Time Domian Electron Paramagnetic Resonance Imaging

PO2成像的动态范围扩展：在前面的研究中，我们检查并确认了自旋探针Ox071(0x063的代版本)在基于R1和R2*的EPR血氧仪中的应用。由于线宽较窄，信号衰减较慢，预计Ox071不仅适用于缺氧组织，也适用于实体器官等缺氧程度较低的组织。在这项工作中，我们提出了我们的第一个3D在体EPR血氧测定研究，使用Ox071和Ox063血氧测定进行比较。用标准体模溶液分别在1、2、5、10 mm和0、2、5、10、21%处校准R2*随[Ox071]和pO2的变化。在体内，使用Ox071或Ox063对携带MIA Paca-2肿瘤的小鼠进行EPR成像。在0x063和0x071之间，与MRI共同配准标记的肿瘤区域的自旋密度、pO2图和pO2直方图相似。使用Ox071和Ox063的裸鼠也进行了健康的肾脏成像。与Ox063相比，Ox071血氧测定仪在肾脏中显示出更均匀的氧分压分布，提示Ox071适合在较高的氧分压范围内进行组织血氧测定仪。影响依福福胺疗效的肿瘤微环境决定因素：胰腺导管腺癌(PDAC)形成低血管和低氧肿瘤，目前的化疗方案难以治疗这些肿瘤。吉西他滨(GEM)经常被用作治疗PDAC的一线药物，但存在化疗耐药和肿瘤内部渗透的问题。埃夫福胺是一种低氧激活的前体药物，已被证明与GEM联合使用有效，尽管两种药物相互作用的机制尚未确定。我们使用了两种不同肿瘤微环境特征的细胞系(MIA Paca-2和SU 86.86)的两种小鼠移植瘤来探讨每种药物对另一种药物的作用。GEM治疗可延长SU 86.86移植瘤小鼠的存活时间(HR=0.35，95%CI=0.13~0.90 P=0.03)，但对MIA PACA-2小鼠无明显影响(HR=0.91，95%CI=0.37~2.25，P=0.84)。相反，依福福胺对SU86.86小鼠没有影响，也没有显著延长小鼠的存活时间(HR=0.57，95%CI=0.23~1.42，p=0.22)。在MIA Paca-2肿瘤中，电子顺磁共振(EPR)成像显示，当GEM治疗时，氧合作用恶化，这为GEM激活依福福胺以及依福福胺和GEM联合治疗的有效性提供了直接机制。在体外，任何一种处理的亚致死量都增强了其他处理的毒性，但在MIAPaca-2中没有。联合治疗可促进Su86.86双链DNA损伤的修复，但不能促进MIA PACA-2的修复。提出了依福福胺与GEM协同作用的可能机制。GEM和依福福胺之间的协同作用似乎源于GEM对肿瘤血管的影响和GEM对同源重组DNA修复过程的抑制的双重作用。每条途径的相对重要性取决于肿瘤微环境，值得进一步研究。低氧激活的前体药物依福福胺改善肿瘤的氧合能力。肿瘤有低位的区域。氧气水平被称为缺氧区，这些区域对放射治疗和化疗具有抵抗力。低氧激活的前体药物，如依福福胺，是为了特异性地杀死缺氧区域的细胞而开发的。在缺氧的肿瘤微环境中，强烈还原的氧化还原状态将依福福胺(TH-302)转化为还原形式，并释放出细胞毒性的溴异磷酰胺(BR-IPM)部分。因此，这种药物优先攻击肿瘤中的缺氧区，而其他标准的抗癌治疗，如化疗和放射治疗，往往无效。已经提出了各种与依福福胺的联合治疗方案，并在临床前和临床环境中进行了测试。然而，单用依福福胺对肿瘤缺氧的治疗效果尚不完全清楚，部分原因是缺乏定量方法来评估体内肿瘤的PO2。在这里，我们使用EPR的pO2定量成像，使用两个胰腺导管腺癌异种移植模型来评估肿瘤缺氧的变化；MIA Paca-2肿瘤对Eofosamide有反应，Su.86.86肿瘤没有反应。EPR成像显示，在低氧MIA Paca-2肿瘤治疗后，氧合功能总体上得到改善，这与体外低氧染色结果一致，与DCE MRI观察到的Ktras降低形成明显对比。观察到依福福胺不仅杀死了肿瘤的缺氧区，而且还改善了残留肿瘤区域的氧合作用，这为在依福福胺后使用放射和抗增殖药物以改善结果提供了理论基础。

Dynamic range extension of pO2 imaging: In the preceding research, we examined and confirmed the utility of spin probe Ox071, deuterated version of Ox063, for both R1 and R2* based EPR oximetry. Due to the narrow line width and slower signal decay, it was expected that pO2 estimation using Ox071 is suitable not only in hypoxic tissue but also in less hypoxic tissue such as solid organs. In this work, we present our first 3D in vivo EPR oximetry study using Ox071 in comparison with Ox063 oximetry. The R2* change with [Ox071] and pO2 was calibrated using standard phantom solutions at 1,2,5,10 mM and 0, 2, 5, 10, 21%, respectively. In vivo EPR imaging of a mouse bearing MIA Paca-2 tumor was performed on successive days by using either Ox071 or Ox063. The spin density, pO2 maps, and pO2 histograms in the tumor regions marked by co-registration with MRI were similar between Ox063 and Ox071. Healthy kidney imaging was also performed on athymic mice using both Ox071 and Ox063. Ox071 oximetry showed more homogeneous pO2 profile in kidney compared with Ox063, suggesting that Ox071 is suitable for oximetry in tissue at higher pO2 range. Tumor microenvironment determinants in evofosfamide efficacy: Pancreatic ductal adenocarcinomas (PDACs) form hypovascular and hypoxic tumors which are difficult to treat with current chemotherapy regimens. Gemcitabine (GEM) is often used as a first line treatment for PDACs, but has issues with chemoresistance and penetration in the interior of the tumor. Evofosfamide, a hypoxia activated prodrug, has been shown to be effective in combination with GEM, although the mechanism of each drug on the other has not been established. We used two mouse xenografts from two cell lines (MIA Paca-2 and SU 86.86) with different tumor microenvironmetal characteristics to probe the action of each drug on the other. GEM treatment enhanced survival times in mice with SU.86.86 xenografts (HR =0.35, 95% CI=0.13 to 0.90 p=0.03) but had no effect on MIA Paca-2 mice (HR =0.91, 95% CI=0.37 to 2.25, p=0.84). Conversely, evofosfamide had no effect on SU86.86 mice and did not improve survival times to a statistically significant degree (HR=0.57, 95% CI=0.23 to 1.42, p=0.22). In MIA Paca-2 tumors, which were initially poorly perfused, electron paramagnetic resonance (EPR) imaging showed that oxygenation worsened when treated with GEM, providing a direct mechanism for the activation of evofosfamide by GEM and the effectiveness of evofosfamide and GEM combinations. Sublethal amounts of either treatment enhanced the toxicity of other treatment in vitro in Su86.86 but not in MIAPaca-2. Repair of double stranded DNA lesions was enhanced in the combination treatment in Su86.86 but not MIA Paca-2. A possible mechanism for the synergy between evofosfamide and GEM has been proposed. The synergy between GEM and evofosfamide appears to stem from the dual action of GEM's effect on tumor vasculature and the GEM inhibition of the homologous recombination DNA repair process. The relative importance of each pathway is dependent on the tumor microenvironment and merits further study. The hypoxia activated prodrug Evofosfamide improves tumor oxygenation. Tumors have regions with low. Levels of oxygen called hypoxic zones, These regions are resistant to radiation therapy and chemotherapy. Hypoxia activated prodrugs such as Evofosfamide are developed to specifically kill cells in hypoxic regions. In hypoxic tumor microenvironments, the strongly reducing redox state converts evofosfamide (TH-302) to a reduced form and releases a cytotoxic bromo-isophosphoramide (Br-IPM) moiety. This drug therefore preferentially attacks hypoxic regions in tumors where other standard anti-cancer treatments such as chemotherapy and radiation therapy are often ineffective. Various combination therapies with evofosfamide have been proposed and tested in preclinical and clinical settings. However, the treatment effect of evofosfamide monotherapy on tumor hypoxia has not been fully understood, partly due to the lack of quantitative methods to assess tumor pO2 in vivo. Here, we use quantitative pO2 imaging by EPR to evaluate the change in tumor hypoxia in response to evofosfamide treatment using two pancreatic ductal adenocarcinoma xenograft models; MIA Paca-2 tumors responding to evofosfamide and Su.86.86 tumors which do not respond. EPR imaging showed oxygenation improved globally after evofosfamide treatment in hypoxic MIA Paca-2 tumors, in agreement with the ex vivo results obtained from hypoxia staining by pimonidazole and in apparent contrast to the decrease in Ktrans observed in DCE MRI. The observation that evofosfamide not only kills the hypoxic region of the tumor but also improves oxygenation in the residual tumor regions provides a rationale for combination therapies using radiation and anti-proliferatives post evofosfamide for improved outcomes.