A novel approach to Mesenchymal Stem Cell Transduction

间充质干细胞转导的新方法

• 批准号: 10325618
• 负责人: KENNETH CORNETTA
• 金额: $ 24.85万
• 依托单位: OSSIUM HEALTH, INC.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-20 至 2024-03-19
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10325618
• 关键词: Address; Adipose tissue; Animal Model; Biological Assay; CD34 gene; Canis familiaris; Cardiovascular system; Cartilage; Cell Survival; Cell Therapy; Cells; Child; Chronic Granulomatous Disease; Clinic; Clinical; Clinical Trials; Clonal Expansion; Clone Cells; Data; Development; Disease; Ensure; Face; Flow Cytometry; Frequencies; GTP-Binding Protein alpha Subunits, Gs; Gammaretrovirus; Gene Transfer; Gene-Modified; Genes; Genetic; Green Fluorescent Proteins; HIV-1; Hematopoietic; Hematopoietic stem cells; High-Throughput Nucleotide Sequencing; Human; Immunologics; Insertional Mutagenesis; Lentivirus Vector; Leukemic Cell; Link; Malignant Neoplasms; Mature T-Lymphocyte; Mesenchymal Stem Cells; Modification; Mus; Neurologic; Orthopedics; Patients; Pattern; Phase; Phenotype; Play; Primary carcinoma of the liver cells; Property; Publishing; Reporting; Research Personnel; Risk; Risk Estimate; Role; Safety; Site; Source; Surface; T-Lymphocyte; Thalassemia; Therapeutic; Transplantation; Vertebral Bone; Vesicular stomatitis Indiana virus; Viral; Wiskott-Aldrich Syndrome; Work; base; bone; bone marrow mesenchymal stem cell; cell bank; cell immortalization; cell type; cellular transduction; chimeric antigen receptor; clinical development; clinical implementation; cost; detection assay; efficacy study; gene therapy clinical trial; gene transfer vector; genetically modified cells; genotoxicity; hematopoietic stem cell expansion; improved; innovative technologies; integration site; lentiviral integration; lentivirally transduced; leukemia; novel; novel strategies; particle; pre-clinical; preference; premature; safety study; senescence; stem cell genes; stem cell model; stem cells; therapeutic effectiveness; vector; vertebra body

PROJECT SUMMARY Mesenchymal stem cells (MSC) are in clinical development for cardiovascular, neurologic, orthopedic, and other indications. Ossium is developing a novel source of MSC from vertebral bodies (vbMSC). Genetic modification of vbMSC using lentiviral vector (LV) has the potential to improve the therapeutic potential. Understanding how genetic modification might alter the vbMSC phenotype will be critical to ensuring LV do not initiate premature senescence or diminish their therapeutic properties. Moreover, an important safety and regulatory barrier to clinical implementation of gene modified vbMSC will be estimating the risk of insertional mutagenesis. Gammaretroviral vectors used to modify hematopoietic stem cells (HSC) led to leukemia in at least 4 clinical trials. While lentiviral vectors appear safer, clonal expansion of HSC and mature T cells have now been reported. Ossium seeks to address important safety and efficacy issues in this Phase I proposal. We hypothesize that (1) A HIV-1 based LV pseudotyped with a novel foamy viral envelope (LV-FV) will efficiently transduce vbMSC with less change in cell phenotype compared to LV pseudotyped with VSV-G; (2), the LV integration pattern, distribution among cancer associated genes and ability to induce cell immortalization, will be similar to that seen in other cell types. To study this, we propose: Specific Aim 1: Assess the Effect of LV Gene Transfer and Vector Pseudotype on vbMSC phenotype. LV expressing green fluorescent protein (GFP) pseudotyped with VSVG and FV will be used to transduce vbMSC. Cells will be assessed for gene transfer (vector copy number) and expression (GFP by flow cytometry), viability, expansion capacity, MSC-associated surface markers, secretome, and ability to differentiate into three lineages (bone, adipose and cartilage). Specific Aim 2: Evaluate Insertional Mutagenesis Risk in LV transduced vbMSC. This aim seeks to investigate the genotoxicity of LV vectors in MSC. Specific Aim 2A. Evaluating LV-transduced vbMSC for Integration Pattern and Cancer-Associated Gene Preferences. Comparisons between LV and gammaretroviral vectors in low and high passage vbMSC will be compared to published data on HSC integrations. Specific Aim 2B. Evaluating LV-transduced vbMSC for Immortalization. In this sub-aim we will seek to develop an assay for assessing IM risk by evaluating vbMSC after gene transfer. The findings will have broad applicability given the number of disease states in which MSC may play a therapeutic role. This proposal also uses a variety of innovative technologies, including a novel LV-FV, a novel stem cell source (vertebral body MSC), and will be the first study aimed at assessing the risk of immortalization in MSC.

项目摘要 间充质干细胞（MSC）在心血管，神经系统，骨科和 其他迹象。 Ossium正在开发椎体（VBMSC）的新型MSC来源。遗传 使用慢病毒载体（LV）对VBMSC进行修饰具有提高治疗潜力的潜力。 了解遗传修饰如何改变VBMSC表型对于确保LV DO至关重要 不要启动过早衰老或降低其治疗特性。而且，重要的安全性和 基因修饰的VBMSC临床实施的监管障碍将估计插入的风险 诱变。用于修饰造血干细胞（HSC）的γ逆转录病毒载体导致白血病AT 至少4次临床试验。尽管慢病毒载体显得更安全，但HSC和成熟T细胞的克隆膨胀具有 现在报道了。 Ossium试图在本一阶段提案中解决重要的安全和效力问题。 我们假设（1）用新型泡沫病毒包膜（LV-FV）将基于HIV-1的LV假型将 与用VSV-G进行的LV伪型相比，有效地转导VBMSC，细胞表型的变化较小； （2），LV整合模式，癌症相关基因之间的分布和诱导细胞的能力 永生化，将与其他细胞类型相似。为了研究这一点，我们提出：特定目标1： 评估LV基因转移和载体假型对VBMSC表型的影响。 LV表达绿色 用VSVG和FV伪型荧光蛋白（GFP）将用于传递VBMSC。细胞将是 评估基因转移（矢量拷贝数）和表达（通过流式细胞仪的GFP），生存力， 扩展能力，MSC相关的表面标记，分泌组和分为三个的能力 谱系（骨，脂肪和软骨）。特定目标2：评估LV中的插入诱变风险 转导的VBMSC。该目标旨在研究MSC中LV载体的遗传毒性。特定目标2a。 评估LV转导的VBMSC的整合模式和与癌症相关的基因偏好。 将比较低通和高通道VBMSC中LV和Gammaretroviral载体之间的比较 发表有关HSC集成的数据。特定目标2B。评估LV转导的VBMSC的 永生化。在此子AIM中，我们将寻求通过评估VBMSC来开发评估IM风险的测定法 基因转移后。鉴于疾病状态的数量，这些发现将具有广泛的适用性 MSC可能会发挥治疗作用。该建议还使用了各种创新技术，包括 新型LV-FV，一种新型的干细胞来源（椎体MSC），将是第一个旨在评估的研究 在MSC中永生化的风险。

项目成果

Gene therapy access: Global challenges, opportunities, and views from Brazil, South Africa, and India.

• DOI: 10.1016/j.ymthe.2022.04.002
• 发表时间: 2022-06-01
• 期刊: MOLECULAR THERAPY
• 影响因子: 12.4
• 作者: Cornetta, Kenneth;Bonamino, Martin;Mahlangu, Johnny;Mingozzi, Federico;Rangarajan, Savita;Rao, Jayandharan
• 通讯作者: Rao, Jayandharan