Multi-modality optical molecular imaging for melanoma tumor margin assessment

描述（由申请人提供）：与任何其他类型的癌症相比，黑色素瘤的发病率和死亡率增长最快（5，6）。目前对所有阶段的黑色素瘤的治疗包括手术，通过目视检查和术后组织病理学评估原发性病变的边缘。然而，对于消除局部复发和/或转移所需的手术切缘宽度仍存在争议（3，12，13），这些情况对患者有严重后果。此外，通过组织病理学实现的无肿瘤边缘的术后评估具有显著的局限性，包括1。是 主观的，2.仅在切除的一小部分上进行边缘的监测，以及3.手术与其成功反馈之间的时间（即，负保证金的确认）可能需要几天时间。光学高分辨率，三维，分子成像提供了一种手段，通过它可以定量，非侵入性，并在真实的时间评估的边缘。为此，该提案侧重于基于定量分子和结构替代生物标志物的多模态光学方法来评估黑色素瘤肿瘤边缘。将使用两种分子成像方法。第一种是泵浦-探测显微镜，这是一种非线性光学方法，可以实现色素（包括真黑素和褐黑素）的定量分子对比度（12）。这种方法特别适合诊断黑色素瘤，因为eu-和pheo-黑色素在这种疾病的恶性进展中起重要作用（12）。第二种方法，泵浦-探测非线性相位色散光谱（PP-NLDS），通过探测由交叉相位调制引起的相位变化产生补充信息，交叉相位调制是提供结构信息的普遍存在的效应（35，36），以及分子重取向效应（即，光学克尔效应），其提供对水性环境的扩散性质的灵敏度（20，32）。额外的结构信息将从多光子自发荧光、二次谐波产生和光学相干显微镜（OCM）中获得。该提案的第一个目标是建立一个综合的多模式系统。该策略是修改现有的定制显微镜，以纳入PP-NLDS和OCM，两者都用相同的仪器检测。目标2将侧重于使用人类皮肤移植小鼠模型制定适当的边缘评估标准。我们预期，边缘阳性的病变将包含多种标志物的组合，包括向正常色素的转变、真皮微卫星、组织结构紊乱、细胞内环境的差异和细胞核增大。目标3将使用来自尸体和接受手术的患者的人体标本来验证该方法。所有成像结果将与组织病理学相关联，以验证该方法。这项工作的成功完成将为提供定量生物标志物以评估黑色素瘤肿瘤边缘的仪器铺平道路，从而在不改变公认的临床范例的情况下，及时提高首次切除手术成功的机会。

DESCRIPTION (provided by applicant): Melanoma has the fastest growing incidence and mortality rates compared to any other type of cancer (5, 6). Current treatment for all stages of melanoma includes surgery with margins of the primary lesions assessed via visual inspection and post-operative histopathology. However, controversy still exists on the width of the surgical margins needed to eliminate local reoccurrence and/or in-transit metastases (3, 12, 13), conditions that have severe consequences for patients. In addition, post-operative assessment of tumor-free margins, achieved via histopathology, has significant limitations, including 1. It is subjective, 2. Surveillance of the margins is performed on just a small portion of the excision, and 3. The time between surgery and feedback on its success (i.e., confirmation of negative margins) can take days. Optical high-resolution, 3D, molecular imaging provides a means by which margins can be assessed quantitatively, non-invasively, and in real time. To this end, this proposal focuses on a multi-modality optical approach to assess melanoma tumor margins based on quantitative molecular and structural surrogate biomarkers. Two molecular imaging methods will be used. The first is pump-probe microscopy, a nonlinear optical method that achieves quantitative molecular contrast of pigments, including eumelanin and phoemelanin (12). This method is uniquely suited for diagnosing melanoma since eu- and pheo-melanin play an important role in the malignant progression of this disease (12). The second method, pump-probe nonlinear phase dispersion spectroscopy (PP-NLDS), yields complementary information by probing phase changes resulting from cross phase modulation, a ubiquitous effect that provides structural information (35, 36), and molecular reorientation effects (i.e., optical Kerr effect), which provides sensitivity to the diffusive properties of aqueous environments (20, 32). Additional structural information will be obtained from multiphoton autofluorescence, second harmonic generation, and optical coherence microscopy (OCM). The first aim of the proposal is to develop a combined, multi-modality system. The strategy is to modify an existing custom-built microscope to incorporate PP-NLDS and OCM, both detected with the same instrumentation. Aim 2 will focus on developing proper criteria for margin assessment using a human grafted skin mouse model. We expect that lesions with positive margins will contain a combination of markers including a shift toward eumlanic pigment, dermal microsatellite, disorganized tissue architecture, differences in intracellular environments, and enlarged cell nuclei. Aim 3 will validate the approach using human specimens from cadavers and patients undergoing surgery. All imaging results will be correlated with histopathology to validate the approach. Successful completion of this work will pave the way for an instrument that provides quantitative biomarkers to assess melanoma tumor margins, thereby improving the chances of surgical success in the first excision and in a timely manner, without altering accepted clinical paradigms.