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.

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.

A clear margin is an important prognostic factor for most solid tumours treated by surgery. Intraoperative fluorescence imaging using exogenous tumour-specific fluorescent agents has shown particular benefit in improving complete resection of tumour tissue. However, signal processing for fluorescence imaging is complex, and fluorescence signal intensity does not always perfectly correlate with tumour location. Raman spectroscopy has the capacity to accurately differentiate between malignant and healthy tissue based on their molecular composition. In Raman spectroscopy, specificity is uniquely high, but signal intensity is weak and Raman measurements are mainly performed in a point-wise manner on microscopic tissue volumes, making whole-field assessment temporally unfeasible. In this review, we describe the state-of-the-art of both optical techniques, paying special attention to the combined intraoperative application of fluorescence imaging and Raman spectroscopy in current clinical research. We demonstrate how these techniques are complementary and address the technical challenges that have traditionally led them to be considered mutually exclusive for clinical implementation. Finally, we present a novel strategy that exploits the optimal characteristics of both modalities to facilitate resection with clear surgical margins.

Background In head and neck oncological surgery the goal is to achieve a complete tumor resection with acceptable remaining function and appearance. For oral cavity squamous cell carcinoma (OCSCC) only 15% of the resections are reported as adequate. Since 2013, we have performed intraoperative assessment of resection margins (IOARM) in our institute, based on palpation and visual inspection of the resected specimens by pathologist and surgeon. This has resulted in an improvement of adequate resection margins from 15% to 50%, underlining the importance of IOARM. However, this method is subjective, labor intensive, and logistically challenging. Objective Our aim is to develop an objective method for fast and reliable IOARM based on Raman spectroscopy (RS). Methods RS is a non-destructive objective optical technique that provides information about the molecular composition of tissues. It can discriminate between healthy tissue and tumors. We developed a prototype Raman instrument employing a fiber-optic needle probe. The fiber-optic needle is driven into the OCSCC specimen, from the resection surface towards the tumor. Based on the Raman spectra collected along the insertion path, the location of the tumor border can be determined. From this the resection margin can be determined. Results First tests of the method show that the instrument accurately predicts the achieved resection margins. Per location the measurement and assessment takes 5 seconds. Conclusions This development signifies an important step towards a fast and objective IOARM. The fast measurement time enables an objective inspection of the margins achieved at a large number of locations of the resection surface. In head and neck oncological surgery the goal is to achieve a complete tumor resection with acceptable remaining function and appearance. For oral cavity squamous cell carcinoma (OCSCC) only 15% of the resections are reported as adequate. Since 2013, we have performed intraoperative assessment of resection margins (IOARM) in our institute, based on palpation and visual inspection of the resected specimens by pathologist and surgeon. This has resulted in an improvement of adequate resection margins from 15% to 50%, underlining the importance of IOARM. However, this method is subjective, labor intensive, and logistically challenging. Our aim is to develop an objective method for fast and reliable IOARM based on Raman spectroscopy (RS). RS is a non-destructive objective optical technique that provides information about the molecular composition of tissues. It can discriminate between healthy tissue and tumors. We developed a prototype Raman instrument employing a fiber-optic needle probe. The fiber-optic needle is driven into the OCSCC specimen, from the resection surface towards the tumor. Based on the Raman spectra collected along the insertion path, the location of the tumor border can be determined. From this the resection margin can be determined. First tests of the method show that the instrument accurately predicts the achieved resection margins. Per location the measurement and assessment takes 5 seconds. This development signifies an important step towards a fast and objective IOARM. The fast measurement time enables an objective inspection of the margins achieved at a large number of locations of the resection surface.

INTRODUCTION:Achieving adequate resection margins during oral cancer surgery is important to improve patient prognosis. Surgeons have the delicate task of achieving an adequate resection and safeguarding satisfactory remaining function and acceptable physical appearance, while relying on visual inspection, palpation, and preoperative imaging. Intraoperative assessment of resection margins (IOARM) is a multidisciplinary effort, which can guide towards adequate resections. Different forms of IOARM are currently used, but it is unknown how accurate these methods are in predicting margin status. Therefore, this review aims to investigate: 1) the IOARM methods currently used during oral cancer surgery, 2) their performance, and 3) their clinical relevance.METHODS:A literature search was performed in the following databases: Embase, Medline, Web of Science Core Collection, Cochrane Central Register of Controlled Trials, and Google Scholar (from inception to January 23, 2020). IOARM performance was assessed in terms of accuracy, sensitivity, and specificity in predicting margin status, and the reduction of inadequate margins. Clinical relevance (i.e., overall survival, local recurrence, regional recurrence, local recurrence-free survival, disease-specific survival, adjuvant therapy) was recorded if available.RESULTS:Eighteen studies were included in the review, of which 10 for soft tissue and 8 for bone. For soft tissue, defect-driven IOARM-studies showed the average accuracy, sensitivity, and specificity of 90.9%, 47.6%, and 84.4%, and specimen-driven IOARM-studies showed, 91.5%, 68.4%, and 96.7%, respectively. For bone, specimen-driven IOARM-studies performed better than defect-driven, with an average accuracy, sensitivity, and specificity of 96.6%, 81.8%, and 98%, respectively. For both, soft tissue and bone, IOARM positively impacts patient outcome.CONCLUSION:IOARM improves margin-status, especially the specimen-driven IOARM has higher performance compared to defect-driven IOARM. However, this conclusion is limited by the low number of studies reporting performance results for defect-driven IOARM. The current methods suffer from inherent disadvantages, namely their subjective character and the fact that only a small part of the resection surface can be assessed in a short time span, causing sampling errors. Therefore, a solution should be sought in the field of objective techniques that can rapidly assess the whole resection surface.

For vulvar squamous cell carcinoma (VSCC), the mainstay of treatment is surgical removal with tumour-free margins. Surgeons still operate without objective tools that provide margin-status. This study assesses Raman spectroscopy potentiality for distinguishing ex-vivo VSCC from healthy tissue in 11 patients. Grid-based Raman maps were obtained from processed spectra. Water content and C-H band ratio (2,910-2,966 cm -1 / 2810-2890 cm -1 ) were calculated per spectrum and used as linear discriminant parameters. Healthy tissue was differentiated from VSCC with 0.90 discriminative power, 0.79 sensitivity and 0.86 specificity.This is an important step towards the development of objective tools for VSCC surgical guidance.

This study reports on the effects of insertion velocity, needle tip geometry and needle diameter on tissue deformation and maximum insertion force. Moreover, the effect of multiple insertions with the same needle on the maximum insertion force is reported. The tissue deformation and maximum insertion force strongly depend on the insertion velocity and the tip geometry. No correlation was found between the outer diameter and the maximum insertion force for small needles (30G - 32G). The endurance experiments showed no remarkable difference in the maximum insertion force during 100 insertions.

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.

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.

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.

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.

Background Specimen-driven intraoperative assessment of the resection margins provides immediate feedback if an additional excision is needed. However, relocation of an inadequate margin in the wound bed has shown to be difficult. The objective of this study is to assess a reliable method for accurate relocation of inadequate tumor resection margins in the wound bed after intraoperative assessment of the specimen. Methods During oral cavity cancer surgery, the surgeon placed numbered tags on both sides of the resection line in a pair-wise manner. After resection, one tag of each pair remained on the specimen and the other tag in the wound bed. Upon detection of an inadequate margin in the specimen, the tags were used to relocate this margin in the wound bed. Results The method was applied during 80 resections for oral cavity cancer. In 31 resections an inadequate margin was detected, and based on the paired tagging an accurate additional resection was achieved. Conclusion Paired tagging facilitates a reliable relocation of inadequate margins, enabling an accurate additional resection during the initial surgery.

Raman spectroscopy in the high-wavenumber spectral region (HWR) is particularly suited for fiber-optic in vivo medical applications. The most-used fiber-optic materials have negligible Raman signal in the HWR. This enables the use of simple and cheap single-fiber-optic probes that can be fitted in endoscopes and needles. The HWR generally shows less tissue luminescence than the fingerprint region. However, the luminescence can still be stronger than the Raman signal. Hardware- and software-based strategies have been developed to correct for these luminescence signals. Typically, hardware-based strategies are more complex and expensive than software-based solutions. Effective software strategies have almost exclusively been developed for the fingerprint region. First-order polynomial baseline fitting (PBF) is the most common background/luminescence estimation employed for the HWR. The goal of this study was to characterize the luminescence background signals of HW spectra of human oral tissue and compare the performance of two algorithms for correction of these background signals: PBF and multiple regression fitting (MRF). In the MRF method, we introduce here, prior knowledge of the range of Raman signals that can be obtained from the tissues of interest is explicitly used. MRF is more robust than PBF because it does not require an a priori choice of the polynomial order for fitting the background signal. This is important because, as we show, no single polynomial order can optimally characterize all backgrounds that are encountered in HW tissue spectra. We conclude that MRF should be the preferred method for background subtraction in the HWR.

Objectives: The aim of this study was to investigate the potential of Raman spectroscopy for detection of oral cavity squamous cell carcinoma (OCSCC) in bone resection surfaces during mandibulectomy. Materials & methods: Raman mapping experiments were performed on fresh mandible resection specimens from patients treated with mandibulectomy for OCSCC. A tumour detection algorithm was created based on water concentration and the high-wavenumber range (2800 cm(-1)-3050 cm(-1)) of the Raman spectra. Results: Twenty-six ex vivo Raman mapping experiments were performed on 26 fresh mandible resection specimens obtained from 22 patients. The algorithm was applied on an independent test set and showed an accuracy of 95%, a sensitivity of 95%, and a specificity of 87%. Conclusion: These results form the basis for further development of a Raman spectroscopy tool as an objective method for intraoperative assessment of bone resection margins. (C) 2018 Elsevier Ltd. All rights reserved.

Oncological applications of Raman spectroscopy have been contemplated, pursued, and developed at academic level for at least 25 years. Published studies aim to detect pre-malignant lesions, detect cancer in less invasive stages, reduce the number of unnecessary biopsies and guide surgery towards the complete removal of the tumour with adequate tumour resection margins. This review summarizes actual clinical needs in oncology that can be addressed by spontaneous Raman spectroscopy and it provides an overview over the results that have been published between 2007 and 2017. An analysis is made of the current status of translation of these results into clinical practice. Despite many promising results, most of the applications addressed in scientific studies are still far from clinical adoption and commercialization. The main hurdles are identified, which need to be overcome to ensure that in the near future we will see the first Raman spectroscopy-based solutions being used in routine oncologic diagnostic and surgical procedures.