Addressing Delicate and Variable Cancer Morphology in Spectral Histopathology Using Canine Visceral Hemangiosarcoma.

Spectral histopathology has shown promise for the classification and diagnosis of tumors with defined morphology, but application in tumors with variable or diffuse morphologies is yet to be investigated. To address this gap, we evaluated the application of Fourier transform infrared (FTIR) imaging as an accessory diagnostic tool for canine hemangiosarcoma (HSA), a vascular endothelial cell cancer that is difficult to diagnose. To preserve the delicate vascular tumor tissue structure, and potential classification of single endothelial cells, paraffin removal was not performed, and a partial least square discrimination analysis (PLSDA) and Random Forest (RF) models to classify different tissue types at individual pixel level were established using a calibration set (24 FTIR images from 13 spleen specimens). Next, the prediction capability of the PLSDA model was tested with an independent test set (n = 11), resulting in 74% correct classification of different tissue types at an individual pixel level. Finally, the performance of the FTIR spectropathology and chemometric algorithm for diagnosis of HSA was established in a blinded set of tissue samples (n = 24), with sensitivity and specificity of 80 and 81%, respectively. Taken together, these results show that FTIR imaging without paraffin removal can be applied to tumors with diffuse morphology, and this technique is a promising tool to assist in canine splenic HSA differential diagnosis.

Candidate Glycoprotein Biomarkers for Canine Visceral Hemangiosarcoma and Validation Using Semi-Quantitative Lectin/Immunohistochemical Assays.

Visceral hemangiosarcoma (HSA) is one of the more frequent cancers in dogs and has a high metastatic rate and poor prognosis, as clinical signs only become apparent in advanced stages of tumor development. In order to improve early and differential diagnostic capabilities and hence, prognosis for dogs with HSA, two types of biomarker are needed: a point-of-care diagnostic biomarker and a prognostic biomarker-preferentially based on samples obtained with minimally invasive methods. In this study, we applied a lectin magnetic bead array-coupled tandem mass spectrometry (LeMBA-MS/MS) workflow through discovery and validation phases to discover serum glycoprotein biomarker candidates for canine HSA. By this approach, we found that Datura stramonium (DSA), wheat germ agglutinin (WGA), Sambucus nigra (SNA), and Pisum sativum (PSA) lectins captured the highest number of validated candidate glycoproteins. Secondly, we independently validated serum LeMBA-MS/MS results by demonstrating the in situ relationship of lectin-binding with tumor cells. Using lectin-histochemistry and immunohistochemistry (IHC) for key proteins on tissues with HSA and semi-quantitation of the signals, we demonstrate that a combination of DSA histochemistry and IHC for complement C7 greatly increases the prospect of a more specific diagnosis of canine HSA.