Photolithographic Tumor DNA Isolation

翻译后摘要：光刻肿瘤DNA分离 个性化肿瘤学是基于癌症中的突变决定最佳治疗的想法。 随着新一代测序仪和生物信息学的发展，突变检测越来越成为可能， 目前，DNA分离的第一步是专门的、手工的和非标准化的。人类肿瘤很复杂 正常细胞和肿瘤细胞的混合物，突变检测将变得更加可靠和可重复， 提取了几乎纯的肿瘤DNA。在这里，我们建议开发一个自动化系统，可以提取 通过集成光刻技术，从常规H&E染色的显微镜载玻片中获得>90%纯度的肿瘤DNA， 高分辨率幻灯片扫描仪、图像分析算法和现代3D打印机。机器学习图像 算法可以区分肿瘤和正常细胞，这些信息将被传输到3D打印机， 其将不透明材料直接放置在载玻片上的肿瘤细胞核上。然后将载玻片暴露在短波中 UV光破坏未受保护的正常细胞中的DNA，而肿瘤DNA被选择性地保护。 戴上面具。可以从整个载玻片中提取DNA，只有被保护的肿瘤细胞中的DNA 可以测序（全外显子组或靶向测序）或测量CpG甲基化。当场 3D打印机的分辨率约为40微米，因此非常不规则复杂的地形和小 像单个肿瘤腺体这样的特征可以通过光刻来保护。整个系统（扫描，分析， 打印、照射、提取）可以在约24小时内从H&E载玻片产生>90%纯的肿瘤DNA。 转型的潜力是，该系统将目前的特设，高度劳动密集型 一个自动化的，有据可查的和可重复的提取技术步骤，可以添加信息， 因为准确提取的细胞数量、它们的表型和空间位置是已知的，所以可以学习。因为 它使用图像算法来选择肿瘤细胞，任何人都可以进行“相同”的DNA分离 世界上任何地方。而且，图像算法可以根据反馈“学习”以更好地提取肿瘤DNA DNA测序的结果将该系统集成到测序管道中将改善 通过提取接近纯（>90%）的肿瘤DNA， 这将推动可重复的癌症研究和精确肿瘤学的临床转化。

Abstract: Photolithographic Tumor DNA Isolation Personalized oncology is based on the idea that the mutations in a cancer determine optimal therapy. Mutation detection is increasingly possible with newer next generation sequencers and bioinformatics, but currently the first step of DNA isolation is ad hoc, manual, and non-standardized. Human tumors are complex mixtures of normal and tumor cells, and mutation detection would become more reliable and reproducible if nearly pure tumor DNA was extracted. Here we propose to develop an automated system that can extract >90% pure tumor DNA from conventional H&E stained microscope slides by integrating photolithography with high resolution slide scanners, image analysis algorithms, and modern 3D printers. Machine learning image algorithms can distinguish tumor from normal cells, and this information will be transferred to the 3D printer, which places opaque material directly over tumor nuclei on the slide. The slide is then exposed to short wave UV light to destroy the DNA in unprotected normal cells whereas tumor DNA is selectively protected by the photolithographic mask. DNA can be extracted from the entire slide, and only DNA in the protected tumor cells can be sequenced (whole exome or targeted sequencing) or measured for CpG methylation. The spot resolution of the 3D printer is about 40 microns, and therefore very irregular complex topography and small features like a single tumor gland can be protected by photolithography. The entire system (scan, analyze, print, irradiate, extract) could yield >90% pure tumor DNA from an H&E slide in about 24 hours. The transformational potential is that the system converts a currently ad hoc, highly labor-intensive technical step into an automated, well-documented and reproducible extraction that can add information and learning because the exact extracted cell numbers, their phenotypes and spatial locations are known. Because it uses an image algorithm to select the tumor cells, the “same” DNA isolation can be performed by anyone anywhere in the world. Moreover, image algorithms can “learn” to better extract tumor DNA based on feedback from the DNA sequencing. The integration of this system into a sequencing pipeline would improve the reliability, documentation, and reproducibility of mutation calling by extracting nearly pure (>90%) tumor DNA, which will advance both reproducible cancer research and the clinical translation of precision oncology.