癌地致小鼠膀胱肿瘤的磁共振成像评价
Summary
小鼠膀胱肿瘤是由 n-丁基-n-(4-羟基丁基) 亚硝胺致癌物 (BBN) 诱导的。膀胱肿瘤的产生是异质性的;因此, 在随机化进行实验治疗之前, 需要对肿瘤负担进行准确的评估。在这里, 我们提出了一个快速, 可靠的 MRI 方案, 以评估肿瘤的大小和阶段。
Abstract
小鼠膀胱肿瘤模型是评估新的治疗方案的关键。n-丁基-n-(4-羟基丁基) 亚硝胺 (BBN) 致癌物质诱导的膀胱肿瘤优于基于细胞的模型, 因为它们密切复制人类肿瘤的基因组特征, 与细胞模型和异种移植不同, 它们提供了很好的机会, 可以研究免疫疗法。然而, 膀胱肿瘤的产生是异质性的;因此, 在随机化进行实验治疗之前, 需要对肿瘤负担进行准确的评估。本文介绍了 BBN 小鼠模型和方案, 以评估膀胱癌肿瘤负担在体内使用快速和可靠的磁共振 (mr) 序列 (真正的 fisp)。这种方法简单可靠, 因为与超声波不同, MR 是独立于操作器的, 并且允许简单的采集后图像处理和审查。利用膀胱的轴向图像, 分析沿膀胱壁和肿瘤感兴趣的区域, 可以计算膀胱壁和肿瘤区域。此测量与体外膀胱重量 (rs= 0.37, p = 0.009) 和肿瘤阶段 (p = 0.0003) 相关。总之, BBN 产生非均质性肿瘤, 是评估免疫治疗的理想选择, MRI 可以在随机化实验治疗臂之前快速可靠地评估肿瘤负担。
Introduction
膀胱癌是全球第五大最常见的癌症, 2017年在美国造成约 80 000个新病例和 16, 000 例死亡1。在经历了大约30年没有取得重大进展的系统治疗膀胱癌2之后, 最近的抗 pd-1 和抗 pd-1 检查点抑制剂试验在晚期患者中表现出令人兴奋的、偶尔持久的反应泌尿系上皮癌 3,4,5。然而, 只有约20% 的患者对这些治疗方法表现出客观的反应, 还需要进一步研究, 以扩大膀胱癌患者免疫治疗的有效使用。
小鼠膀胱癌模型是临床前评价新治疗方法 6、7 的关键工具。为了控制小鼠在随机进行不同治疗时的肿瘤大小, 必须在治疗组之间进行评估和控制。以前的研究使用超声波或生物发光来评估原位细胞线基础膀胱癌模型8,9,10,11。但是, 这两种技术都有几个缺点。超声测量可能会受到操作人员技能的影响, 缺乏三维特征和较高的空间分辨率。生物发光方法只能提供肿瘤细胞的半定量评价, 不允许膀胱解剖和形态的可视化。此外, 生物发光只能用于基于细胞系的模型, 这些模型在无毛小鼠或白外套小鼠中表达生物发光基因。
另一方面, 磁共振成像 (MRI) 在获取高分辨率解剖图像方面具有独特的灵活性, 具有广泛的组织对比度, 可实现对肿瘤负担的准确可视化和定量评估无需表示生物发光特性。MR 图像更易于重现与适当的分析管道和保证的膀胱三维可视化。MRI 最大的限制是检查所需的时间长度以及相关的高成本, 这些成本限制了高吞吐量检测。然而, 一些研究表明, MR 序列可以提供高质量的诊断图像, 可用于有效地检测和监测细胞线为基础的膀胱肿瘤;因此, 它们可用于高吞吐量分析9,12。
在这里, 我们描述了一种非侵入性的基于 mr 的方法, 以可靠和有效地表征致癌物诱导的小鼠膀胱肿瘤。为了实现这一目标, 我们使用了具有稳态进动 MR 技术 (true FISP) 的快速成像技术, 该技术保证了较短的扫描过程, 同时仍可提供高质量和高空间分辨率 (~ 100 微米), 用于检测和测量膀胱肿瘤13。此外, 为了确认这种非侵入性 MRI 检测的准确性, 我们描述了 mri 衍生参数与体外膀胱重量以及病理证实的肿瘤阶段之间的相关性。
Protocol
Representative Results
Discussion
准确的肿瘤模型成像是必要的适当的安乐死分期和动物随机化之前开始实验治疗。利用这里介绍的程序, 我们演示了方法, 以产生膀胱肿瘤使用 BBN 致癌物和 (2) 分层膀胱肿瘤负担, 通过使用 mr. a rr 衍生面积测量 (BLA壁)显着相关体外膀胱重量与病理肿瘤分期有关。
通过采用在高空间分辨率 (真正的 FISP) 和高诊断质量下获取时间短的快速成像方法, 我们可以在治疗?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
J. J. M. 由退伍军人卫生管理局资助。J. J. M. 还得到西北大学罗伯特·H·卢里综合癌症中心约翰·汉森癌症研究基金会的支持。我们感谢翻译成像中心提供的 MRI 采集和处理。资金来源在撰写手稿或决定提交出版方面没有任何作用。
Materials
| C57BL/6 mice | The Jackson Laboratory | 664 | Mice |
| N-butyl-N-(4-hydroxybutyl)nitrosamine carcinogen (BBN) | TCI American | B0938 | Carcinogen |
| 0.9% normal saline | Hospira, Inc | NDC 0409-488-02 | |
| Isoflurane | Piramal HealthCare | 60307-120-25 | Anesthetic |
| 7Tesla ClinScan MRI | Bruker | NA | Dedicated Small Animal Imaging MRI |
| Syngo | Siemens | NA | MR Integrated Imaging Software |
| Model 1030 Monitoring & Gating System | Small Animal Instruments, Inc. (SAII) | NA | Small animal physiologic monitoring |
| Formalin, Neutral Buffered, 10% | Sigma | HT501128 | Fixative |
| Eosin Y | Fisher Scientific | NC1093844 | Histologic staining agent |
| Hematoxylin | Fisher Scientific | 23-245651 | Histologic staining agent |
| Jim7 | Xinapse Systems | NA | Medical image analysis software |
| GraphPad Prism v7.04 | Graphpad | NA | Graphing software |
| R v3.4.2 | The R Project for Statistical Computing | NA | Statistical software |
| R package pROC v1.10.0. | The R Project for Statistical Computing | NA | ROC analysis |
References
- Siegel, R. L., Miller, K. D., Jemal, A. Cancer Statistics, 2017. CA: A Cancer Journal for Clinicians. 67 (1), 7-30 (2017).
- Abdollah, F., et al. Incidence, survival and mortality rates of stage-specific bladder cancer in United States: a trend analysis. Cancer Epidemiology. 37 (3), 219-225 (2013).
- Rosenberg, J. E., et al. Atezolizumab in patients with locally advanced and metastatic urothelial carcinoma who have progressed following treatment with platinum-based chemotherapy: a single-arm, multicentre, phase 2 trial. The Lancet. 387 (10031), 1909-1920 (2016).
- Sharma, P., et al. Nivolumab monotherapy in recurrent metastatic urothelial carcinoma (CheckMate 032): a multicentre, open-label, two-stage, multi-arm, phase 1/2 trial. The Lancet Oncology. 17 (11), 1590-1598 (2016).
- Bellmunt, J., et al. Pembrolizumab as Second-Line Therapy for Advanced Urothelial Carcinoma. New England Journal of Medicine. 376 (11), 1015-1026 (2017).
- Chan, E., Patel, A., Heston, W., Larchian, W. Mouse orthotopic models for bladder cancer research. BJU International. 104 (9), 1286-1291 (2009).
- Zhang, N., Li, D., Shao, J., Wang, X. Animal models for bladder cancer: The model establishment and evaluation (Review). Oncology Letters. 9 (4), 1515-1519 (2015).
- Patel, A. R., et al. Transabdominal micro-ultrasound imaging of bladder cancer in a mouse model: a validation study. Urology. 75 (4), 799-804 (2010).
- Chin, J., Kadhim, S., Garcia, B., Kim, Y. S., Karlik, S. Magnetic resonance imaging for detecting and treatment monitoring of orthotopic murine bladder tumor implants. The Journal of Urology. 145 (6), 1297-1301 (1991).
- Jurczok, A., Fornara, P., Soling, A. Bioluminescence imaging to monitor bladder cancer cell adhesion in vivo: a new approach to optimize a syngeneic, orthotopic, murine bladder cancer model. BJU International. 101 (1), 120-124 (2008).
- Vandeveer, A. J., et al. Systemic Immunotherapy of Non-Muscle Invasive Mouse Bladder Cancer with Avelumab, an Anti-PD-L1 Immune Checkpoint Inhibitor. Cancer Immunology Research. 4 (5), 452-462 (2016).
- Kikuchi, E., et al. Detection and quantitative analysis of early stage orthotopic murine bladder tumor using in vivo magnetic resonance imaging. Journal of Urology. 170, 1375-1378 (2003).
- Chung, H. W., et al. T2-weighted fast MR imaging with true FISP versus HASTE: comparative efficacy in the evaluation of normal fetal brain maturation. American Journal of Roentgenology. 175 (5), 1375-1380 (2000).
- Miyamoto, H., et al. Promotion of bladder cancer development and progression by androgen receptor signals. Journal of the National Cancer Institute. 99 (7), 558-568 (2007).
- Bertram, J. S., Craig, A. W. Specific induction of bladder cancer in mice by butyl-(4-hydroxybutyl)-nitrosamine and the effects of hormonal modifications on the sex difference in response. European Journal of Cancer. 8 (6), 587-594 (1972).
- Nagao, M., et al. Mutagenicity of N-butyl-N-(4-hydroxybutyl)nitrosamine, a bladder carcinogen, and related compounds. 암 연구학. 37, 399-407 (1977).
- Hirose, M., Fukushima, S., Hananouchi, M., Shirai, T., Ogiso, T. Different susceptibilities of the urinary bladder epithelium of animal species to three nitroso compounds. Gan. Gann; The Japanese Journal of Cancer Research. 67 (2), 175-189 (1976).
- Shin, K., et al. Cellular origin of bladder neoplasia and tissue dynamics of its progression to invasive carcinoma. Nature Cell Biology. 16 (5), 469-478 (2014).
- Epstein, J. I. Chapter 17: Immunohistology of the Bladder, Kidney, and Testis. Diagnostic Immunohistochemistry. , 624-661 (2019).
- Cohen, S. M., Ohnishi, T., Clark, N. M., He, J., Arnold, L. L. Investigations of rodent urinary bladder carcinogens: collection, processing, and evaluation of urine and bladders. Toxicologic Pathology. 35 (3), 337-347 (2007).
- Wood, D. P. Tumors of the bladder. Campbell-Walsh Urology. 11 (92), 2184-2204 (2016).
- Zitvogel, L., Pitt, J. M., Daillere, R., Smyth, M. J., Kroemer, G. Mouse models in oncoimmunology. Nature Reviews Cancer. , (2016).
- Kaneko, S., Li, X. X chromosome protects against bladder cancer in females via a KDM6A-dependent epigenetic mechanism. Science Advances. 4 (6), eaar5598 (2018).
- Smilowitz, H. M., et al. Biodistribution of gold nanoparticles in BBN-induced muscle-invasive bladder cancer in mice. International Journal of Nanomedicine. 12, 7937-7946 (2017).
- Dai, Y. C., et al. The interaction of arsenic and N-butyl-N-(4-hydroxybutyl)nitrosamine on urothelial carcinogenesis in mice. PLoS One. 12 (10), e0186214 (2017).
- Williams, P. D., Lee, J. K., Theodorescu, D. Molecular Credentialing of Rodent Bladder Carcinogenesis Models. Neoplasia. 10 (8), (2008).
- Fantini, D., et al. A Carcinogen-induced mouse model recapitulates the molecular alterations of human muscle invasive bladder cancer. Oncogene. 37 (14), 1911-1925 (2018).
- . NCCN Guidelines in Clinical Oncology – Bladder Cancer 5.2018 Available from: https://www.nccn.org/professionals/physician_gls/pdf/bladder.pdf (2018)
- Costa, M. J., Delingette, H., Novellas, S., Ayache, N. Automatic segmentation of bladder and prostate using coupled 3-D deformable models. Medical Image Computing and Computer-Assisted Intervention. 10 (Pt 1), 252-260 (2007).
- Rosenkrantz, A. B., et al. Utility of quantitative MRI metrics for assessment of stage and grade of urothelial carcinoma of the bladder: preliminary results. American Journal of Roentgenology. 201 (6), 1254-1259 (2013).
