Targeting neovascularization and respiration of tumor grafts grown on chick embryo chorioallantoic membranes

Abstract

Since growing tumors stimulate angiogenesis, via vascular endothelial growth factor (VEGF), angiogenesis inhibitors (AIs, blockers of the VEGF signaling pathway) have been introduced to cancer therapy. However, AIs often yielded only modest and short-lived gains in cancer patients and more invasive tumor phenotypes in animal models. Combining anti-VEGF strategies with lactate uptake blockers may boost both efficacy and safety of AIs. We assessed this hypothesis by using the ex ovo chorioallantoic membrane (CAM) assay. We show that AI-based monotherapy (Avastin®, AVA) increases tumor hypoxia in human CAM cancer cell xenografts and cell spread in human as well as canine CAM cancer cell xenografts. In contrast, combining AVA treatment with lactate importer MCT1 inhibitors (α-cyano-4-hydroxycinnamic acid (CHC) or AZD3965 (AZD)) reduced both tumor growth and cell dissemination of human and canine explants. Moreover, combining AVA+AZD diminished blood perfusion and tumor hypoxia in human explants. Thus, the ex ovo CAM assay as an easy, fast and cheap experimental setup is useful for pre-clinical cancer research. Moreover, as an animal-free experimental setup the CAM assay can reduce the high number of laboratory animals used in pre-clinical cancer research.

Development and Validation of Lactate Metabolism-Related lncRNA Signature as a Prognostic Model for Lung Adenocarcinoma

“…For instance, preclinical studies have demonstrated that integrating the MCT1 inhibitor AZD3965 and anti-PD-1 treatment suppresses the infiltration of excessive PD-1 + Tim-3 + T cells in solid tumors while simultaneously improving antitumor immunity ( 48 ). In another preclinical investigation, the combination of angiogenesis inhibitors (which block the VEGF signaling pathway) and AZD3965 decreases tumor development while simultaneously decreasing both hypoxia and blood perfusion in tumor tissues ( 49 ). Despite limited research available on the combined application of tyrosine kinase inhibitors (TKIs) and MCT1 inhibitors, one preclinical study illustrated that AZD3965 strongly impedes cell proliferation and motility in both TKI-resistant and -sensitive NSCLC cells-implicating excellent potential of combination therapies to treat this disease ( 50 ).…”

Bridging the In Vitro to In Vivo gap: Using the Chick Embryo Model to Accelerate Nanoparticle Validation and Qualification for In Vivo studies

“…This imaging modality records dynamic blood flow in real time and can reveal tumor hypoxia and angiogenic vessel growth induced by the xenograft. LSPI was used to gain insight into the response of glioblastoma xenografts to antiangiogenic and lactate uptake inhibitors in the CAM . Antiangiogenic treatments in clinical trials have not led to the expected increase in survival and cure for cancer patients and have demonstrated increased metastasis in animal models.…”

“…When a second treatment was added, a lactate uptake blocker, active angiogenesis was reduced without the increase in tumor hypoxia seen with antiangiogenic treatment alone. Further, dual treatment reduced the spread of tumor cells out from the xenograft . While this research focused on drug therapies, this same methodology could be applied to nanomaterials designed to alter angiogenesis or target tumor neovasculature.…”

“…One of the clear advantages to the CAM system is the ability to visualize both the xenograft and the associated vasculature without the need for surgery to expose the tumor. This also makes the CAM xenograft model system ideal for technologies that monitor vascular perfusion in real time, such as laser-speckle perfusion imaging (LSPI; Figure A) . This imaging modality records dynamic blood flow in real time and can reveal tumor hypoxia and angiogenic vessel growth induced by the xenograft.…”

Replacement in angiogenesis research: Studying mechanisms of blood vessel development by animal-free in vitro, in vivo and in silico approaches

“…These can either be topically applied to the CAM ( Liu et al, 2022 ) or administered systemically by intravascular injection ( Storgard et al, 2005 ). In addition, the CAM frequently serves as host tissue to study the vascularization of implanted biomaterials as well as benign and malignant tissue grafts ( Gescher et al, 2005 ; Ademi et al, 2021 ; Ara et al, 2022 ). The latter ones can be analyzed independently of their species origin without the risk of immunological rejection, because the early chicken embryo lacks a functional immune system ( Baiguera et al, 2012 ).…”