Shear wave elastography and photoacoustic imaging to non-invasively identify resistant regions in the tumor microenvironment

Increased stiffness is a hallmark of malignant tumors and can impede chemotherapy delivery due to elevated interstitial pressure. Clinically, tumor stiffness is assessed using Shear Wave Elastography (SWE); however, small animal imaging systems combining SWE with anatomical, functional, and molecular information have been lacking. Here, we present, for the first time, high-frequency ultrasound with ultrafast plane wave acquisition combined with assessments of capillary function and tumor oxygenation, offering non-invasive, detailed insight into the tumor microenvironment.CT26 murine colorectal carcinoma cells were implanted subcutaneously into the flanks of Balb/C mice to establish solid tumors. Two weeks later, tumor imaging was performed using the Vevo® F2 LAZR-X system. During a single imaging session, tumor structure, vascularization (after intravenous injection of the MicroMarker contrast agent), oxygenation/hypoxia (via photoacoustic imaging), and stiffness (via SWE) were evaluated without moving the animals. An oxygen challenge identified regions responsive to changing conditions (potentially treatable “responder” regions), which were compared with SWE-measured stiffness and vascular data.Our results demonstrate that combining SWE with ultrasound and photoacoustic imaging provides real-time, non-invasive insights into the tumor microenvironment. Regions of high stiffness, identified by SWE, are linked to increased interstitial pressure, which may restrict drug delivery and affect treatment efficacy. By mapping vascular and hypoxic features alongside stiffness, this multimodal approach revealed intra- and inter-tumor heterogeneity. Areas with higher stiffness correlated with non-responsive oxygen regions and lower vascular density, providing a non-invasive representation of evolving tumor characteristics.This study establishes SWE as a powerful tool for non-invasively assessing tumor stiffness and its relationship with hypoxia and vascularity. Integrating SWE with photoacoustic and ultrasound imaging provides a comprehensive, multidimensional method to characterize the tumor microenvironment, moving beyond traditional measures based solely on tumor volume. This advancement enhances understanding of tumor heterogeneity and supports the development of personalized therapeutic strategies, ultimately improving treatment planning and outcomes