Cancer Resistant Mice

项目摘要 生殖系突变能对其他致命癌症产生强大的抵抗力吗？某些种系基因型可能 对肿瘤血管形成、营养需求或对免疫攻击的抵抗力的支持性较差， 与宿主的生存相适应。特别令人感兴趣的是，一些突变可能会助长宿主对neo的反应， 抗原，或甚至同源肿瘤中高度表达的自身抗原。抗性鉴定 突变可以为癌症治疗提供新的方法和靶点。至少在人类中， 抗药性突变就很难识别。人类生殖系遗传变异性， 癌症基因组，和高频率的人谁从来没有患癌症在他们的生活将 使得绘制新的人类耐药等位基因几乎不可能。在小鼠中，发现这样的突变要容易得多。 同基因肿瘤系（具有相对稳定的基因组）存在于许多近交系的小家鼠中。近交 小鼠本身具有确定的生殖系参考序列。每个个体在几乎所有的基因座上都是纯合的， 几乎与其他物种的基因完全相同过去几年，我们利用这种情况， 鉴定突变赋予癌症抗性的基因。使用随机生殖系诱变剂ENU，我们 建立了第三代（G3）种系突变小鼠（C57 BL/6 J品系）。第三代（G3）共23,751辆 来自561个谱系的小鼠，总共携带32，039个非同义编码/剪接变化，被纳入 - 筛选，其中每只小鼠皮下注射2 e5 B16 F10黑素瘤细胞，和抗PD-1 在第5、8和11天施用抗体。在第13天和第20天测量肿瘤体积。G1男性 对每个谱系的创始者进行测序，以鉴定所有非同义编码/剪接突变， 诱变，并且在筛选之前在所有诱导的突变位点对所有G3后代进行基因分型。 自动减数分裂作图允许快速检测甚至微妙的表型和分配到致病 突变。这种筛选产生了几种突变，导致对可移植癌症的抗性。14.2%饱和度 在筛选中实现了常染色体基因组的（具有严重损伤或缺失的常染色体基因的分数）。 在纯合状态下测试三次或更多次的破坏性等位基因）。因此，还有很多东西未被发现。 从我们已经知道的情况来看，有一个现实的机会将基因发现从这个屏幕上转化为 人类癌症治疗该提案旨在扩大癌症耐药性的筛查，并进一步推动 两种耐药突变的机制和翻译研究，每一种都在具有人类直向同源物的基因中， 测试围绕每种蛋白质靶点建立的治疗方法之间的协同作用，并为 临床应用。

PROJECT SUMMARY Can germline mutations cause strong resistance to otherwise lethal cancers? Certain germline genotypes might be poorly supportive of tumor vascularization, nutritional demands, or resistance to immune attack, yet compatible with host survival. Of particular interest, some mutations might abet the host response to neo- antigens, or even to self-antigens highly expressed in syngeneic tumors. The identification of resistance mutations could provide new approaches and targets for cancer therapy. At least in human populations, resistance mutations would be very difficult to identify. Human germline genetic variability, stem variability among cancer genomes, and the high frequency of humans who never develop cancer throughout their lives would make mapping novel human resistance alleles all but impossible. In mice, finding such mutations is much easier. Syngeneic tumor lines (with relatively stable genomes) exist for many inbred strains of Mus musculus. The inbred mice themselves have a defined germline reference sequence. Each individual is homozygous at nearly all loci, and almost genetically identical to all others. Over the past several years, we took advantage of this situation to identify genes in which mutations confer cancer resistance. Using the random germline mutagen ENU, we created third generation (G3) germline mutant mice (C57BL/6J strain). A total of 23,751 third-generation (G3) mice from 561 pedigrees, bearing a total of 32,039 non-synonymous coding/splicing changes were enrolled into a screen in which each mouse was injected subcutaneously with 2e5 B16F10 melanoma cells, and anti-PD-1 antibody was administered on days 5, 8, and 11. Tumor volume was measured on days 13 and 20. The G1 male founder of each pedigree was sequenced to identify all non-synonymous coding/splicing mutations induced by mutagenesis, and all G3 descendants were genotyped at all induced mutation sites in advance of screening. Automated meiotic mapping allowed quick detection of even subtle phenotypes and assignment to causative mutations. This screen yielded several mutations causing resistance to transplantable cancers. 14.2% saturation of the autosomal genome was achieved in screening (fraction of autosomal genes with severely damaging or destructive alleles tested in the homozygous state three times or more). Therefore, much remains undiscovered. From what we know already, there is a realistic chance of translating genetic discoveries from this screen to human cancer therapy. This proposal aims to extend screening for cancer resistance, and to further advance mechanistic and translational studies of two resistance mutations, each in a gene with a human orthologue, testing synergy between therapeutic approaches built around each protein target, and laying groundwork for clinical applications.