Indocyanine green uptake by human tumor and non­tumor cell lines and tissue.

Indocyanine green (ICG) is a potential promising dye for a better intraoperative tumor border definition and an improved patient outcome by potentially improving tumor border visualization compared with traditional white light guided surgery. Here, the cellular uptake of ICG in human squamous cell carcinoma (SCC026) and immortalized non-cancer skin (HaCaT) cell lines was evaluated to study the tumor-specific cellular uptake of ICG. The spatial distribution of ICG inside tumor tissue was investigated in tissue sections of head and neck squamous cell carcinoma at a microscopic level. ICG uptake and internalization was observed in living cells after 2.5 h and in the nucleus after 24 h. In dead cells, higher and faster uptake was observed. In the tissue sections, higher ICG signal intensity could be detected in connective tissue and surrounding clusters and blood vessels. In conclusion, no distinct ICG uptake by tumor cells was detected in cancer cell lines and tumor tissue. ICG localization in certain regions of tumor tissue appears to be a result of enhanced tissue permeability and retention, but not specific to tumor cells.

Hyperspectral imaging and evaluation of surgical margins: where do we stand?

PURPOSE OF REVIEW: To highlight the recent literature on the use of hyperspectral imaging (HSI) for cancer margin evaluation ex vivo, for head and neck cancer pathology and in vivo during head and neck cancer surgery. RECENT FINDINGS: HSI can be used ex vivo on unstained and stained tissue sections to analyze head and neck tissue and tumor cells in combination with machine learning approaches to analyze head and neck cancer cell characteristics and to discriminate the tumor border from normal tissue. Data on in vivo applications during head and neck cancer surgery are preliminary and limited. Even now an accuracy of 80% for tumor versus nonneoplastic tissue classification can be achieved for certain tasks, within the current in vivo settings. SUMMARY: Significant progress has been made to introduce HSI for ex vivo head and neck cancer pathology evaluation and for an intraoperative use to define the tumor margins. To optimize the accuracy for in vivo use, larger HSI databases with annotations for head and neck cancer are needed.

Multi-Class Cancer Subtyping in Salivary Gland Carcinomas with MALDI Imaging and Deep Learning.

Salivary gland carcinomas (SGC) are a heterogeneous group of tumors. The prognosis varies strongly according to its type, and even the distinction between benign and malign tumor is challenging. Adenoid cystic carcinoma (AdCy) is one subgroup of SGCs that is prone to late metastasis. This makes accurate tumor subtyping an important task. Matrix-assisted laser desorption/ionization (MALDI) imaging is a label-free technique capable of providing spatially resolved information about the abundance of biomolecules according to their mass-to-charge ratio. We analyzed tissue micro arrays (TMAs) of 25 patients (including six different SGC subtypes and a healthy control group of six patients) with high mass resolution MALDI imaging using a 12-Tesla magnetic resonance mass spectrometer. The high mass resolution allowed us to accurately detect single masses, with strong contributions to each class prediction. To address the added complexity created by the high mass resolution and multiple classes, we propose a deep-learning model. We showed that our deep-learning model provides a per-class classification accuracy of greater than 80% with little preprocessing. Based on this classification, we employed methods of explainable artificial intelligence (AI) to gain further insights into the spectrometric features of AdCys.

Intraoperative Assessment of Tumor Margins in Tissue Sections with Hyperspectral Imaging and Machine Learning.

The intraoperative assessment of tumor margins of head and neck cancer is crucial for complete tumor resection and patient outcome. The current standard is to take tumor biopsies during surgery for frozen section analysis by a pathologist after H&E staining. This evaluation is time-consuming, subjective, methodologically limited and underlies a selection bias. Optical methods such as hyperspectral imaging (HSI) are therefore of high interest to overcome these limitations. We aimed to analyze the feasibility and accuracy of an intraoperative HSI assessment on unstained tissue sections taken from seven patients with oral squamous cell carcinoma. Afterwards, the tissue sections were subjected to standard histopathological processing and evaluation. We trained different machine learning models on the HSI data, including a supervised 3D convolutional neural network to perform tumor detection. The results were congruent with the histopathological annotations. Therefore, this approach enables the delineation of tumor margins with artificial HSI-based histopathological information during surgery with high speed and accuracy on par with traditional intraoperative tumor margin assessment (Accuracy: 0.76, Specificity: 0.89, Sensitivity: 0.48). With this, we introduce HSI in combination with ML hyperspectral imaging as a potential new tool for intraoperative tumor margin assessment.

High-resolution MRI of the human palatine tonsil and its schematic anatomic 3D reconstruction.

The palatine tonsils form an important part of the human immune system. Together with the other lymphoid tonsils of Waldeyer's tonsillar ring, they act as the first line of defense against ingested or inhaled pathogens. Although histologically stained sections of the palatine tonsil are widely available, they represent the tissue only in two dimensions and do not provide reference to three-dimensional space. Such a representation of a tonsillar specimen based on imaging data as a 3D anatomical reconstruction is lacking both in scientific publications and especially in textbooks. As a first step in this direction, the objective of the present work was to image a resected tonsil specimen with high spatial resolution in a 9.4 T small-bore pre-clinical MRI and to combine these data with data from the completely sectioned and H&E stained same palatine tonsil. Based on the information from both image modalities, a 3D anatomical sketch was drawn by a scientific graphic artist. In perspective, such studies could help to overcome the difficulty of capturing the spatial extent and arrangement of anatomical structures from 2D images and to establish a link between three-dimensional anatomical preparations and two-dimensional sections or illustrations, as they have been found so far in common textbooks and anatomical atlases.

Fluorescein-Guided Panendoscopy for Head and Neck Cancer Using Handheld Probe-Based Confocal Laser Endomicroscopy: A Pilot Study.

BACKGROUND: White-light endoscopy and microscopy combined with histological analysis is currently the mainstay for intraprocedural tissue diagnosis during panendoscopy for head and neck cancer. However, taking biopsies leads to selection bias, ex vivo histopathology is time-consuming, and the advantages of in-vivo intraoperative decision making cannot be used. Confocal laser endomicroscopy (CLE) has the potential for a rapid and histological assessment in the head and neck operating room. METHODS: Between July 2019 and January 2020, 13 patients (69% male, median age: 61 years) with newly diagnosed head and neck cancer (T3/T4: 46%) underwent fluorescein-guided panendoscopy. CLE was performed from both the tumor and margins followed by biopsies from the CLE spots. The biopsies were processed for histopathology. The CLE images were ex vivo classified blinded with a CLE cancer score (DOC score). The classification was compared to the histopathological results. RESULTS: Median additional time for CLE during surgery was 9 min. A total of 2,565 CLE images were taken (median CLE images: 178 per patient; 68 per biopsy; evaluable 87.5%). The concordance between histopathology and CLE images varied between the patients from 82.5 to 98.6%. The sensitivity, specificity, and accuracy to detect cancer using the classified CLE images was 87.5, 80.0, and 84.6%, respectively. The positive and negative predictive values were 87.0 and 80.0%, respectively. CONCLUSION: CLE with a rigid handheld probe is easy and intuitive to handle during panendoscopy. As next step, the high accuracy of ex vivo CLE image classification for tumor tissue suggests the validation of CLE in vivo. This will evolve CLE as a complementary tool for in vivo intraoperative diagnosis during panendoscopy.