Towards reliable hyperspectral imaging biomarkers of CT26 murine tumor model

The non-invasive monitoring of tumor growth can offer invaluable diagnostic insights and enhance our understanding of tumors and their microenvironment. Integrating hyperspectral imaging (HSI) with three-dimensional optical profilometry (3D OP) makes contactless and non-invasive tumor diagnosis possible by utilizing the inherent tissue contrast provided by visible (VIS) and near-infrared (NIR) light. Consequently, valuable information regarding tumors and healthy tissues can be extracted from the acquired hyperspectral images. Until now, very few methods have been used to monitor tumor models in vivo daily and non-invasively. In this research, we conducted a 14-day study monitoring BALB/c mice with subcutaneously grown CT26 murine colon carcinomas in vivo, commencing on the day of tumor cell injection. We extracted physiological properties such as total hemoglobin (THB) and tissue oxygenation ( $$ ) using the inverse adding-doubling (IAD) algorithm and manually segmented the tissues. We then selected the ten most relevant features describing tumors using the Max-Relevance Min-Redundancy (MRMR) algorithm and utilized 30 classic and advanced machine learning (ML) algorithms to discriminate tumors from healthy tissues. Finally, we tested the robustness of feature selection and model performance by smoothing tissue parameter maps extracted by IAD with a variable kernel and omitting selected training data. We could discriminate CT26 tumor models from surrounding healthy tissues with an area under the curve (AUC) of up to 1 for models based on the gradient boosting method, linear discriminant analysis, and random forests. Our findings help pave the way for precise and robust imaging biomarkers that could aid tumor diagnosis and advance clinical practice.