Novel Human Cancer Cell Isolation System

描述（由申请人提供）：细胞系是许多癌症研究的关键试剂。 人们可能会凭直觉想象，癌细胞具有体内选择性生长的优势，在培养中相对容易建立。 然而，产生癌细胞系是令人惊讶地困难的。 无胸腺（“裸”）和严重联合免疫缺陷（SCID）小鼠是支持人类癌细胞生长的有价值的工具。 当收获癌症用于体外细胞系生产时，成纤维细胞和其他基质细胞通常在培养皿上过度生长，并且在大多数情况下阻止恶性细胞的分离。 对侵袭和转移（癌症的标志）的研究同样受到无法恢复已经完成该过程的少量癌细胞的阻碍。 为了避免这些问题，我们正在制造hprt缺陷的免疫缺陷小鼠，并且已经构建了裸hprt缺失小鼠。 与其他免疫缺陷小鼠类似，它们将支持许多异种移植癌症的生长。 在肿瘤生长过程中，小鼠细胞取代了人类基质细胞。 当我们希望恢复人类细胞时，我们将通过在经典的选择性培养基HAT中培养来消除小鼠基质细胞。 在该提案中，我们计划：生成SCID hprt-无效小鼠，证明生化缺陷小鼠用于细胞系生产的优越性，并证明自发转移细胞的恢复增强。 具体目标#1：产生生物化学选择的免疫缺陷小鼠，用于人类癌细胞恢复。 具体地，产生SCID hprt-无效小鼠。 具体目标#2：记录hprt缺失小鼠优于标准裸鼠和SCID小鼠。 具体而言，记录小鼠基质细胞替代，并从患者来源的原发性癌症（乳腺癌、前列腺癌、肺癌、胰腺癌和神经胶质癌）建立的异种移植物中分离纯的人癌细胞系群体。 比较有无基质细胞补充的恢复情况。 具体目标#3：记录回收少量转移性癌细胞的效用。 具体而言，确定加标样品的回收率，并将器官特异性转移灶与实验转移灶和原位植入后的自发转移灶分离。 小鼠将存放在杰克逊实验室，分发给包括NIH在内的非营利机构的研究人员。

DESCRIPTION (provided by applicant): Cell lines are critical reagents for much of cancer research. One might intuitively imagine that cancer cells, with their in vivo selective growth advantage, would be relatively easy to establish in culture. However, generating cancer cell lines is surprisingly difficult. Athymic ("nude") and severe combined immunodeficient (SCID) mice are valuable tools that support human cancer cell growth. When the cancer is harvested for in vitro cell line production, fibroblasts and other stromal cells commonly overgrow the culture dish and in most cases prevent isolation of the malignant cells. The study of invasion and metastasis (hallmarks of cancer) is similarly hindered by the inability to recover small numbers of cancer cells that have completed the process. To circumvent these problems, we are generating immunodeficient mice that are hprt defective, and have already constructed a nude hprt-null mouse. Similar to other immunodeficient mice, they will support growth of many xenografted cancers. During tumor growth, mouse cells replace the human stromal cells. When we wish to recover the human cells, we will eliminate the mouse stromal cells by growing the culture in the classic selective media, HAT. In this proposal, we plan to: generate SCID hprt-null mice, demonstrate superiority of the biochemically defective mice for cell line production, and demonstrate enhanced recovery of cells from spontaneous metastases. Specific Aim #1: Generate biochemically selectable immunodefective mice designed for human cancer cell recovery. Specifically, generate SCID hprt-null mice. Specific Aim #2: Document superiority of the hprt-null mice over standard nude and SCID mice. Specifically, document mouse stromal cell replacement and isolate pure populations of human cancer cell lines from xenografts established from patient-derived primary cancers (breast, prostate, lung, pancreatic and glial cancers). Compare recovery with and without stromal cell supplementation. Specific Aim #3: Document utility in recovering small numbers of metastatic cancer cells. Specifically, determine recovery efficiency from spiked samples, and isolate organ-specific metastases from experimental metastases and from spontaneous metastases after orthotopic implantation. Mice will be deposited at Jackson Labs for distribution to investigators at non-profit institutions, including the NIH.