Intraoperative Tumor Visualization Research Articles

Improved ex vivo fluorescence imaging of human head and neck cancer using the peptide tracer TPP-IRDye800 targeting membrane-bound Hsp70 on tumor cells.

The primary goal of surgery in HNSCC is the complete resection of tumor cells with maximum preservation of normal tissue. The membrane Hsp70-targeting fluorescence labelled peptide TPP-IRDye800 represents a promising tool for real-time intraoperative tumor visualization, enabling the detection of true tumor margins, critical isles of high-grade dysplasia and LN metastases. Membrane Hsp70 (mHsp70) expression on HNSCC cell lines and primary HNSCC was determined by flow cytometry and fluorescence microscopy using FITC-conjugated mAb cmHsp70.1 and TPP. TPP-IRDye800 was sprayed on freshly resected tumor material of immunohistochemically confirmed HNSCC and LN metastases for tumor imaging. TBRs were compared using TPP-IRDye800 and Cetuximab-IRDye680, recognizing EGFR. mHsp70 expressing HNSCC cells specifically bind and internalize TPP in vitro. The TBR (2.56 ± 0.39) and AUC [0.98 CI, 0.95-1.00 vs. 0.91 CI, 0.85-0.97] of TPP-IRDye800 on primary HNSCC was significantly higher than Cetuximab-IRDye680 (1.61 ± 0.39) (p = 0.0068) and TPP-IRDye800 provided a superior tumor delineation. Fluorescence imaging showed higher AUC values than a visual inspection by surgeons [0.97 CI, 0.94-1.00 vs. 0.92 CI, 0.88-0.97] (p = 0.048). LN metastases could be visualized using TPP-IRDye800. Real-time tissue delineation was confirmed using the clinically applied KARL-STORZ imaging system. TPP-IRDye800 is a promising fluorescence imaging probe for HNSCC.

Immunohistochemical Evaluation of Cathepsin B, L, and S Expression in Breast Cancer Patients.

Cysteine cathepsins are proteases that play a role in normal cellular physiology and neoplastic transformation. Elevated expression and enzymatic activity of cathepsins in breast cancer (BCa) indicates their potential as a target for tumor imaging. In particular cathepsin B (CTSB), L (CTSL), and S (CTSS) are used as targets for near-infrared (NIR) fluorescence imaging (FI), a technique that allows real-time intraoperative tumor visualization and resection margin assessment. Therefore, this immunohistochemical study explores CTSB, CTSL, and CTSS expression levels in a large breast cancer patient cohort, to investigate in which BCa patients the use of cathepsin-targeted NIR FI may have added value. Protein expression was analyzed in tumor tissue microarrays (TMA) of BCa patients using immunohistochemistry and quantified as a total immunostaining score (TIS), ranging from 0-12. In total, the tissues of 557 BCa patients were included in the TMA. CTSB, CTSL, and CTSS were successfully scored in respectively 340, 373 and 252 tumors. All tumors showed CTSB, CTSL, and/or CTSS expression to some extent (TIS > 0). CTSB, CTSL, and CTSS expression was scored as high (TIS > 6) in respectively 28%, 80%, and 18% of tumors. In 89% of the tumors scored for all three cathepsins, the expression level of one or more of these proteases was scored as high (TIS > 6). Tumors showed significantly higher cathepsin expression levels with advancing Bloom-Richardson grade (p < 0.05). Cathepsin expression was highest in estrogen receptor (ER)-negative, human epidermal growth factor receptor 2(HER2)-positive and triple-negative (TN) tumors. There was no significant difference in cathepsin expression between tumors that were treated with neoadjuvant systemic therapy and tumors that were not. The expression of at least one of the cysteine cathepsins B, L and S in all breast tumor tissues tested suggests that cathepsin-activatable imaging agents with broad reactivity for these three proteases will likely be effective in the vast majority of breast cancer patients, regardless of molecular subtype and treatment status. Patients with high grade ER-negative, HER2-positive, or TN tumors might show higher imaging signals.

AB025. Efficacy of 5-ALA brightness analysis for malignant brain tumor surgery.

For fluorescence-guided neurosurgery, 5-aminolevulinic acid (5-ALA) is widely used for intraoperative tumor visualization. We quantified the brightness of 5-ALA by Image J and report the pathological results of glioma, and non-tumor lesions. From 2019 to 2023, we investigated 27 high-grade glioma patients who underwent surgery with 5-ALA. Twenty-three cases of glioblastoma (GBM) and four cases of anaplastic astrocytoma (AA) were examined. The pathological diagnosis was based on the classification of World Health Organization (WHO) 2016. The 5-ALA was administered before surgery, and the 5-ALA brightness was quantified. Other than high-grade glioma, four low-grade gliomas (LGGs), two radiation necrosis, and one inflammation patients were evaluated by Image J. In GBM, the mean brightness was 134.5±65.4, except for one negative case. AA showed the mean brightness with 180.5±51.6. All LGGs showed negative in the brightness. In two radiation necrosis cases, the mean brightness was 139.5±37.4. In one inflammatory case, the brightness was 239, but after the lesion removed, the adjacent brain parenchyma showed bright, and the border was not clear. In one GBM case, the ventricle was opened, and the brightness difference between the tumor and the ventricular wall was observed. 5-ALA brightness analysis by Image J would be helpful to distinguish between malignant glioma and LGG, and other non-tumor lesions to support rapid pathological diagnosis. Also, it would be useful to distinguish between the tumor and the ventricle wall. As for radiation necrosis and inflammation, border of the lesion is unclear.

Preclinical evaluation of AGTR1-Targeting molecular probe for colorectal cancer imaging in orthotopic and liver metastasis mouse models

Despite advancements in colorectal cancer (CRC) treatment, the prognosis remains unfavorable for patients with distant liver metastasis. Fluorescence molecular imaging with specific probes is increasingly used to guide CRC surgical resection in real-time and treatment planning. Here, we demonstrate the targeted imaging capacity of an MPA-PEG4-N3-Ang II probe labeled with near-infrared (NIR) fluorescent dye targeting the angiotensin II (Ang II) type 1 receptor (AGTR1) that is significantly upregulated in CRC. MPA-PEG4-N3-Ang II was highly selective and specific to in vitro tumor cells and in vivo tumors in a mouse CRC xenograft model. The favorable ex vivo imaging and in vivo biodistribution of MPA-PEG4-N3-Ang II afforded tumor-specific accumulation with low background and >10 contrast tumor-to-colorectal values in multiple subcutaneous CRC models at 8 h following injection. Biodistribution analysis confirmed the probe's high uptake in HT29 and HCT116 orthotopic and liver metastatic models of CRC with signal-to-noise ratio (SNR) values of tumor-to-colorectal and -liver fluorescence of 5.8 ± 0.6, 5.3 ± 0.7, and 2.7 ± 0.5, 2.6 ± 0.5, respectively, enabling high-contrast intraoperative tumor visualization for surgical navigation. Given its rapid tumor targeting, precise tumor boundary delineation, durable tumor retention and docking study, MPA-PEG4-N3-Ang II is a promising high-contrast imaging agent for the clinical detection of CRC.

NIMG-49. INTRAOPERATIVE 5-ALA INTENSITY ANALYSIS OF GLIOMA USING IMAGE J

Abstract PURPOSE: The 5 aminolevulinic acid (5-ALA) is used for intraoperative tumor visualization in neurosurgery. We hypothesized that glioma may vary in malignancy according to the brightness of 5-ALA. We quantified the brightness of 5-ALA and compared it with the pathological results of glioma. MATERIALS AND METHODS: From 2019 to 2023, we investigated 35 glioma patients who underwent surgery with 5-ALA. Twenty-three cases of glioblastoma (GBM), four cases of anaplastic astrocytoma (AA), three cases of oligodendroglioma (OL), one case of anaplastic oligodendroglioma (AO), and four cases of low grade glioma (LGG) were examined. The pathological diagnosis was based on classification of WHO 2016. The 5-ALA was administered before surgery, and the 5-ALA brightness were quantified. RESULTS: In GBM, the mean brightness was 134.5 ± 65.4, except for one negative case. AA showed the mean brightness with 180.5 ± 51.6. Three cases of OL showed the mean brightness with 78.6 ± 32.7. AO showed the brightness with 153. All LGGs showed negative in the brightness. CONCLUSION: The 5-ALA showed higher brightness in GBM , AA, AO and negative in LGG. In the future, we will accumulate cases and add further studies.

Predictive factors and repetition numbers for intraoperative additional resection of initially involved soft tissue resection margins in oral squamous cell carcinoma: a retrospective study

BackgroundIntraoperative additional resection (IAR) of initially microscopically involved soft tissue resection margins negatively impacts tumor recurrence in oral squamous cell carcinoma (OSCC). Increasing the selected initial macroscopic resection margin distance beyond the tumor tissue may help prevent IAR; however, the existence of predictive factors for IAR and IAR repetition numbers remains unclear. This study aimed to identify predictive factors for IAR and to evaluate the IAR repetition numbers in soft tissue for surgically treated OSCC.MethodsA cohort of 197 patients surgically treated for OSCC between 2008 and 2019 was retrospectively reviewed (44 patients with IAR and 153 patients without IAR). Clinical parameters (tumor location, midline involvement, clinical T-status, time between staging imaging and surgery, bone resection, monopolar use, and reconstruction flap size) and histopathological parameters (pathologic T-status [pT-status], grading, vascular invasion, and lymphatic invasion) of the two groups were compared.ResultsPatients with and without IAR differed in their histopathological parameters, such as pT-status above 2 (47.7% vs. 28.1%, p = 0.014) and lymphatic invasion (13.6% vs. 4.6%, p = 0.033); however, their clinical parameters were similar (all p > 0.05). Only pT-status above 2 was predictive for IAR in a multivariable regression analysis (odds ratio 2.062 [confidence interval 1.008–4.221], p = 0.048). The IAR repetition numbers varied from zero to two (zero = 84.4%, one = 11.4%, and two = 2.3%).ConclusionsOnly postoperative available pT-status was identified as a predictive factor for IAR, underscoring the importance of improving preoperative or intraoperative tumor visualization in OSCC before selecting the initial macroscopic resection margin distance to avoid IAR.

OS03.7.A AUTOFLUORESCENCE BASED RECOGNITION OF BRAIN TUMORS WITH A CONVOLUTIONAL NEURAL NETWORK

Abstract BACKGROUND The delineation of malignant brain tumors from surrounding tissue is crucial for maximal safe resection. Existing tools for intraoperative tumor visualization lack the possibility of real-time tissue classification. We therefore built a coherent fiber bundle (CFB) endoscope for autofluorescence imaging and trained a convolutional neural network (CNN) to distinguish tumor tissue (T) and non-tumor tissue (nT) in an ex vivo proof of concept study. MATERIAL AND METHODS The T-cohort included frozen tissue sections of 35 glioblastoma and 30 brain metastasis patients (10 breast cancer, 10 melanoma, 10 lung cancer). The nT-cohort comprised frozen tissue sections of hippocampi and temporal lobes from 23 patients undergoing surgery for epilepsy treatment. For image acquisition, a laser with excitation wavelength of 473 nm was coupled into a CFB and tissue autofluorescence between 500-550 nm was collected using a bandpass filter. The acquisition time of the camera was set to 700 ms. A CNN (VGG-19) was trained to differentiate the labels T from nT wherein the ground truth was defined by histopathological diagnosis. Training, validation and test sets were randomly generated across all patients in 66.6% / 16.6% / 16.6% proportion. Set generation was repeated 5 times and reported results for the independent test sets are mean values of all iterations. RESULTS In total, 3259 CFB images were acquired (T:2408, nT:851), with usually 40 CFB images per sample. Removal of CFB background was not improving classification accuracy and thus, was not performed. The classification accuracy of the CNN with a threshold probability of 0.5 for T-label assignment was 0.986 (sensitivity: 0.988, specificity: 0.982) for analysis of images and 0.98 (sensitivity: 0.971, specificity: 1) when calculating the result for each patient. To simulate clinical application where correct identification of healthy tissue is essential, we created a specificity-tuned CNN with adjusted threshold probability of 0.9 for T-label assignment. This led to a decreased accuracy of 0.945 but increased specificity on the image evaluation approach to 1 (sensitivity: 0.92). CONCLUSION Autofluorescence imaging through a CFB is capable of ex vivo recognition of brain tumor tissue with a CNN. The simplicity of the setup and minimal need for data pre-processing allows for intraoperative implementation and potentially enables real-time in vivo brain tumor delineation. To realize intraoperative transition, analysis of ex vivo bulk tissue will be upcoming to capture the signal complexity caused by tissue depth.

Advancements in Intraoperative Near-Infrared Fluorescence Imaging for Accurate Tumor Resection: A Promising Technique for Improved Surgical Outcomes and Patient Survival.

Clear surgical margins for solid tumor resection are essential for preventing cancer recurrence and improving overall patient survival. Complete resection of tumors is often limited by a surgeon's ability to accurately locate malignant tissues and differentiate them from healthy tissue. Therefore, techniques or imaging modalities are required that would ease the identification and resection of tumors by real-time intraoperative visualization of tumors. Although conventional imaging techniques such as positron emission tomography (PET), computed tomography (CT), magnetic resonance imaging (MRI), or radiography play an essential role in preoperative diagnostics, these cannot be utilized in intraoperative tumor detection due to their large size, high cost, long imaging time, and lack of cancer specificity. The inception of several imaging techniques has paved the way to intraoperative tumor margin detection with a high degree of sensitivity and specificity. Particularly, molecular imaging using near-infrared fluorescence (NIRF) based nanoprobes provides superior imaging quality due to high signal-to-noise ratio, deep penetration to tissues, and low autofluorescence, enabling accurate tumor resection and improved survival rates. In this review, we discuss the recent developments in imaging technologies, specifically focusing on NIRF nanoprobes that aid in highly specific intraoperative surgeries with real-time recognition of tumor margins.

5-aminolevulinic acid, fluorescein sodium, and indocyanine green for glioma margin detection: analysis of operating wide-field and confocal microscopy in glioma models of various grades.

Surgical resection remains the first-line treatment for gliomas. Several fluorescent dyes are currently in use to augment intraoperative tumor visualization, but information on their comparative effectiveness is lacking. We performed systematic assessment of fluorescein sodium (FNa), 5-aminolevulinic acid (5-ALA)-induced protoporphyrin IX (PpIX), and indocyanine green (ICG) fluorescence in various glioma models using advanced fluorescence imaging techniques. Four glioma models were used: GL261 (high-grade model), GB3 (low-grade model), and an in utero electroporation model with and without red fluorescence protein (IUE +RFP and IUE -RFP, respectively) (intermediate-to-low-grade model). Animals underwent 5-ALA, FNa, and ICG injections and craniectomy. Brain tissue samples underwent fluorescent imaging using a wide-field operative microscope and a benchtop confocal microscope and were submitted for histologic analysis. Our systematic analysis showed that wide-field imaging of highly malignant gliomas is equally efficient with 5-ALA, FNa, and ICG, although FNa is associated with more false-positive staining of the normal brain. In low-grade gliomas, wide-field imaging cannot detect ICG staining, can detect FNa in only 50% of specimens, and is not sensitive enough for PpIX detection. With confocal imaging of low-intermediate grade glioma models, PpIX outperformed FNa. Overall, compared to wide-field imaging, confocal microscopy significantly improved diagnostic accuracy and was better at detecting low concentrations of PpIX and FNa, resulting in improved tumor delineation. Neither PpIX, FNa, nor ICG delineated all tumor boundaries in studied tumor models, which emphasizes the need for novel visualization technologies and molecular probes to guide glioma resection. Simultaneous administration of 5-ALA and FNa with use of cellular-resolution imaging modalities may provide additional information for margin detection and may facilitate maximal glioma resection.

878 Development of a Near Infrared Somatostatin Receptor Type 2 (SSTR2)-Targeted Probe for Fluorescence-Guided Surgery in Meningiomas

INTRODUCTION: Meningioma frequently recur at the primary resection site requiring improvement in the intraoperative tumor visualization and surgical technique. To guide the resection, this study aims to develop a highly specific and sensitive optical method for intraoperative meningioma imaging. By suitable cameras fluorescent dyes in the near infrared (NIR, 700-1000nm) can be detected up to few millimeters in depth in real time. METHODS: We invented a SSTR2 targeting probe by linking a somatostatin analogue (TATE, Tyr(3)Thr(8)-Octreotid) to a new developed dye (sNIR). In the optical characterization, our probe showed similarities to indocyanine green and stability as well as bleaching kinetic were investigated in vitro. The probe was intravenously injected in native mice and detected by a custom-built camera setup to evaluate pharmacokinetics and biodistribution, focusing on physiologically SSTR2 overexpressing organs that were validated by immunohistochemistry (IHC). As proof-of-principle, fluorescence-guided meningioma resection was performed in an ectopic and orthotopic IOMM-Lee mouse model. RESULTS: Our probe is stable in aqueous solution up to 12h with favorable optical properties regarding pharmacokinetics, biodistribution and photostability compared to its IRDye800 analogue. In vivo, we observed a highly specific signal uptake in the gastric epithelium, pancreas and pituitary (all are overexpressing SSTR2) compared to autofluorescence and negative controls. Given the intermediate SSTR2 expression in the meningioma mouse model, we could still detect a mean tumor-to-brain ratio of 6.12 (n = 5; SD 1.8) allowing the fluorescence-guidance to be feasible. CONCLUSIONS: Our preclinical results indicate that the new SSTR2-targeted fluorescent probe is stable in various media over time and capable of specifically targeting the SSTR2 in vitro and in vivo potentially enabling sensitive and specific meningioma fluorescent-guided surgery in the future.

KS04.5.A Development of a somatostatin receptor type 2 (SSTR2)-targeted probe for near infrared fluorescence guided meningioma surgery

Abstract Background Meningioma mostly recur at the primary resection bed requiring improvement in the intraoperative tumor visualization and surgical technique. This study aims to develop a highly specific and sensitive optical probe for intraoperative meningioma imaging to guide the tumor resection. Fluorescent dyes in the near infrared (NIR I, 700-1000nm) and shortwave infrared (NIR II, 1000-2000nm) allow detection by suitable cameras up to few millimeters in depth in real time. Our objective is to invent a fluorescent NIR I and II probe specifically targeting the somatostatin receptor type 2 (SSTR2) reliably overexpressed in meningioma independently of WHO grade and subtype. Material and Methods By linking a somatostatin analogue (TATE, Tyr(3)Thr(8)-Octreotid) to a new developed dye (sNIR), we synthesized a SSTR2 specific probe. Its optical properties are comparable to indocyanine green and stability as well as bleaching kinetic were observed in vitro. The probe was intravenously injected in native mice to evaluate pharmacokinetics and biodistribution and detected by a custom-built camera setup focusing on physiologically highly expressing SSTR2 tissues validated by immunohistochemistry (IHC). As proof-of-principle fluorescence guided tumor resection was performed in an ectopic and orthotopic meningioma IOMM-Lee mouse model. Results The SSTR2-sNIR probe is stable in aqueous solution up to 12h with favorable optical properties regarding kinetics, biodistribution and photostability compared to its IRDye800 analogue. In vivo, we observed a highly specific physiological signal uptake in the gastric epithelium, the pancreas and pituitary (they are highly expressing SSTR2 on IHC) compared to autofluorescence and negative controls (unlabeled dye and scrambled version). Given the moderate SSTR2 expression in the preclinical meningioma model we could still detect a mean tumor-to-brain ratio of 6.12 (n = 5; SD 1.8) in the orthotopic meningioma mouse model, allowing the fluorescence guidance to be feasible. Conclusion Our preclinical results demonstrate that the innovative SSTR2-targeted fluorescent probe is stable in various biological media over time and capable of specifically targeting the SSTR2 in vitro and in vivo potentially enabling sensitive and specific meningioma fluorescent guided surgery in the future.

The application of fluorescein sodium for the resection of medulloblastoma

IntroductionSurgical resection of medulloblastoma (MB) remains a challenge. At present, a variety of tracers have been used for intraoperative tumor visualization. However, there are few reports on the intraoperative visualization of MB. Hence, we reported our experience of applying fluorescein sodium (FS) in MB surgery.MethodsWe retrospectively analyzed the clinical information of patients with MB confirmed by surgery and pathology from January 2016 to December 2020 from Sun Yat-sen University Cancer Center. A total of 62 patients were enrolled, of which 27 received intraoperative FS and 35 did not. The intraoperative dose of FS was 3 mg/kg.ResultsAmong the 62 patients, 42 were males, and twenty were females. The age of onset in the FS group was 9.588 ± 7.322, which in the non-fluorescein sodium group was 13.469 ± 10.968, p = 0.198. We did not find significant differences in tumor location, tumor size, tumor resection, tumor histology, and preoperative symptoms (hydrocephalus, headache, vomit, balance disorder) between the groups. There was no significant difference in the postoperative symptoms (hydrocephalus, headache, vomiting, balance disorder, and cerebellar mutism). However, patients in the FS group had a relatively low incidence of balance disorder and cerebellar mutism. There was definite fluorescence of tumor in all cases of the FS group, and even the tiny metastatic lesion was visible. No case had side effects related to the use of FS.ConclusionsFS is safe and effective in MB surgery. Whether the application of FS for surgery can reduce complications remains to be studied in the future.

Utility of intraoperative real-time near-infrared fluorescence surgery for spinal schwannoma.

The authors report the first cases of fluorescence-guided spinal surgery of schwannomas using near-infrared fluorescence imaging with the delayed window indocyanine (ICG) green (DWIG) technique for accurate real-time intraoperative tumor visualization.Patients with intradural spinal schwannomas received 0.5 mg/kg ICG at the beginning of surgery. After 1 hour, using the DWIG technique, near-infrared spectroscopy (NIRS) detected the spinal schwannomas, showing the exact tumor location and boundaries. DWIG with NIRS microscopy confirmed the exact location of spinal schwannomas before and after opening of the dura mater, thereby facilitating successful tumor dissection from the surrounding tissues, tumor resection, and confirmation of tumor removal.The video can be found here: https://stream.cadmore.media/r10.3171/2021.10.FOCVID21158

Application of an indocyanine green-mediated fluorescence imaging navigation system in detecting mice tumors.

BackgroundSurgical operation plays an important role in the treatment of cancer. The success of the operation lies in the complete removal of the primary and disseminated tumor tissue while preserving the normal tissue. The development of optical molecular image navigation technology has provided a new option for intraoperative tumor visualization. In this study, a fluorescence imaging navigation system was used to detect the diameter of mice tumors and provide experimental evidence for the further development of digital diagnosis and treatment equipment.MethodsThe minimum detection concentration in vitro of the fluorescence imaging navigation system for indocyanine green (ICG) was first detected, then 120 female Institute of Cancer Research (ICR) mice and 120 female BALB/c nude mice were randomly divided into three groups by weight, high-dose (H, 4 mg/kg), middle-dose (M, 2 mg/kg), and low-dose (L, 1 mg/kg) groups of ICG solution. After inoculating solid tumors, high, medium, and low doses of ICG were injected via the tail vein, and the tumor diameter was measured by a fluorescence imaging navigation system and vernier caliper within 24 hours of injection.ResultsThe minimum detectable diameter of the system could reach 0.2 mm compared with the vernier caliper, and the actual measurement error was within 0.2 mm.ConclusionsA fluorescence imaging navigation system has high accuracy and sensitivity in the application of tumor detection, which may assist the clinical diagnosis and treatment of tumors.

C-MET Receptor-Targeted Fluorescence on the Road to Image-Guided Surgery in Penile Squamous Cell Carcinoma Patients.

In penile squamous cell carcinoma (pSCC), primary surgery aims to obtain oncologically safe margins while minimizing mutilation. Surgical guidance provided by receptor-specific tracers could potentially improve margin detection and reduce unnecessary excision of healthy tissue. Here, we present the first results of a prospective feasibility study for real-time intraoperative visualization of pSCC using a fluorescent mesenchymal-epithelial transition factor (c-MET) receptor targeting tracer (EMI-137). Methods: EMI-137 tracer performance was initially assessed ex vivo (n = 10) via incubation of freshly excised pSCC in a solution containing EMI-137 (500 nM). The invivo potential of c-MET targeting and intraoperative tumor visualization was assessed after intravenous administration of EMI-137 to 5 pSCC patients scheduled for surgical resection using a cyanine-5 fluorescence camera. Fluorescence imaging results were related to standard pathologic tumor evaluation and c-MET immunohistochemistry. Three of the 5 invivo patients also underwent a sentinel node resection after local administration of the hybrid tracer indocyanine green- 99mTc-nanocolloid, which could be imaged using a near-infrared fluorescence camera. Results: No tracer-related adverse events were encountered. Both ex vivo and invivo, EMI-137 enabled c-MET-based tumor visualization in all patients. Histopathologic analyses showed that all pSCCs expressed c-MET, with expression levels of at least 70% in 14 of 15 patients. Moreover, the highest c-MET expression levels were seen on the outside rim of the tumors, and a visual correlation was found between c-MET expression and fluorescence signal intensity. No complications were encountered when combining primary tumor targeting with lymphatic mapping. As such, simultaneous use of cyanine-5 and indocyanine green in the same patient proved to be feasible. Conclusion: Fluorescence imaging of c-MET receptor- expressing pSCC tumors after intravenous injection of EMI-137 was shown to be feasible and can be combined with fluorescence-based lymphatic mapping. This combination is unique and paves the way toward further development of this surgical guidance approach.

5-ALA fluorescence for intraoperative visualization of spinal ependymal tumors and identification of unexpected residual tumor tissue: experience in 31 patients.

Gross-total resection (GTR) is the treatment of choice in the majority of patients suffering from spinal ependymal tumors. In such tumors, the extent of resection (EOR) is considered the key factor for tumor recurrence and thus patient prognosis. However, incomplete resection is not uncommon and leads to increased risk of tumor recurrence. One important cause of incomplete resection is insufficient intraoperative visualization of tumor tissue as well as residual tumor tissue. Therefore, the authors investigated the value of 5-aminolevulinic acid (5-ALA)-induced fluorescence in a series of spinal ependymal tumors for improved tumor visualization. Adult patients who underwent preoperative 5-ALA administration and surgery for a spinal ependymal tumor were included in this study. For each tumor, a conventional white-light microsurgical resection was performed. Additionally, the fluorescence status (strong, vague, or no fluorescence) and fluorescence homogeneity (homogenous or inhomogeneous) of the spinal ependymal tumors were evaluated during surgery using a modified neurosurgical microscope. In intramedullary tumor cases with assumed GTR, the resection cavity was investigated for potential residual fluorescing foci under white-light microscopy. In cases with residual fluorescing foci, these areas were safely resected and the corresponding samples were histopathologically screened for the presence of tumor tissue. In total, 31 spinal ependymal tumors, including 27 intramedullary tumors and 4 intradural extramedullary tumors, were included in this study. Visible fluorescence was observed in the majority of spinal ependymal tumors (n = 25, 81%). Of those, strong fluorescence was noted in 23 of these cases (92%), whereas vague fluorescence was present in 2 cases (8%). In contrast, no fluorescence was observed in the remaining 6 tumors (19%). Most ependymal tumors demonstrated an inhomogeneous fluorescence effect (17 of 25 cases, 68%). After assumed GTR in intramedullary tumors (n = 15), unexpected residual fluorescing foci within the resection cavity could be detected in 5 tumors (33%). These residual fluorescing foci histopathologically corresponded to residual tumor tissue in all cases. This study indicates that 5-ALA fluorescence makes it possible to visualize the majority of spinal ependymal tumors during surgery. Unexpected residual tumor tissue could be detected with the assistance of 5-ALA fluorescence in approximately one-third of analyzed intramedullary tumors. Thus, 5-ALA fluorescence might be useful to increase the EOR, particularly in intramedullary ependymal tumors, in order to reduce the risk of tumor recurrence.

Fluorescence guided surgery for pituitary adenomas.

Resection of pituitary adenomas presents a number of unique challenges in neuro-oncology. The proximity of these lesions to key vascular and endocrine structures as well as the need to interpret neuronavigation in the context of shifting tumor position increases the complexity of the operation. More recently, substantial advances in fluorescence-guided surgery have been demonstrated to facilitate the identification of numerous tumor types and result in increased rates of complete resection and overall survival. A review of the literature was performed, and data regarding the mechanism of the fluorescence agents, their administration, and intraoperative tumor visualization were extracted. Both in vitro and in vivo studies were assessed. The application of these agents to pituitary tumors, their advantages and limitations, as well as future directions are presented here. Numerous laboratory and clinical studies have described the use of 5-ALA, fluorescein, indocyanine green, and OTL38 in pituitary lesions. All of these drugs have been demonstrated to accumulate in tumor cells. Several studies have reported the successful use of the majority of the agents in inducing intraoperative tumor fluorescence. However, their sensitivity and specificity varies across the literature and between functioning and non-functioning adenomas. At present, numerous studies have shown the feasibility and safety of these agents for pituitary adenomas. However, further research is needed to assess the applicability of fluorescence-guided surgery across different tumor subtypes as well as explore the relationship between their use and postoperative clinical outcomes.

IRDye800CW labeled uPAR-targeting peptide for fluorescence-guided glioblastoma surgery: Preclinical studies in orthotopic xenografts.

Glioblastoma (GBM) is a devastating cancer with basically no curative treatment. Even with aggressive treatment, the median survival is disappointing 14 months. Surgery remains the key treatment and the postoperative survival is determined by the extent of resection. Unfortunately, the invasive growth with irregular infiltrating margins complicates an optimal surgical resection. Precise intraoperative tumor visualization is therefore highly needed and molecular targeted near-infrared (NIR) fluorescence imaging potentially constitutes such a tool. The urokinase-type Plasminogen Activator Receptor (uPAR) is expressed in most solid cancers primarily at the invading front and the adjacent activated peritumoral stroma making it an attractive target for targeted fluorescence imaging. The purpose of this study was to develop and evaluate a new uPAR-targeted optical probe, IRDye800CW-AE344, for fluorescence guided surgery (FGS).Methods: In the present study we characterized the fluorescent probe with regard to binding affinity, optical properties, and plasma stability. Further, in vivo imaging characterization was performed in nude mice with orthotopic human patient derived glioblastoma xenografts, and we performed head-to-head comparison within FGS between our probe and the traditional procedure using 5-ALA. Finally, the blood-brain barrier (BBB) penetration was characterized in a 3D BBB spheroid model.Results: The probe effectively visualized GBM in vivo with a tumor-to-background ratio (TBR) above 4.5 between 1 to 12 h post injection and could be used for FGS of orthotopic human glioblastoma xenografts in mice where it was superior to 5-ALA. The probe showed a favorable safety profile with no evidence of any acute toxicity. Finally, the 3D BBB model showed uptake of the probe into the spheroids indicating that the probe crosses the BBB.Conclusion: IRDye800CW-AE344 is a promising uPAR-targeted optical probe for FGS and a candidate for translation into human use.

Contrast-Enhanced Ultrasound for Intraoperative Tumor Contour Definition During Complex Partial Nephrectomy

Contrast-enhanced ultrasound (CEUS) is an imaging modality that utilizes a non-nephrotoxic, intravascular microbubble agent to capture real-time perfusion of the target organ. CEUS has demonstrated utility for the evaluation and surveillance of indeterminate renal lesions in patients who are unable to receive conventional contrast agents, yet its potential utility in renal surgery remains poorly defined in the current literature. We present a series of 4 patients who underwent successful intraoperative CEUS during partial nephrectomy (PN) to help define renal mass contour and proximity to vascular structures in an attempt to optimize renal function after PN. All cases were endophytic with moderate-high complexity (R.E.N.A.L. scores>9). CEUS was safely performed in all cases providing excellent tumor contour definition. In one case, CEUS provided better visualization of tumor proximity within the hilum than conventional US, facilitating surgical resection and reconstruction. In another case, CEUS provided better definition of tumor contour, allowing wedge resection rather than heminephrectomy. Margins were negative in all cases. The average global renal function preserved following PN was 90%. While grey-scale US with color Doppler is usually sufficient for intraoperative tumor visualization, CEUS can be considered during PN for complex endophytic, infiltrative or hilar tumors where accurate delineation of tumor contour is critical.

Development of a drug-device combination for fluorescence-guided surgery in neuroendocrine tumors.

.Significance: The use of cancer-targeted contrast agents in fluorescence-guided surgery (FGS) has the potential to improve intraoperative visualization of tumors and surgical margins. However, evaluation of their translational potential is challenging.Aim: We examined the utility of a somatostatin receptor subtype-2 (SSTR2)-targeted fluorescent agent in combination with a benchtop near-infrared fluorescence (NIRF) imaging system to visualize mouse xenografts under conditions that simulate the clinical FGS workflow for open surgical procedures.Approach: The dual-labeled somatostatin analog, -MMC(IR800)-TOC, was injected into mice () implanted with SSTR2-expressing tumors and imaged with the customized OnLume NIRF imaging system (Madison, Wisconsin). In vivo and ex vivo imaging were performed under ambient light. The optimal dose (0.2, 0.5, and 2 nmol) and imaging time point (3, 24, 48, and 72 h) were determined using contrast-to-noise ratio (CNR) as the image quality parameter. Video captures of tumor resections were obtained to provide an FGS readout that is representative of clinical utility. Finally, a log-transformed linear regression model was fitted to assess congruence between fluorescence readouts and the underlying drug distribution.Results: The drug–device combination provided high in vivo and ex vivo contrast (, except lung at 3 h) at all time points with the optimal dose of 2 nmol. The optimal imaging time point was 24-h post-injection, where were achieved in tissues of interest (i.e., pancreas, small intestine, stomach, and lung). Intraoperative FGS showed excellent utility for examination of the tumor cavity pre- and post-resection. The relationship between fluorescence readouts and gamma counts was linear and strongly correlated (, ; ; ).Conclusion: The innovative OnLume NIRF imaging system enhanced the evaluation of -MMC(IR800)-TOC in tumor models. These components comprise a promising drug–device combination for FGS in patients with SSTR2-expressing tumors.