A stem cell strategy to induce tumor stasis and inhibit metastasis of breast canc

描述(申请人提供)：乳腺癌细胞的恶性表型可以通过胚胎环境逆转为正常的细胞表型，这突显了一种可能的新的细胞转化策略，用于治疗恶性乳腺癌而不是杀死癌细胞。诱导多能干细胞(IPSC)技术可以将恶性癌细胞重新编程为类似胚胎干细胞(ESC)的细胞，提供活的胚胎环境。然而，目前IPSC技术的低转化率限制了其在癌症研究和治疗中的广泛应用。我们开发了一种蛋白质诱导的PSC(PIPSC)技术，该技术利用Sox2/Oct4/Nanog(SON)蛋白将异质恶性乳腺癌细胞(MalBCC)重新编程为PiPSC，转化率为90±4%。将4T1-piPSCs植入小鼠乳腺，可导致肿瘤停滞、转移抑制和向正常乳腺细胞分化。4T1细胞与4T1-piPSCs共培养提示了一种旁观者效应，使少量4T1-piPSCs能够逆转周围大量4T1细胞的恶性表型。在4T1荷瘤小鼠体内直接注射“SON”蛋白可导致肿瘤停滞和转移抑制，提示一种原位细胞转化癌症治疗方法。尾静脉注射QQ-SON蛋白将这些蛋白靶向乳腺癌肿瘤细胞的细胞核。我们的中心假说是：“注射”SON“蛋白，在原位将MalBCC转化为piPSCs，通过piPSCs的旁观者效应，将周围MalBCC的恶性表型逆转为正常细胞。”我们将首先优化乳腺上皮细胞和MalBCC的PIPSC生成。将进行MalBCC与piPSCs的共培养，以研究共培养的MalBCC在体外能否降低致瘤性(目标1)。为了研究体内的旁观者效应，我们将使用不同的 在不同时间点将(少量)piPSCs注入荷瘤小鼠的肿瘤内(大量)，以研究这些piPSCs是否能在体内诱导肿瘤停滞和抑制转移(目标2)。为了模拟乳腺癌治疗的临床环境，我们将直接注射 通过尾静脉或肿瘤内的“SON”蛋白在不同的时间点进入乳腺癌荷瘤小鼠，研究原位细胞转化是否可以诱导肿瘤停滞和抑制转移，从而延缓乳腺癌的复发和延长生存期(目标3)。与目前基于分子的细胞杀伤疗法不同，我们将产生组织特异性的 原位pPSCs作为活体胚胎环境，将乳腺癌细胞恶性表型逆转为正常细胞。这将通过尾静脉和肿瘤内注射SON蛋白到乳腺癌转移小鼠模型中来实现。虽然这项建议侧重于原则验证，但蛋白质诱导的原位细胞转化策略将很容易转化为人类临床应用。这项提议的成功可能会为这种拟议的细胞转化乳腺癌疗法的未来人类临床试验奠定基础，因为它可以显著推迟癌症复发，延长生存时间，并改善乳腺癌患者的预后。

DESCRIPTION (provided by applicant): Malignant phenotypes of breast cancer cells can be reversed into normal cell phenotypes by embryonic environments, highlighting a possible novel cell-converting strategy to treat malignant breast cancers instead of killing cancerous cells. The induced Pluripotent Stem Cell (iPSC) technique can reprogram malignant cancer cells into Embryonic Stem Cell (ESC)-like cells, providing living embryonic environments. However, the low conversion of the current iPSC technique limits its broad applications in cancer research and treatment. We developed a protein-induced PSC (piPSC) technique that reprogrammed heterogeneous malignant Breast Cancer Cells (malBCCs) into piPSCs using the Sox2/Oct4/Nanog (SON) proteins with 90±4% conversion. Implantations of the 4T1-piPSCs into mouse mammary glands resulted in tumor stasis, metastasis inhibition and differentiation into normal mammary cells. Co-cultures of 4T1 cells with 4T1-piPSCs suggest a bystander effect that enables a small number of 4T1-piPSCs to reverse malignant phenotype of large numbers of surrounding 4T1 cells. Direct intra-tumor injections of the "SON" proteins in the 4T1 tumor bearing mice resulted in tumor stasis and metastasis inhibition, suggesting an in situ cell-converting cancer therapy. Tail vein injections of QQ-SON proteins targeted these proteins into the nuclei of tumor cells of breast cancer. Our central hypothesis is: "Injections of the "SON" proteins convert malBCCs into piPSCs in situ that reverse malignant phenotype of the surrounding malBCCs into normal cells via a bystander effect of piPSCs". We will first optimize piPSC generation from mammary epithelial cells and malBCCs. Co-cultures of malBCCs with piPSCs will be performed to study if oncogenic properties of the co-cultured malBCCs can be reduced in vitro (Aim 1). To investigate the bystander effect in vivo, we will administer different doses of (small numbers) piPSCs into the tumor (large numbers) of the tumor-bearing mice at different time points to study if these piPSCs can induce tumor stasis and inhibit metastasis in vivo (Aim 2). To mimic the clinical setting of breast cancer treatment, we will directly inject the "SON" proteins either via tail vein or via intra-tumor into the breast cancer-bearing mice at different time points to investigate if the in situ cell conversion can induce tumor stasis and inhibit metastasis, therefore delaying breast cancer recurrence and prolonging survival (Aim 3). Different from the current molecular-based cell-killing therapies, we will generate tissue-specific piPSCs in situ as living embryonic environments to reverse malignant phenotype of breast cancer cells into normal cells. This will be achieved by injections of the SON proteins via tail vein and via intra-tumor into breast cancer metastatic mouse models. Although this proposal focuses on a proof-of-principle verification, a protein-induced in situ cell-conversion strategy wil be easily translatable to human clinical applications. The success of this proposal may set a stage for future human clinical trials of this proposed cell converting breast cancer therapy by significantly delaying cancer recurrence, prolonging survival and improving outcomes of breast cancer patients.