The effect of oncogenic Ras signaling strength on cancer

描述(申请人提供)：在三分之一的人类癌症中，KRAS、较小程度的NRAS和罕见的HRAS发生突变，这是公认的促进肿瘤发生的机制。使用两种独立的方法在内源性水平上操纵致癌KRAS信号，我们表明这种信号的大小会导致相反的影响，这取决于肿瘤发生的阶段。具体地说，我们在第一份拨款申请中报告了野生型HRAS和NRAS蛋白在致癌KRAS下游被激活，当被击倒时，抑制了癌细胞系的肿瘤生长。相反，我们现在发现，在小鼠中，野生型Hras1的遗传消融促进了早期致癌的KRAS驱动的肿瘤发生。除了这方面的研究外，我们最近发现，由于稀有密码子的偏向，KRAS的表达比HRAS表达得很差。将稀有密码子改为常见密码子可产生更多致癌KRAS蛋白，促进已建立的细胞系的肿瘤生长，但抑制小鼠早期病变的发展。由于致癌RAS可以诱导正常细胞衰老生长停滞，我们假设KRAS在正常细胞中的表达范围非常窄-高到足以促进增殖，低到足以避免过度衰老-在突变时导致增殖。我们建议在AIM 1中通过确定在早期肿瘤形成过程中操纵野生型RAS蛋白的表达和KRAS的密码子偏向对衰老的影响来检验这一假说。然而，低致癌KRAS信号仍然抑制已建立的细胞系的肿瘤生长。因此，我们假设，在肿瘤发展的后期阶段，存在克服罕见密码子施加的KRAS表达不足的选择性压力。在这方面，在许多人类癌症中，突变的KRAS等位基因的拷贝数增加，这与更差的预后有关。因此，我们建议在AIM 2中测试在耐受高癌基因表达的环境中将罕见密码子改为常见密码子是否会抑制肿瘤发生过程中Kras拷贝数的增加。我们的初步发现表明，拷贝数的增加并不是增加KRAS蛋白表达的唯一机制。因此，我们还建议确定在肿瘤发生过程中KRAS蛋白的表达是如何升高的，以及密码子偏爱是否也调节其他癌症相关基因的表达。这项研究的完成将阐明内源性致癌KRAS信号水平如何不同地影响早期和晚期肿瘤发生的分子机制。

DESCRIPTION (provided by applicant): KRAS, to a lesser extent NRAS, and rarely HRAS, are mutated in a third of human cancers, which is well established to promote tumorigenesis. Using two independent approaches to manipulate oncogenic KRas signaling at the endogenous level, we show that the magnitude of this signaling causes opposite effects depending on the stage of tumorigenesis. Specifically, we reported in the first grant submission that wild-type HRas and NRas proteins are activated downstream of oncogenic KRas, and when knocked down, inhibited the tumor growth of cancer cell lines. Conversely, we now find that genetic ablation of wild-type Hras1 in mice promotes early oncogenic KRas-driven tumorigenesis. Independent of this line of investigation, we recently discovered that KRas is expressed very poorly compared to HRas owing to a bias of rare codons. Changing the rare to common codons produced more oncogenic KRas protein, enhanced tumor growth of established cell lines, but inhibited development of early lesions in mice. Since oncogenic Ras can induce a senescent growth arrest in normal cells, we hypothesize that KRas is expressed in a very narrow range in normal cells - high enough to promote proliferation but low enough to avoid excessive senescence - leading to hyperplasia when mutated. We propose to test this hypothesis in AIM 1 by determining the impact of manipulating the expression of wild-type Ras proteins and the codon bias of KRAS on senescence during early tumorigenesis. However, low oncogenic KRas signaling nevertheless inhibited tumor growth of established cell lines. As such, we hypothesize that there is selective pressure to overcome poor KRas expression imposed by rare codons during later stages of tumor development. In this regard, there is a gain in the copy number of the mutant KRAS allele in many human cancers, which is associated with a worse prognosis. We thus propose in AIM 2 to test whether changing the rare to common codons in settings that tolerate high oncogene expression suppresses a gain in Kras copy number during tumorigenesis. Our preliminary findings indicate that a gain in copy number is not the only mechanism to increase KRas protein expression. Thus, we also propose to identify how else KRas protein expression is elevated during tumorigenesis, and whether codon bias also regulates the expression of other cancer-related genes. Completion of this study will elucidate the molecular mechanisms underlying how the level of endogenous oncogenic KRas signaling differentially impacts early and late tumorigenesis.