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Background: Anastomotic leakage is a severe postoperative complication in colorectal surgery and compromised bowel perfusion is considered a major contributing factor. Conventional methods to assess bowel perfusion have a low predictive value for anastomotic leakage. We therefore aimed to evaluate the efficacy of real-time assessment with near-infrared (NIR) fluorescence imaging with indocyanine green (ICG) in the prevention of anastomotic leakage., Methods: This multicentre, randomised, controlled, phase 3 trial was done in eight hospitals in the Netherlands. We included adults (aged >18 years) who were scheduled for laparoscopic or robotic colorectal surgery (with planned primary anastomosis) for benign and malignant diseases. Preoperatively, patients were randomly assigned (1:1) to fluorescence-guided bowel anastomosis (FGBA) or conventional bowel anastomosis (CBA) by variable block randomisation (block sizes 4, 6, and 8) and stratified by site. The operating surgeon and investigators analysing the data were not masked to group assignment. Patients were unmasked after the surgical procedure or after study end. In the FGBA group, surgeons marked anastomosis levels per conventional perfusion assessment and then administered 5 mg of ICG by 2 mL intravenous bolus. They assessed bowel perfusion using NIR fluorescence imaging and adjusted (or kept) transection lines accordingly. Only conventional methods for bowel perfusion assessment were used in the CBA group. The primary outcome was the difference in the rate of clinically relevant anastomotic leakage (ie, requiring active therapeutic intervention but manageable without reoperation [grade B] or requiring reoperation [grade C], per the International Study Group of Rectal Cancer) between the FGBA group and the CBA group within 90 days post-surgery. The primary outcome and safety were assessed in the intention-to-treat population. This study was registered with ToetsingOnline.nl (NL7502) and ClinicalTrials.gov (NCT04712032) and is complete., Findings: Between July 2, 2020, and Feb 21, 2023, 982 patients were enrolled, of whom 490 were assigned to FGBA and 492 were assigned to CBA. After excluding 51 patients, the intention-to-treat population comprised 931 (463 assigned FGBA and 468 assigned CBA). Patients had a median age of 68·0 years (IQR 59·0-75·0) and 485 (52%) were male and 446 (48%) were female. Ethnicity data were not available. The overall 90-day rate of clinically relevant anastomotic leakage was not significantly different between the FGBA group (32 [7%] of 463 patients) and the CBA group (42 [9%] of 468 patients; relative risk 0·77 [95% CI 0·50-1·20]; p=0·24). No adverse events related to ICG use were observed. 313 serious adverse events in 229 (25%) patients were at 90-day follow-up (159 serious adverse events in 113 [24%] patients in the FGBA group and 154 serious adverse events in 116 [25%] patients in the CBA group). 18 (2%) people died by 90 days (ten in the FGBA group and eight in the CBA group)., Interpretation: ICG NIR fluorescence imaging did not reduce 90-day anastomotic leakage rates in this trial across all types of colorectal surgeries. Further research should be done in subgroups, such as rectosigmoid resections, for which evidence suggests ICG NIR might be beneficial., Funding: Olympus Medical, Diagnostic Green, and Intuitive Foundation., Competing Interests: Declaration of interests We declare no competing interests., (Copyright © 2024 Elsevier Ltd. All rights reserved, including those for text and data mining, AI training, and similar technologies.)

Detailed knowledge of the brain's nerve fiber network is crucial for understanding its function in health and disease. However, mapping fibers with high resolution remains prohibitive in most histological sections because state-of-the-art techniques are incompatible with their preparation. Here, we present a micron-resolution light-scattering-based technique that reveals intricate fiber networks independent of sample preparation for extended fields of view. We uncover fiber structures in both label-free and stained, paraffin-embedded and deparaffinized, newly-prepared and archived, animal and human brain tissues - including whole-brain sections from the BigBrain atlas. We identify altered microstructures in demyelination and hippocampal neurodegeneration, and show key advantages over diffusion magnetic resonance imaging, polarization microscopy, and structure tensor analysis. We also reveal structures in non-brain tissues - including muscle, bone, and blood vessels. Our cost-effective, versatile technique enables studies of intricate fiber networks in any type of histological tissue section, offering a new dimension to neuroscientific and biomedical research., Competing Interests: Competing interests The authors declare no competing interests.

The aim of the study is to assess whether indocyanine green (ICG) fluorescence can replace technetium in the preoperative detection of sentinel lymph nodes (SLN) from cutaneous melanoma. The current golden standard for SLN detection is the radioisotope technetium. A promising alternative is fluorescence imaging (FLI) using ICG. In this study, we enrolled patients undergoing sentinel lymph node biopsy (SLNB) for skin melanoma at the Erasmus Medical Center between November 2022 and July 2023. The SLNB procedure was performed as a standard of care. After general anesthesia, ICG was injected intradermally around the primary tumor site. Both the patient and the surgeon were not blinded for the location of the SLN. FLI was performed before incision, in vivo after incision, and ex vivo. Fluorescent SLNs were confirmed using the gamma probe in all cases. Thirty-two patients were included in this study, and a total of 39 SLNs were harvested. The transcutaneous detection rate of ICG was 21.9%. The combined ex vivo ICG fluorescence and technetium uptake was 94.9%. One SLN contained only ICG (2.6%) and one SLN contained only technetium-uptake (2.6%). FLI using ICG resulted in a relatively low transcutaneous detection, which means that exclusive use of this technique in its present form is not feasible. However, we did find a high accumulation of ICG in the SLN, indicating the potential of ICG in combination with other imaging techniques., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc.)

Purpose: Metastasectomy is a common treatment option for patients with colorectal lung metastases (CLM). Challenges exist with margin assessment and identification of small nodules, especially during minimally invasive surgery. Intraoperative fluorescence imaging has the potential to overcome these challenges. The aim of this study was to assess feasibility of targeting CLM with the carcinoembryonic antigen (CEA) specific fluorescent tracer SGM-101., Methods: This was a prospective, open-label feasibility study. The primary outcome was the number of CLM that showed a true positive fluorescence signal with SGM-101. Fluorescence positive signal was defined as a signal-to-background ratio (SBR) ≥ 1.5. A secondary endpoint was the CEA expression in the colorectal lung metastases, assessed with the immunohistochemistry, and scored by the total immunostaining score., Results: Thirteen patients were included in this study. Positive fluorescence signal with in vivo, back table, and closed-field bread loaf imaging was observed in 31%, 45%, and 94% of the tumors respectively. Median SBRs for the three imaging modalities were 1.00 (IQR: 1.00-1.53), 1.45 (IQR: 1.00-1.89), and 4.81 (IQR: 2.70-7.41). All tumor lesions had a maximum total immunostaining score for CEA expression of 12/12., Conclusion: This study demonstrated the potential of fluorescence imaging of CLM with SGM-101. CEA expression was observed in all tumors, and closed-field imaging showed excellent CEA specific targeting of the tracer to the tumor nodules. The full potential of SGM-101 for in vivo detection of the tracer can be achieved with improved minimal invasive imaging systems and optimal patient selection., Trial Registration: The study was registered in ClinicalTrial.gov under identifier NCT04737213 at February 2021., (© 2023. The Author(s).)

Background: Oral and laryngeal squamous cell carcinoma (OSCC and LSCC) and papillary thyroid carcinoma (PTC) are common head and neck cancers (HNCs) typically treated surgically. Challenges in tumour delineation often lead to inadequate resection margins in OSCC and LSCC, and missed multifocality in PTC. Fluorescence imaging (FLI) using near-infrared tumour-targeting tracers may improve intraoperative identification of malignancy, facilitating precise excision. This study evaluates six potential FLI targets in OSCC, LSCC and PTC., Materials and Methods: Immunohistochemical staining was performed on OSCC (n = 20), LSCC (n = 10) and PTC (n = 10), assessing CEA, c-Met, EpCAM, EGFR, integrin αvβ6 and VEGF-α. Expression was scored (0-12) using the total immunostaining score (TIS) system, and categorized into absent (TIS 0), low (TIS 1-5), moderate (TIS 6-8) or high (TIS 9-12)., Results: Integrin αvβ6 showed significant overexpression in OSCC (TIS: 12; p < 0.001) and LSCC (TIS: 8; p = 0.002), with 80% of OSCC and 90% of LSCC exhibiting moderate-high expression. Similarly, EGFR expression was moderate-high in most OSCC (87.5%; TIS: 8) and universally high in LSCC (100%; TIS: 12). In PTC, EGFR and VEGF-α expressions were low-moderate, but significantly higher than in healthy tissue (TIS: 6; p < 0.006)., Conclusion: This study highlights integrin αvβ6 and EGFR as viable FLI targets in OSCC and LSCC, especially integrin αvβ6 for tumour margin delineation. In PTC, despite lower expressions, the significant overexpression of VEGF-α, c-MET, and EGFR suggests their potential as FLI targets. Our findings support the development of tumour-targeted FLI tracers to improve surgical precision in HNC., (© 2024. The Author(s).)

Radiotherapy in the head-and-neck area is one of the main curative treatment options. However, this comes at the cost of varying levels of normal tissue toxicity, affecting up to 80% of patients. Mucositis can cause pain, weight loss and treatment delays, leading to worse outcomes and a decreased quality of life. Therefore, there is an urgent need for an approach to predicting normal mucosal responses in patients prior to treatment. We here describe an assay to detect irradiation responses in healthy oral mucosa tissue. Mucosa specimens from the oral cavity were obtained after surgical resection, cut into thin slices, irradiated and cultured for three days. Seven samples were irradiated with X-ray, and three additional samples were irradiated with both X-ray and protons. Healthy oral mucosa tissue slices maintained normal morphology and viability for three days. We measured a dose-dependent response to X-ray irradiation and compared X-ray and proton irradiation in the same mucosa sample using standardized automated image analysis. Furthermore, increased levels of inflammation-inducing factors-major drivers of mucositis development-could be detected after irradiation. This model can be utilized for investigating mechanistic aspects of mucositis development and can be developed into an assay to predict radiation-induced toxicity in normal mucosa.

Background: Patients with head and neck squamous cell carcinoma (HNSCC) can develop second primary tumors (SPTs) in the esophagus. Endoscopic screening could lead to detection of SPTs at early stages and improve survival., Methods: We performed a prospective endoscopic screening study in patients with curably treated HNSCC diagnosed between January 2017-July 2021 in a Western country. Screening was performed synchronously (< 6 months) or metachronously (≥ 6 months) after HNSCC diagnosis. Routine imaging for HNSCC consisted of flexible transnasal endoscopy with positron emission tomography/computed tomography or magnetic resonance imaging, depending on primary HNSCC location. The primary outcome was prevalence of SPTs, defined as presence of esophageal high grade dysplasia or squamous cell carcinoma., Results: 202 patients (mean age 65 years, 80.7 % male) underwent 250 screening endoscopies. HNSCC was located in the oropharynx (31.9 %), hypopharynx (26.9 %), larynx (22.2 %), and oral cavity (18.5 %). Endoscopic screening was performed within 6 months (34.0 %), 6 months to 1 year (8.0 %), 1-2 years (33.6 %), and 2-5 years (24.4 %) after HNSCC diagnosis. We detected 11 SPTs in 10 patients (5.0 %, 95 %CI 2.4 %-8.9 %) during synchronous (6/85) and metachronous (5/165) screening. Most patients had early stage SPTs (90 %) and were treated with curative intent with endoscopic resection (80 %). No SPTs in screened patients were detected with routine imaging for HNSCC before endoscopic screening., Conclusion: In 5 % of patients with HNSCC, an SPT was detected with endoscopic screening. Endoscopic screening should be considered in selected HNSCC patients to detect early stage SPTs, based on highest SPT risk and life expectancy according to HNSCC and comorbidities., Competing Interests: The authors declare that they have no conflict of interest., (The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution License, permitting unrestricted use, distribution, and reproduction so long as the original work is properly cited. (https://creativecommons.org/licenses/by/4.0/).)

Background: Indocyanine green near-infrared fluorescence bowel perfusion assessment has shown its potential benefit in preventing anastomotic leakage. However, the surgeon's subjective visual interpretation of the fluorescence signal limits the validity and reproducibility of the technique. Therefore, this study aimed to identify objective quantified bowel perfusion patterns in patients undergoing colorectal surgery using a standardized imaging protocol., Method: A standardized fluorescence video was recorded. Postoperatively, the fluorescence videos were quantified by drawing contiguous region of interests (ROIs) on the bowel. For each ROI, a time-intensity curve was plotted from which perfusion parameters (n = 10) were derived and analyzed. Furthermore, the inter-observer agreement of the surgeon's subjective interpretation of the fluorescence signal was assessed., Results: Twenty patients who underwent colorectal surgery were included in the study. Based on the quantified time-intensity curves, three different perfusion patterns were identified. Similar for both the ileum and colon, perfusion pattern 1 had a steep inflow that reached its peak fluorescence intensity rapidly, followed by a steep outflow. Perfusion pattern 2 had a relatively flat outflow slope immediately followed by its plateau phase. Perfusion pattern 3 only reached its peak fluorescence intensity after 3 min with a slow inflow gradient preceding it. The inter-observer agreement was poor-moderate (Intraclass Correlation Coefficient (ICC): 0.378, 95% CI 0.210-0.579)., Conclusion: This study showed that quantification of bowel perfusion is a feasible method to differentiate between different perfusion patterns. In addition, the poor-moderate inter-observer agreement of the subjective interpretation of the fluorescence signal between surgeons emphasizes the need for objective quantification., (© 2023. The Author(s).)

As for many solid cancers, laryngeal cancer is treated surgically, and adequate resection margins are critical for survival. Raman spectroscopy has the capacity to accurately differentiate between cancer and non-cancerous tissue based on their molecular composition, which has been proven in previous work. The aim of this study is to investigate whether Raman spectroscopy can be used to discriminate laryngeal cancer from surrounding non-cancerous tissue. Patients surgically treated for laryngeal cancer were included. Raman mapping experiments were performed ex vivo on resection specimens and correlated to histopathology. Water concentration analysis and CH-stretching region analysis were performed in the high wavenumber range of 2500-4000 cm -1 . Thirty-four mapping experiments on 22 resection specimens were used for analysis. Both laryngeal cancer and all non-cancerous tissue structures showed high water concentrations of around 75%. Discriminative information was only found to be present in the CH-stretching region of the Raman spectra of the larynx (discriminative power of 0.87). High wavenumber region Raman spectroscopy can discriminate laryngeal cancer from non-cancerous tissue structures. Contrary to the findings for oral cavity cancer, water concentration is not a discriminating factor for laryngeal cancer., (© 2023. The Author(s).)

Patients with oral cavity cancer are almost always treated with surgery. The goal is to remove the tumor with a margin of more than 5 mm of surrounding healthy tissue. Unfortunately, this is only achieved in about 15% to 26% of cases. Intraoperative assessment of tumor resection margins (IOARM) can dramatically improve surgical results. However, current methods are laborious, subjective, and logistically demanding. This hinders broad adoption of IOARM, to the detriment of patients. Here we present the development and validation of a high-wavenumber Raman spectroscopic technology, for quick and objective intraoperative measurement of resection margins on fresh specimens. It employs a thin fiber-optic needle probe, which is inserted into the tissue, to measure the distance between a resection surface and the tumor. A tissue classification model was developed to discriminate oral cavity squamous cell carcinoma (OCSCC) from healthy oral tissue, with a sensitivity of 0.85 and a specificity of 0.92. The tissue classification model was then used to develop a margin length prediction model, showing a mean difference between margin length predicted by Raman spectroscopy and histopathology of -0.17 mm.

Background: Radiotherapy (RT) is the standard of care for most advanced head and neck squamous cell carcinoma (HNSCC) and results in an unfavorable 5-year overall survival of 40%. Despite strong biological rationale, combining RT with immune checkpoint inhibitors does not result in a survival benefit. Our hypothesis is that the combination of these individually effective treatments fails because of radiation-induced immunosuppression and lymphodepletion. By integrating modern radiobiology and innovative radiotherapy concepts, the patient's immune system could be maximally retained by (1) increasing the dose per fraction so that the total dose and number of fractions can be reduced (HYpofractionation), (2) redistributing the radiation dose towards a higher peak dose within the tumor center and a lowered elective lymphatic field dose (Dose-redistribution), and (3) using RAdiotherapy with protons instead of photons (HYDRA)., Methods: The primary aim of this multicenter study is to determine the safety of HYDRA proton- and photon radiotherapy by conducting two parallel phase I trials. Both HYDRA arms are randomized with the standard of care for longitudinal immune profiling. There will be a specific focus on actionable immune targets and their temporal patterns that can be tested in future hypofractionated immunoradiotherapy trials. The HYDRA dose prescriptions (in 20 fractions) are 40 Gy elective dose and 55 Gy simultaneous integrated boost on the clinical target volume with a 59 Gy focal boost on the tumor center. A total of 100 patients (25 per treatment group) will be recruited, and the final analysis will be performed one year after the last patient has been included., Discussion: In the context of HNSCC, hypofractionation has historically only been reserved for small tumors out of fear for late normal tissue toxicity. To date, hypofractionated radiotherapy may also be safe for larger tumors, as both the radiation dose and volume can be reduced by the combination of advanced imaging for better target definition, novel accelerated repopulation models and high-precision radiation treatment planning and dose delivery. HYDRA's expected immune-sparing effect may lead to improved outcomes by allowing for future effective combination treatment with immunotherapy., Trial Registration: The trial is registered at ClinicalTrials.gov; NCT05364411 (registered on May 6th, 2022)., (© 2023. The Author(s).)

Near-infrared (NIR) fluorescence imaging using exogenous fluorescent agents provides whole-field images in real-time to assist the surgeon in the excision of a tumor. Although the method has high sensitivity, the specificity can sometimes be lower than expected. Raman spectroscopy can detect tumors with high specificity. Therefore, a combination of both techniques can be advantageous. A complication that must be addressed is that the NIR spectral region is favored by both techniques for ( in vivo ) tissue analysis. When fluorescence and Raman emissions spectrally overlap, it becomes challenging or impossible to detect the Raman signal. In this paper, by avoiding this overlap, we describe a Raman spectroscopy setup capable of recording high-quality Raman spectra from tissue containing NIR exogenous fluorescent agents. We identify an optimal wavelength interval (900-915 nm) for Raman excitation, which avoids both excitation of fluorescent dyes and Raman signal self-absorption by the tissue. In this way, Raman spectroscopy can be combined with the currently most-used NIR fluorescent dyes. This combined novel setup could pave the way for clinical trials benefiting from both fluorescence imaging and Raman spectroscopy to avoid positive margins in cancer surgery.

Background: Head and neck squamous cell carcinoma (HNSCC) displays a large heterogeneity in treatment response, and consequently in patient prognosis. Despite extensive efforts, no clinically validated model is available to predict tumor response. Here we describe a functional test for predicting tumor response to radiation and chemotherapy on the level of the individual patient., Methods: Resection material of 17 primary HNSCC patients was cultured ex vivo, irradiated or cisplatin-treated, after which the effect on tumor cell vitality was analyzed several days after treatment., Results: Ionizing radiation (IR) affected tumor cell growth and viability with a clear dose-response relationship, and marked heterogeneity between tumors was observed. After a single dose of 5Gy, proliferation in IR-sensitive tumors dropped below 30% of the untreated level, while IR-resistant tumors maintained at least 60% of proliferation. IR-sensitive tumors showed on average a twofold increase in apoptosis, as well as an increased number and size of DNA damage foci after treatment. No differences in the homologous recombination (HR) proficiency between IR-sensitive and -resistant tumors were detected. Cisplatin caused a decrease in proliferation, as well as induction of apoptosis, again with marked variation between the samples., Conclusions: Our functional ex vivo assay discriminated between IR-sensitive and IR-resistant HNSCC tumors, and may also be suitable for predicting response to cisplatin. Its predictive value is currently under investigation in a prospective clinical study.

Background: Considerable interindividual variation in meniscal microvascularization has been reported. The purpose of this review was to identify which patient characteristics affect meniscal microvascularization and provide a structured overview of angiogenic therapies that influence meniscal neovascularization., Methods: A systematic literature search was undertaken using PubMed, Embase, Web of Science, Cochrane library and Emcare from inception to November 2021. Studies reporting on (1) Patient characteristics that affect meniscal microvascularization, or (2) Therapies that induce neovascularization in meniscal tissue were included. Studies were graded in quality using the Anatomical Quality Assessment (AQUA) tool. The study was registered with PROSPERO(ID:CRD42021242479)., Results: Thirteen studies reported on patient characteristics and eleven on angiogenic therapies. The influence of Age, Degenerative knee, Gender, and Race was reported. Age is the most studied factor. The entire meniscus is vascularized around birth. With increasing age, vascularization decreases from the inner to the peripheral margin. Around 11 years, blood vessels are primarily located in the peripheral third of the menisci. There seems to be a further decrease in vascularization with increasing age in adults, yet conflicting literature exists. Degenerative changes of the knee also seem to influence meniscal vascularization, but evidence is limited. Angiogenic therapies to improve meniscal vascularization have only been studied in preclinical setting. The use of synovial flap transplantation, stem cell therapy, vascular endothelial growth factor, and angiogenin has shown promising results., Conclusion: To decrease failure rates of meniscal repair, a better understanding of patient-specific vascular anatomy is essential. Translational clinical research is needed to investigate the clinical value of angiogenic therapies., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Purpose: The purpose of this study was to assess whether the vascularisation of the meniscus could be visualised intra-operatively using near-infrared fluorescence (NIRF) imaging with indocyanine green (ICG) in patients undergoing total knee arthroplasty (TKA)., Methods: The anterior horn (i.e., Cooper classification: zones C and D) of the meniscus that was least affected (i.e., least degenerative) was removed during TKA surgery in ten patients to obtain a cross section of the inside of the meniscus. Thereafter, 10 mg of ICG was injected intravenously, and vascularisation of the cross section of the meniscus was assessed using the Quest spectrum NIRF camera system. We calculated the percentage of patients in whom vascularisation was observed intra-operatively using NIRF imaging compared to immunohistochemistry., Results: Meniscal vascularisation using NIRF imaging was observed in six out of eight (75%) patients in whom vascularisation was demonstrated with immunohistochemistry. The median extent of vascularisation was 13% (interquartile range (IQR) 3-28%) using NIRF imaging and 15% (IQR 11-23%) using immunohistochemistry., Conclusion: This study shows the potential of NIRF imaging to visualise vascularisation of the meniscus, as vascularisation was observed in six out of eight patients with histologically proven meniscal vascularisation., Level of Evidence: IV., (© 2021. The Author(s).)

Background: Colorectal cancer is the fourth most diagnosed malignancy worldwide and surgery is one of the cornerstones of the treatment strategy. Near-infrared (NIR) fluorescence imaging is a new and upcoming technique, which uses an NIR fluorescent agent combined with a specialised camera that can detect light in the NIR range. It aims for more precise surgery with improved oncological outcomes and a reduction in complications by improving discrimination between different structures., Methods: A systematic search was conducted in the Embase, Medline and Cochrane databases with search terms corresponding to 'fluorescence-guided surgery', 'colorectal surgery', and 'colorectal cancer' to identify all relevant trials., Results: The following clinical applications of fluorescence guided surgery for colorectal cancer were identified and discussed: (1) tumour imaging, (2) sentinel lymph node imaging, (3) imaging of distant metastases, (4) imaging of vital structures, (5) imaging of perfusion. Both experimental and FDA/EMA approved fluorescent agents are debated. Furthermore, promising future modalities are discussed., Conclusion: Fluorescence-guided surgery for colorectal cancer is a rapidly evolving field. The first studies show additional value of this technique regarding change in surgical management. Future trials should focus on patient related outcomes such as complication rates, disease free survival, and overall survival., Competing Interests: Declaration of Competing Interest Alexander L Vahrmeijer and Cornelis Verhoef are local principal investigators of several industry driven studies initiated by Surgimab (Montpellier, France) using SGM-101 without any personal financial interests. Surgimab was not involved in the process of creating this manuscript. The other authors declare no competing interests. Study concepts: S.K., D.H. Study design: H.G., R.M., S.K., D.H. Data acquisition: H.G., R.M. Quality control of data and algorithms: H.G., R.M. Data analysis and interpretation: na Statistical analysis: na Manuscript preparation: H.G., R.M., S.K., D.H. Manuscript editing: H.G., R.M., S.K., D.H. Manuscript review: L.L., C.V., J.B., A.V., M.H., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Photodynamic therapy (PDT) has a great therapeutic potential because it induces local cellular cytotoxicity upon application of a laser light that excites a photosensitizer, leading to toxic reactive oxygen species. Nevertheless, PDT still is underutilized in the clinic, mostly because of damage induced to normal surrounding tissues. Efforts have been made to improve the specificity. Nanobody-targeted PDT is one of such approaches, in which the variable domain of heavy-chain antibodies, i.e., nanobodies, are used to target photosensitizers selectively to cancer cells. In vitro studies are certainly very valuable to evaluate the therapeutic potential of PDT approaches, but many aspects such as bio-distribution of the photosensitizers, penetration through tissues, and clearance are not taken into account. In vivo studies are therefore essential to assess the influence of such factors, in order to gain more insights into the therapeutic potential of a treatment under development. This chapter describes the development of an orthotopic model of head and neck cancer, to which nanobody-targeted PDT is applied, and the therapeutic potential is assessed by immunohistochemistry one day after PDT., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)

Introduction: A challenge in the treatment of patients with head and neck cancer is the management of occult cervical lymph node (LN) metastases. Single-fiber reflectance (SFR) spectroscopy has the potential to detect physiological tissue changes that occur in a positive LN. This pilot study aimed to investigate whether SFR spectroscopy could serve as an alternative or additional technique to detect cervical lymph node metastases., Materials and Methods: We performed intraoperative SFR spectroscopy measurements of LNs with and without malignancies. We analyzed if physiological and scattering parameters were significantly altered in positive LNs., Results: Nine patients with a total of nineteen LNs were included. Three parameters, blood volume fraction (BVF), microvascular saturation (StO 2 ), and Rayleigh amplitude, were significantly lower in positive LNs. They were combined into one optical parameter 'delta', using discriminant analysis. Delta was significantly decreased in positive LNs, p = 0,0006. It had a high diagnostic accuracy where the sensitivity, specificity, PPV, and NPV were 90,0%, 88.9%, 90,0%, and 88.9%, respectively. The area under the ROC curve was 96.7% (95% confidence interval 89.7-100.0%)., Conclusion: This proof of principle study is a first step in the development of an SFR spectroscopy technique to detect LN metastases in real time. A next step towards this goal is replicating these results in LNs with smaller metastases and in a larger cohort of patients. This future study will combine SFR spectroscopy with fine-needle aspiration, using the same needle, to perform preoperative in vivo measurements., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Patients with head and neck squamous cell carcinoma (HNSCC) have an increased risk of developing esophageal second primary tumors (ESPTs). We aimed to determine the incidence, stage, and outcome of synchronous ESPTs in patients with HNSCC in a Western population. We performed a prospective, observational, and cohort study. Patients diagnosed with HNSCC in the oropharynx, hypopharynx, any other sub-location in combination with alcohol abuse, or patients with two synchronous HNSCCs, between February 2019 and February 2020 underwent screening esophagogastroduodenoscopy (EGD). ESPT was defined as presence of esophageal squamous cell carcinoma (ESCC) or high grade dysplasia (HGD). Eighty-five patients were included. A lesion suspected for ESPT was detected in 14 of 85 patients, which was pathologically confirmed in five patients (1 ESCC and 4 HGD). The radiotherapy field was extended to the esophagus in two of five patients, HGD was treated with endoscopic resection in three of five patients. None of the ESPTs were detected on MRI and/or CT-scan prior to EGD. Of the remaining nine patients, three had low grade dysplasia on histology whereas the other six patients had benign lesions. Incidence of synchronous ESPT was 5.9% in our cohort of HNSCC patients. All ESPTs were diagnosed at an early stage and treated with curative intent. We recommend that screening for synchronous ESPTs should be considered in a selected group of patients with HNSCC., (© The Author(s) 2021. Published by Oxford University Press on behalf of International Society for Diseases of the Esophagus. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Objectives: To define metastatic categories based on their prognostic significance. We hypothesized that oligometastasis in patients with head and neck squamous cell carcinoma (HNSCC) is associated with better post-distant metastasis disease specific survival (post-DM DSS) compared to patients with polymetastasis. Furthermore, the impact on survival of synchronous versus metachronous distant metastasis (DM) occurrence was assessed., Materials and Methods: Retrospective cohort study in which patients with DM were stratified into three groups: oligometastasis (maximum of 3 metastatic foci in ≤2 anatomic sites), explosive metastasis (≥4 metastatic foci at one anatomic site) and explosive-disseminating metastasis (spread to ≥3 anatomic sites or >3 metastatic foci in 2 anatomic sites). In addition, patients were divided into synchronous versus metachronous DM., Results: Between January 1, 2006 and December 31, 2013, a total of 2687 patients with HNSCC were identified, of which 324 patients developed DM. In this group, 115 (35.5%) patients had oligometastasis, 64 (19.8%) patients had explosive metastasis and 145 (44.8%) patients had explosive-disseminating metastasis. Their median post-DM DSS were 4.7 months, 4.1 months and 1.7 months respectively (p < .001). Synchronous DM was associated with more favorable survival rates in univariable and multivariable analyses than metachronous DM with recurrence of the index tumor (6-month post-DM DSS probability of 0.51 vs 0.17, p < .001)., Conclusion: Oligometastasis in HNSCC signifies a better prognosis than a polymetastatic pattern. Metachronous DM occurrence with recurrence of the primary index tumor is associated with an unfavorable prognosis., (Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The goal of head and neck oncological surgery is complete tumor resection with adequate resection margins while preserving acceptable function and appearance. For oral cavity squamous cell carcinoma (OCSCC), different studies showed that only 15%-26% of all resections are adequate. A major reason for the low number of adequate resections is the lack of information during surgery; the margin status is only available after the final histopathologic assessment, days after surgery. The surgeons and pathologists at the Erasmus MC University Medical Center in Rotterdam started the implementation of specimen-driven intraoperative assessment of resection margins (IOARM) in 2013, which became the standard of care in 2015. This method enables the surgeon to turn an inadequate resection into an adequate resection by performing an additional resection during the initial surgery. Intraoperative assessment is supported by a relocation method procedure that allows accurate identification of inadequate margins (found on the specimen) in the wound bed. The implementation of this protocol resulted in an improvement of adequate resections from 15%-40%. However, the specimen-driven IOARM is not widely adopted because grossing fresh tissue is counter-intuitive for pathologists. The fear exists that grossing fresh tissue will deteriorate the anatomical orientation, shape, and size of the specimen and therefore will affect the final histopathologic assessment. These possible negative effects are countered by the described protocol. Here, the protocol for specimen-driven IOARM is presented in detail, as performed at the institute.

Fluorescence-guided surgery is an intraoperative optical imaging method that provides surgeons with real-time guidance for the delineation of tumours. Currently, in phase 1 and 2 clinical trials, evaluation of fluorescence-guided surgery is primarily focused on its diagnostic performance, although the corresponding outcome variables do not inform about the added clinical benefit of fluorescence-guided surgery and are challenging to assess objectively. Nonetheless, the effect of fluorescence-guided surgery on intraoperative decision making is the most objective outcome measurement to assess the clinical value of this imaging method. In this Review, we explore the study designs of existing trials of fluorescence-guided surgery that allow us to extract information on potential changes in intraoperative decision making, such as additional or more conservative resections. On the basis of this analysis, we offer recommendations on how to report changes in intraoperative decision making that result from fluorescence imaging, which is of utmost importance for the widespread clinical implementation of fluorescence-guided surgery., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Laryngeal cancer is a prevalent head and neck malignancy, with poor prognosis and low survival rates for patients with advanced disease. Treatment consists of unimodal therapy through surgery or radiotherapy in early staged tumors, while advanced stage tumors are generally treated with multimodal chemoradiotherapy or (total) laryngectomy followed by radiotherapy. Still, the recurrence rate for advanced laryngeal cancer is between 25 and 50%. In order to improve surgical resection of laryngeal cancer and reduce local recurrence rates, various intraoperative optical imaging techniques have been investigated. In this systematic review, we identify these technologies, evaluating the current state and future directions of optical imaging for this indication. Narrow-band imaging (NBI) and autofluorescence (AF) are established tools for early detection of laryngeal cancer. Nonetheless, their intraoperative utility is limited by an intrinsic inability to image beyond the (sub-)mucosa. Likewise, contact endoscopy (CE) and optical coherence tomography (OCT) are technically cumbersome and only useful for mucosal margin assessment. Research on fluorescence imaging (FLI) for this application is sparse, dealing solely with nonspecific fluorescent agents. Evidently, the imaging modalities that have been investigated thus far are generally unsuitable for deep margin assessment. We discuss two optical imaging techniques that can overcome these limitations and suggest how they can be used to achieve adequate margins in laryngeal cancer at all stages.

Objectives: To compare the impact on survival and late radiation toxicity of patients with squamous cell carcinoma of unknown primary who were treated with ipsilateral neck dissection and ipsilateral postoperative radiotherapy (PORT) and patients treated with ipsilateral neck dissection and PORT to both sides of the neck plus the pharyngeal axis., Methods: Retrospective cohort study performed at the Erasmus University Medical Center in which 78 patients with squamous cell carcinoma of unknown primary were identified. Thirty-nine patients received PORT to both sides of the neck plus the pharyngeal axis (BILAX) and another 39 patients were irradiated on the ipsilateral neck (IPSI) only. The endpoints of the present study were 3-year overall survival (OS), 3-year disease-free survival (DFS), and overall late radiation toxicity., Results: The 3-year OS rate of the entire group of 78 patients was 74.2% and the 3-year DFS rate was 72.7%. The 3-year OS rates for the IPSI and the BILAX groups were 74.4% and 74.1%, respectively ( P = .654). The most common late radiation toxicity experienced was xerostomia (64.8%), which was significantly more often seen in the BILAX group than in the IPSI group (83.8% and 44.1%, respectively, P = .001). Overall late radiation toxicity was significantly higher in the BILAX group ( P = .003)., Conclusion: There was no significant difference in OS and DFS rates between the IPSI and the BILAX group. Late radiation toxicity was significantly higher in the BILAX group., Level of Evidence: Level 2b: Individual retrospective cohort study., Competing Interests: 6The authors declare no potential conflict of interest., (© 2021 The Authors. Laryngoscope Investigative Otolaryngology published by Wiley Periodicals LLC. on behalf of The Triological Society.)

Objective: The depth of invasion (DOI) is considered an independent risk factor for occult lymph node metastasis in oral cavity squamous cell carcinoma (OCSCC). It is used to decide whether an elective neck dissection (END) is indicated in the case of a clinically negative neck for early stage carcinoma (pT1/pT2). However, there is no consensus on the cut-off value of the DOI for performing an END. The aim of this study was to determine a cut-off value for clinical decision making on END, by assessing the association of the DOI and the risk of occult lymph node metastasis in early OCSCC., Methods: A retrospective cohort study was conducted at the Erasmus MC, University Medical Centre Rotterdam, The Netherlands. Patients surgically treated for pT1/pT2 OCSCC between 2006 and 2012 were included. For all cases, the DOI was measured according to the 8 th edition of the American Joint Committee on Cancer guideline. Patient characteristics, tumor characteristics (pTN, differentiation grade, perineural invasion, and lymphovascular invasion), treatment modality (END or watchful waiting), and 5-year follow-up (local recurrence, regional recurrence, and distant metastasis) were obtained from patient files., Results: A total of 222 patients were included, 117 pT1 and 105 pT2. Occult lymph node metastasis was found in 39 of the 166 patients who received END. Univariate logistic regression analysis showed DOI to be a significant predictor for occult lymph node metastasis (odds ratio (OR) = 1.3 per mm DOI; 95% CI: 1.1-1.5, p = 0.001). At a DOI of 4.3 mm the risk of occult lymph node metastasis was >20% (all subsites combined)., Conclusion: The DOI is a significant predictor for occult lymph node metastasis in early stage oral carcinoma. A NPV of 81% was found at a DOI cut-off value of 4 mm. Therefore, an END should be performed if the DOI is >4 mm., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Aaboubout, van der Toom, de Ridder, De Herdt, van der Steen, van Lanschot, Barroso, Nunes Soares, ten Hove, Mast, Smits, Sewnaik, Monserez, Keereweer, Caspers, Baatenburg de Jong, Bakker Schut, Puppels, Hardillo and Koljenović.)

Surgery is the mainstay of treatment for localized soft tissue sarcomas (STS). The curative treatment highly depends on complete tumor resection, as positive margins are associated with local recurrence (LR) and prognosis. However, determining the tumor margin during surgery is challenging. Real-time tumor-specific imaging can facilitate complete resection by visualizing tumor tissue during surgery. Unfortunately, STS specific tracers are presently not clinically available. In this review, STS-associated cell surface-expressed biomarkers, which are currently already clinically targeted with monoclonal antibodies for therapeutic purposes, are evaluated for their use in near-infrared fluorescence (NIRF) imaging of STS. Clinically targeted biomarkers in STS were extracted from clinical trial registers and a PubMed search was performed. Data on biomarker characteristics, sample size, percentage of biomarker-positive STS samples, pattern of biomarker expression, biomarker internalization features, and previous applications of the biomarker in imaging were extracted. The biomarkers were ranked utilizing a previously described scoring system. Eleven cell surface-expressed biomarkers were identified from which 7 were selected as potential biomarkers for NIRF imaging: TEM1, VEGFR-1, EGFR, VEGFR-2, IGF-1R, PDGFRα, and CD40. Promising biomarkers in common and aggressive STS subtypes are TEM1 for myxofibrosarcoma, TEM1, and PDGFRα for undifferentiated soft tissue sarcoma and EGFR for synovial sarcoma.

Background: Inadequate resection margins in oral cavity squamous cell carcinoma have an adverse effect on patient outcome. Intraoperative assessment provides immediate feedback enabling the surgeon to achieve adequate resection margins. The goal of this study was to evaluate the value of specimen-driven intraoperative assessment by comparing the margin status in the period before and the period after the introduction of specimen-driven assessment as a standard of care (period 2010-2012 vs period 2013-2017)., Methods: A cohort of patients surgically treated for oral squamous cell carcinoma at the Erasmus MC Cancer Institute, Rotterdam, between 2010-2012 was studied retrospectively and compared to results of a prospectively collected cohort between 2013-2017. The frequency, type and results of intraoperative assessment of resection margins were analyzed., Results: One hundred seventy-four patients were included from 2010-2012, 241 patients were included from 2013-2017. An increase in the frequency of specimen-driven assessment was seen between the two periods, from 5% in 2010-2012 to 34% in 2013-2017. When performing specimen-driven assessment, 16% tumor-positive resection margins were found in 2013-2017, compared to 43% tumor-positive resection margins overall in 2010-2012. We found a significant reduction of inadequate resection margins for specimen-driven intraoperative assessment (p < 0.001). Also, tumor recurrence significantly decreased, and disease-specific survival improved when performing specimen-driven intraoperative assessment., Conclusions: Specimen-driven intraoperative assessment improves resection margins and consequently, the outcome of oral cancer patients. We advocate this method as standard of care., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2020 Smits, van Lanschot, Aaboubout, de Ridder, Hegt, Barroso, Meeuwis, Sewnaik, Hardillo, Monserez, Keereweer, Mast, Hove, Bakker Schut, Baatenburg de Jong, Puppels and Koljenović.)

Objectives: Depth of invasion (DOI) is the most important predictor for lymph node metastasis (LNM) in early stage (T1-T2) oral cancer. The aim of this study is to validate the cut-off value of 4 mm on which the decision to perform an Elective Neck Dissection (END) is made., Materials and Methods: We performed a retrospective study in patients with pathologically proven early stage oral cavity squamous cell carcinoma (OCSCC) without clinical or radiological signs of LNM, who were treated between 2013 and 2018. An END was performed when DOI was ≥ 4 mm and a watchful waiting protocol was applied in patients with DOI < 4 mm., Results: Three hundred patients were included. END was performed in 77% of patients with DOI ≥ 4 mm, of which 36% had occult LNM (pN+). Patients in the watchful waiting group (48%) developed a regional recurrence in 5.2% for DOI < 4 mm and 24.1% for DOI ≥ 4 mm. For DOI ≥ 4 mm, regional recurrence free survival was higher for patients who were treated with END compared to watchful waiting (p = 0.002). A Receiver-Operator-Curve -analysis showed that a DOI cut-off value of 4.0 mm was the optimal threshold for the prediction of occult LNM (95.1% sensitivity, 52.9% specificity)., Conclusion: A DOI of ≥ 4 mm is an accurate cut-off value for performing an END in early stage OCSCC. END results in higher survival rates and lower regional recurrence rates in patients with DOI ≥ 4 mm., (Copyright © 2020. Published by Elsevier Ltd.)

Objectives: Tumour-positive resection margins are a major problem during oral cancer surgery. gGlu-HMRG is a tracer that becomes fluorescent upon activation by gamma-glutamyltranspeptidase (GGT). This study aims to investigate the combination of gGlu-HMRG and a clinical fluorescence imaging system for the detection of tumour-positive resection margins., Materials and Methods: The preclinical Maestro and clinical Artemis imaging systems were compared in vitro and ex vivo with cultured human head and neck cancer cells (OSC19, GGT-positive; and FaDu, GGT negative) and tumour-bearing nude mice. Subsequently, frozen sections of normal and oral cancer tissues were ex vivo sprayed with gGlu-HMRG to determine the sensitivity and specificity. Finally, resection margins of patients with suspected oral cancer were ex vivo sprayed with gGlu-HMRG to detect tumour-positive resection margins., Results: Both systems could be used to detect gGlu-HMRG activation in vitro and ex vivo in GGT positive cancer cells. Sensitivity and specificity of gGlu-HMRG and the Artemis on frozen tissue samples was 80% and 87%, respectively. Seven patients undergoing surgery for suspected oral cancer were included. In three patients fluorescence was observed at the resection margin. Those margins were either tumour-positive or within 1 mm of tumour. The margins of the other patients were clear (≥8 mm)., Conclusion: This study demonstrates the feasibility to detect tumour-positive resection margins with gGlu-HMRG and a clinical fluorescence imaging system. Applying this technique would enable intraoperative screening of the entire resection margin and allow direct re-resection in case of tumour-positivity., (Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Navigation with fluorescence guidance has emerged in the last decade as a promising strategy to improve the efficacy of oncologic surgery. To achieve routine clinical use, the onus is on the surgical community to objectively assess the value of this technique. This assessment may facilitate both Food and Drug Administration approval of new optical imaging agents and reimbursement for the imaging procedures. It is critical to characterize fluorescence-guided procedural benefits over existing practices and to elucidate both the costs and the safety risks. This report is the result of a meeting of the International Society of Image Guided Surgery (www.isigs.org) on February 6, 2015, in Miami, Florida, and reflects a consensus of the participants' opinions. Our objective was to critically evaluate the imaging platform technology and optical imaging agents and to make recommendations for successful clinical trial development of this highly promising approach in oncologic surgery., (© 2016 by the Society of Nuclear Medicine and Molecular Imaging, Inc.)

Purpose: Antiangiogenic therapy, mostly targeting VEGF, has been applied in cancer patients for the last decade. However, resistance to anti-VEGF therapy and/or no significant benefit as monotherapeutic agent is often observed. Therefore, new antiangiogenic strategies are needed. In the current study, we investigated the therapeutic effect of interfering with the bone morphogenetic protein (BMP)9/activin receptor-like kinase (ALK)1 signaling pathway by using an ALK1-Fc ligand trap., Experimental Design: We analyzed the potential antiangiogenic and antitumor effects of ALK1-Fc protein as monotherapy and in combination with chemotherapy in vivo in mouse models of melanoma, head and neck cancer, and invasive lobular breast carcinomas. ALK1-Fc sequesters BMP9 and 10 and prevents binding of these ligands to endothelial ALK1, which regulates angiogenesis., Results: Treatment of mice with ALK1-Fc strongly decreased the tumors' microvascular density in the three different mouse cancer models. However, this effect was not accompanied by a reduction in tumor volume. An immunohistochemical analysis of the tumor samples revealed that ALK1-Fc treatment increased the pericyte coverage of the remaining tumor vessels and decreased the hypoxia within the tumor. Next, we observed that combining ALK1-Fc with cisplatin inhibited tumor growth in the breast and head and neck cancer models more efficiently than chemotherapy alone., Conclusions: The addition of ALK1-Fc to the cisplatin treatment was able to enhance the cytotoxic effect of the chemotherapy. Our results provide strong rationale to explore combined targeting of ALK1 with chemotherapy in a clinical setting, especially in the ongoing phase II clinical trials with ALK1-Fc., (©2015 American Association for Cancer Research.)

Quantification of tumor necrosis in cancer patients is of diagnostic value as the amount of necrosis is correlated with disease prognosis and it could also be used to predict early efficacy of anti-cancer treatments. In the present study, we identified two near infrared fluorescent (NIRF) carboxylated cyanines, HQ5 and IRDye 800CW (800CW), which possess strong necrosis avidity. In vitro studies showed that both dyes selectively bind to cytoplasmic proteins of dead cells that have lost membrane integrity. Affinity for cytoplasmic proteins was confirmed using quantitative structure activity relations modeling. In vivo results, using NIRF and optoacoustic imaging, confirmed the necrosis avid properties of HQ5 and 800CW in a mouse 4T1 breast cancer tumor model of spontaneous necrosis. Finally, in a mouse EL4 lymphoma tumor model, already 24 h post chemotherapy, a significant increase in 800CW fluorescence intensity was observed in treated compared to untreated tumors. In conclusion, we show, for the first time, that the NIRF carboxylated cyanines HQ5 and 800CW possess strong necrosis avid properties in vitro and in vivo. When translated to the clinic, these dyes may be used for diagnostic or prognostic purposes and for monitoring in vivo tumor response early after the start of treatment.

Purpose: Hyperbaric oxygen therapy (HBOT) is used in the treatment of radiation-induced tissue injury but its effect on (residual) tumor tissue is indistinct and therefore investigated in this study., Procedures: Orthotopic FaDu tumors were established in mice, and the response of the (irradiated) tumors to HBOT was monitored by bioluminescence imaging. Near infrared fluorescence imaging using AngioSense750 and Hypoxisense680 was applied to detect tumor vascular permeability and hypoxia., Results: HBOT treatment resulted in accelerated growth of non-irradiated tumors, but mouse survival was improved. Tumor vascular leakiness and hypoxia were enhanced after HBOT, whereas histological characteristics, epithelial-to-mesenchymal transition markers, and metastatic incidence were not influenced., Conclusions: Squamous cell carcinoma responds to HBOT with respect to tumor growth, vascular permeability, and hypoxia, which may have implications for its use in cancer patients. The ability to longitudinally analyze tumor characteristics highlights the versatility and potential of optical imaging methods in oncological research.

Objectives: To date, over 200 000 cochlear implants (CIs) have been implanted worldwide and the incidence is still increasing. We present a case of traumatic displacement of CI magnet to raise awareness about this complication and to highlight the need for vigilance during surgery as well as for proper counseling., Methods: The clinical presentation of a 1.5-year-old boy with a traumatic displacement of a CI magnet was presented and the literature was reviewed for this rare complication., Results: After minor head injury, the sound processor could no longer connect to the CI. X-ray imaging demonstrated displacement of the CI magnet. During revision surgery, the magnet was replaced by a new magnet in the silicon holding cap. Intraoperative impedance measurements were normal and the CI was successfully activated 4 weeks postoperatively., Conclusions: Clinicians and patients should be aware of the risk of displacement of the CI magnet after (minor) head injury. Young boys tend to have a higher risk for this complication.

Cancer patients could benefit from a surgical procedure that helps the surgeon to determine adequate tumor resection margins. Systemic injection of tumor-specific fluorescence agents with subsequent intraoperative optical imaging can guide the surgeon in this process. However, tumor heterogeneity hampers tumor-specific targeting. In addition, determination of adequate resection margins can be very challenging due to invasive tumor strands that are difficult to resolve and because of the confounding effect of variations in tissue optical properties in the surgical margin. We provide an overview of the "classic approach" of imaging tumor-specific targets or tumor-associated pathophysiological processes, and explain the limitations of these targeting strategies. It is proposed that problems of tumor heterogeneity can theoretically be circumvented by shifting focus of tumor targeting towards the follicle-stimulating hormone receptor (FSHR). Furthermore, we discuss why objective determination of resection margins is required to improve resection of the invasive strands, a goal that may be achieved by targeting the FSHR. When invasive strands would nevertheless extend beyond such a standardized resection margin, we suggest that adjuvant photodynamic therapy would be a very suitable therapeutic regimen. Finally, we describe how point optical spectroscopy can be used to scrutinize suspect tissue that is difficult to differentiate from normal tissue by measuring the local tissue optical properties to recover a local intrinsic fluorescence measurement.

Optical image-guided cancer surgery is a promising technique to adequately determine tumor margins by tumor-specific targeting, potentially resulting in complete resection of tumor tissue with improved survival. However, identification of the photons coming from the fluorescent contrast agent is complicated by autofluorescence, optical tissue properties, and accurate fluorescent targeting agents and imaging systems. All these factors have an important influence on the image that is presented to the surgeon. Considering the clinical consequences at stake, it is a prerequisite to answer the questions that are essential for the surgeon. What is optical image-guided surgery and how can it improve patient care? What should the oncologic surgeon know about the fundamental principles of optical imaging to understand which conclusions can be drawn from the images? And how do the limitations influence clinical decision making? This article discusses these questions and provides a clear overview of the basic principles and practical applications. Although there are limitations to the intrinsic capacity of the technique, when practical and technical surgical possibilities are considered, optical imaging can be a very powerful intraoperative tool in guiding the future oncologic surgeon toward radical resection and optimal clinical results.

Objectives: Elective neck dissection is frequently performed during surgery in head and neck cancer patients. The sentinel lymph node (SLN) procedure can prevent the morbidity of a neck dissection and improve lymph node staging by fine pathology. Near-infrared (NIR) fluorescence imaging is a promising technique to identify the sentinel lymph node (SLN) intraoperatively. This feasibility study explored the use of indocyanine green adsorbed to human serum albumin (ICG:HSA) for SLN mapping in head and neck cancer patients., Materials and Methods: A total of 10 consecutive patients with oral cavity or oropharyngeal cancer and a clinical N0 neck were included. After exposure of the neck, 1.6 mL of ICG:HSA (500 μM) was injected at four quadrants around the tumor. During the neck dissection, levels I-IV were measured for fluorescence using the Mini-FLARE imaging system., Results: In all 10 patients, NIR fluorescence imaging enabled visualization of one or more SLNs. A total of 17 SLNs were identified. The mean contrast between the fluorescent signal of the lymph nodes and of the surrounding tissue was 8.7±6.4. In 3 patients, of which 1 was false-negative, lymph node metastases were found. After administration of ICG:HSA, the average number of fluorescent lymph nodes significantly increased over time (P<0.001)., Conclusion: This study demonstrated feasibility to detect draining lymph nodes in head and neck cancer patients using NIR fluorescence imaging. However, the fluorescent tracer quickly migrated beyond the SLN to higher tier nodes., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Optical imaging is a promising technique to visualize cancer tissue during surgery. In this study, we explored the use of combinations of near-infrared (NIR) fluorescence agents that emit fluorescence signal at different wavelengths and each target specific tumor characteristics. Two combinations of agents (ProSense680 combined with 2DG CW800 and MMPSense680 combined with EGF CW800) were used to detect hypopharyngeal cancer in an animal model. ProSense680 and MMPSense680 detect increased activity of cathepsins and matrix metalloproteinases, respectively. These enzymes are mainly found in the invasive tumor border due to degradation of the extracellular matrix. 2DG CW800 detects tumor cells with high glucose metabolism and EGF CW800 is internalized by the epidermal growth factor receptor of tumor cells. Whole-body imaging revealed clear demarcation of tumor tissue using all four agents. The tumor-to-background ratio (standard deviation, p-value) was 3.69 (0.72, p < 0.001) for ProSense680; 4.26 (1.33, p < 0.001) for MMPSense680; 5.81 (3.59, p = 0.02) for 2DG CW800 and 4.84 (1.56, p < 0.001) for EGF CW800. Fluorescence signal corresponded with histopathology and immunohistochemistry, demonstrating signal of ProSense680 and MMPSense680 in the invasive tumor border, and signal of 2DG CW800 and EGF CW800 in the tumor tissue. In conclusion, we demonstrated the feasibility of dual wavelength tumor detection using different targeting strategies simultaneously in an animal model. Combined targeting at different wavelengths allowed simultaneous imaging of different tumor characteristics. NIR fluorescence optical imaging has the potential to be translated into the clinic in order to improve the complete removal of tumors by real-time image-guided surgery., (Copyright © 2012 UICC.)

Background: In oral cancer surgery, intraoperative optical imaging could help the surgeon to determine adequate tumor-free margins., Methods: Tumor-specific near-infrared fluorescence agents targeting epidermal growth factor receptor (CW800 EGF) or glucose transporter system (CW800 2-DG) were administered to mice with tongue carcinoma and cervical lymph node metastases. Tumor growth was followed by bioluminescence imaging. Fluorescence signals were compared with a control group of healthy animals., Results: Significantly higher fluorescence was found in tongue tumors and cervical lymph node metastases compared with that in control animals. Fluorescence correlated with histopathology. Tumor-to-background ratio of CW800 EGF in the tongue was 13.8 (SD = 6.1) and in the lymph nodes 15.7 (SD = 8.8). For CW800 2-DG, the tumor-to-background ratio in the tongue was 4.6 (SD = 2.1) and in the lymph nodes 33.9 (SD = 18.4)., Conclusions: Optical imaging can be used to detect oral cancer and cervical lymph node metastases and could potentially improve complete surgical resection by real-time image-guided surgery., (Copyright © 2011 Wiley Periodicals, Inc.)

Background and Objectives: Near-infrared (NIR) fluorescence optical imaging is a promising technique to assess the tumor margins during cancer surgery. This technique requires targeting by specific fluorescence agents to differentiate tumor from normal surrounding tissue. We assessed the feasibility of cancer detection using NIR fluorescence agents that target either αvβ3 integrins or the enhanced permeability and retention (EPR) effect in an orthotopic mouse model of oral cancer., Methods: Binding of the integrin-targeted agent to tumor cells was assessed in vitro. Oral cancer was induced in 6 BALB/c nu/nu mice by submucosal inoculation of human OSC19-luc cells into the tongue. Tumor growth was followed with bioluminescence imaging. A combination of agents targeting integrins or EPR effect was injected followed by fluorescence imaging in vivo and ex vivo after resection of the tongues., Results: Oral cancer was clearly demarcated in vitro; in vivo; and on histological analysis with sufficient tumor-to-background ratios of the contrast agents., Conclusion: This study demonstrates the feasibility of optical imaging of oral squamous cell carcinoma based on targeting of αvβ3 integrins and the EPR effect. Once these NIR fluorescence agents become available for clinical testing, optical image-guided surgery could reduce residual disease after oral cancer surgery., (Copyright © 2011 Wiley Periodicals, Inc.)

Chemoradiation (CRT) is a valuable treatment option for advanced hypopharyngeal squamous cell cancer (HSCC). However, long-term toxicity and quality of life (QOL) is scarcely reported. Therefore, efficacy, acute and long-term toxic effects, and long-term QOL of CRT for advanced HSCC were evaluated,using retrospective study and post-treatment quality of life questionnaires. in a tertiary hospital setting. Analysis was performed of 73 patients that had been treated with CRT. Toxicity was rated using the CTCAE score list. QOL questionnaires EORTC QLQ-C30, QLQ-H&N35, and VHI were analyzed. The most common acute toxic effects were dysphagia and mucositis. Dysphagia and xerostomia remained problematic during long-term follow-up. After 3 years, the disease-specific survival was 41%, local disease control was 71%, and regional disease control was 97%. The results indicated that CRT for advanced HSCC is associated with high locoregional control and disease-specific survival. However, significant acute and long-term toxic effects occur, and organ preservation appears not necessarily equivalent to preservation of function and better QOL.

In cancer imaging, many different modalities are used that each have their specific features, leading to the combined use of different techniques for the detection, staging and treatment evaluation of cancer. Optical imaging using near-infrared fluorescence light is a new imaging modality that has recently emerged in the field of cancer imaging. After extensive preclinical research, the first steps of translation to the clinical practice are currently being made. In this article, we discuss the preclinical and clinical results of near-infrared optical imaging for non-invasive detection and classification of tumors, therapy monitoring, sentinel lymph node procedures, and image-guided cancer surgery. Widespread availability of imaging systems and optical contrast agents will enable larger studies on their clinical benefit and can help establish a definitive role in clinical practice.

Conclusions: Although organ preservation can be achieved with chemoradiation protocols for laryngeal or pharyngeal cancers, salvage surgery is accompanied by high complication rates., Objectives: To determine the rate of complications associated with salvage surgery after chemoradiation for laryngeal and pharyngeal cancers., Methods: A multicenter retrospective study was performed of 24 patients treated with total laryngectomy combined with total or partial pharyngectomy between 1995 and 2004 who had previously been treated with chemoradiation. The main outcome measures were early and late complication rates. Quality of life analysis was determined by two questionnaires., Results: The complication rate after salvage surgery was 92% in the direct postoperative period. The most frequent complication was pharyngocutaneous fistula formation. Narrowing of the esophagus and tracheostoma were the most common late sequelae. The quality of life, measured at least 2 years after salvage surgery, showed a social dysfunctioning.

Bioluminescence imaging (BLI) has shown its appeal as a sensitive technique for in vivo whole body optical imaging. However, the development of injectable tumor-specific near-infrared fluorescent (NIRF) probes makes fluorescence imaging (FLI) a promising alternative to BLI in situations where BLI cannot be used or is unwanted (e.g., spontaneous transgenic tumor models, or syngeneic mice to study immune effects).In this study, we addressed the questions whether it is possible to detect tumor progression using FLI with appropriate sensitivity and how FLI correlates with BLI measurements. In addition, we explored the possibility to simultaneously detect multiple tumor characteristics by dual-wavelength FLI (~700 and ~800 nm) in combination with spectral unmixing. Using a luciferase-expressing 4T1-luc2 mouse breast cancer model and combinations of activatable and targeting NIRF probes, we showed that the activatable NIRF probes (ProSense680 and MMPSense680) and the targeting NIRF probes (IRDye 800CW 2-DG and IRDye 800CW EGF) were either activated by or bound to 4T1-luc2 cells. In vivo, we implanted 4T1-luc2 cells orthotopically in nude mice and were able to follow tumor progression longitudinally both by BLI and dual-wavelength FLI. We were able to reveal different probe signals within the tumor, which co-localized with immuno-staining. Moreover, we observed a linear correlation between the internal BLI signals and the FLI signals obtained from the NIRF probes. Finally, we could detect pulmonary metastases both by BLI and FLI and confirmed their presence histologically.Taken together, these data suggest that dual-wavelength FLI is a feasible approach to simultaneously detect different features of one tumor and to follow tumor progression with appropriate specificity and sensitivity. This study may open up new perspectives for the detection of tumors and metastases in various experimental models and could also have clinical applications, such as image-guided surgery.

Objective: To assess the feasibility of optical imaging using activatable near-infrared fluorescence (NIRF) agents to detect oral cancer and cervical lymph node metastasis in vivo., Design: In vivo study., Setting: University medical center., Subjects: Female nude mice aged 4 to 6 weeks., Intervention: Luciferase-expressing OSC-19-luc cells were injected into the tongues of nude mice. A control group of nude mice was injected in the tongue with a physiologic saline solution. Tumor growth was followed by bioluminescence imaging. After 3 weeks, animals were randomly allocated to intravenous administration of 1 of 2 activatable NIRF agents: ProSense680 or MMPSense680. Fluorescence imaging of the mice was performed, and the tumor to background ratio (TBR) was determined on histologic sections of the tongue and cervical lymph nodes after resection at necropsy., Main Outcome Measure: Fluorescence signals., Results: The fluorescence signals in tongue tumor and cervical lymph node metastases were significantly higher than those in control animals. The mean (SD) TBR of ProSense680 in the tongue was 15.8 (8.1) and in the lymph nodes was 11.8 (3.6). For MMPSense680, the mean (SD) TBR in the tongue was 18.6 (9.4) and in the lymph nodes was 10.5 (4.0)., Conclusions: Oral cancer and cervical lymph node metastases can be detected by targeting increased proteolytic activity at the tumor borders using NIRF optical imaging. These NIRF agents could be used for real-time image-guided surgery, which has the potential to improve the complete surgical resection of oral cancer.

In cancer surgery, intra-operative assessment of the tumor-free margin, which is critical for the prognosis of the patient, relies on the visual appearance and palpation of the tumor. Optical imaging techniques provide real-time visualization of the tumor, warranting intra-operative image-guided surgery. Within this field, imaging in the near-infrared light spectrum offers two essential advantages: increased tissue penetration of light and an increased signal-to-background-ratio of contrast agents. In this article, we review the various techniques, contrast agents, and camera systems that are currently used for image-guided surgery. Furthermore, we provide an overview of the wide range of molecular contrast agents targeting specific hallmarks of cancer and we describe perspectives on its future use in cancer surgery.