on my finding that activation of M3 muscarinic acetylcholine receptors (mAChR) inhibits cell cycle progression in SCLC cells. Cell cycle regulatory proteins will be distinguished by two criteria. First, the proteins must be modified (by changes in transcription, translation, or posttranslational processing) when SCLC cells progress through the cell cycle. Second, these modifications must not occur when cell cycle progression is inhibited by mAChR activation. This novel approach may also identify oncogenic proteins in SCLC cells. mAChR stimulation may uniquely inhibit SCLC proliferation because it normalizes the aberrant expression or activation of proteins contributing to SCLC transformation. For example, mAChR stimulation may induce the expression or activation of tumor suppressor proteins that are underexpressed or inactivated in SCLC cells. Characterization of these affected proteins may shed light on processes contributing to SCLC transformation. Proteins undergoing cell cycle-dependent, post-transcriptional modifications that are inhibited by mAChR activation will be detected by their unique 35S- or 32P-radiolabelling in cycling SCLC cells, compared to cells that are quiescent or treated with carbachol (an mAChR agonist). These proteins will be identified using antibodies directed against known regulatory proteins and by protein micro-sequence analysis. Northern blot and nuclear run-off transcription assays will identify mRNA transcripts whose cell cycle-dependent accumulation is altered by mAChR activation. To expedite the search for regulatory proteins, southern blot analysis will be used to identify proteins unable to participate in mAChR-mediated growth inhibition because of genetic deletion or rearrangement. Once potential regulatory proteins are identified, antibody and oligonucleotide probes will be made in order to compare the expression and activity of the identified proteins in SCLC and other tissues. Unique expression or activation of these proteins in SCLC cells, compared to other cell types, will provide compelling evidence that the identified proteins contribute to neoplastic transformation in SCLC.