Andreou Chrysafis, Oseledchyk Anton, Nicolson Fay, Berisha Naxhije, Pal Suchetan, Kircher Moritz F
Department of Radiology, Memorial Sloan Kettering Cancer Center.
Department of Radiology, Memorial Sloan Kettering Cancer Center; Department of Chemistry, The Graduate Center of the City University of New York.
J Vis Exp. 2019 Mar 25(145). doi: 10.3791/58389.
Ovarian cancer represents the deadliest gynecologic malignancy. Most patients present at an advanced stage (FIGO stage III or IV), when local metastatic spread has already occurred. However, ovarian cancer has a unique pattern of metastatic spread, in that tumor implants are initially contained within the peritoneal cavity. This feature could enable, in principle, the complete resection of tumor implants with curative intent. Many of these metastatic lesions are microscopic, making them hard to identify and treat. Neutralizing such micrometastases is believed to be a major goal towards eliminating tumor recurrence and achieving long-term survival. Raman imaging with surface enhanced resonance Raman scattering nanoprobes can be used to delineate microscopic tumors with high sensitivity, due to their bright and bioorthogonal spectral signatures. Here, we describe the synthesis of two 'flavors' of such nanoprobes: an antibody-functionalized one that targets the folate receptor - overexpressed in many ovarian cancers - and a non-targeted control nanoprobe, with distinct spectra. The nanoprobes are co-administered intraperitoneally to mouse models of metastatic human ovarian adenocarcinoma. All animal studies were approved by the Institutional Animal Care and Use Committee of Memorial Sloan Kettering Cancer Center. The peritoneal cavity of the animals is surgically exposed, washed, and scanned with a Raman microphotospectrometer. Subsequently, the Raman signatures of the two nanoprobes are decoupled using a Classical Least Squares fitting algorithm, and their respective scores divided to provide a ratiometric signal of folate-targeted over untargeted probes. In this way, microscopic metastases are visualized with high specificity. The main benefit of this approach is that the local application into the peritoneal cavity - which can be done conveniently during the surgical procedure - can tag tumors without subjecting the patient to systemic nanoparticle exposure. False positive signals stemming from non-specific binding of the nanoprobes onto visceral surfaces can be eliminated by following a ratiometric approach where targeted and non-targeted nanoprobes with distinct Raman signatures are applied as a mixture. The procedure is currently still limited by the lack of a commercial wide-field Raman imaging camera system, which once available will allow for the application of this technique in the operating theater.
卵巢癌是最致命的妇科恶性肿瘤。大多数患者就诊时已处于晚期（国际妇产科联盟（FIGO）分期为III期或IV期），此时局部转移已经发生。然而，卵巢癌具有独特的转移模式，即肿瘤种植最初局限于腹腔内。从理论上讲，这一特征能够实现以治愈为目的的肿瘤种植完全切除。这些转移病灶中有许多是微小的，难以识别和治疗。中和此类微转移被认为是消除肿瘤复发并实现长期生存的主要目标。由于具有明亮且生物正交的光谱特征，表面增强共振拉曼散射纳米探针的拉曼成像可用于高灵敏度地描绘微小肿瘤。在此，我们描述了两种此类纳米探针的合成：一种是抗体功能化的纳米探针，靶向许多卵巢癌中过度表达的叶酸受体；另一种是非靶向对照纳米探针，具有不同的光谱。将这些纳米探针经腹腔共同给药于转移性人类卵巢腺癌小鼠模型。所有动物研究均经纪念斯隆凯特琳癌症中心机构动物护理和使用委员会批准。通过手术暴露动物的腹腔，冲洗后用拉曼显微光谱仪进行扫描。随后，使用经典最小二乘法拟合算法解耦两种纳米探针的拉曼特征，并将它们各自的分数相除，以提供叶酸靶向探针与非靶向探针的比率信号。通过这种方式，微小转移灶得以高特异性地可视化。这种方法的主要优点是在手术过程中可方便地将其局部应用于腹腔，能够标记肿瘤，而不会使患者受到全身性纳米颗粒暴露。通过采用比率法，将具有不同拉曼特征的靶向和非靶向纳米探针混合应用，可以消除纳米探针在内脏表面非特异性结合产生的假阳性信号。目前，该方法仍受限于缺乏商业宽场拉曼成像相机系统，一旦有了该系统，将可在手术室应用这项技术。
