The immune system is a fine modulator of the tumor biology supporting or inhibiting its progression, growth, invasion and conveys the pharmacological treatment effect. Tumors, on their side, have developed escaping mechanisms from the immune system action ranging from the direct secretion of biochemical signals to an indirect reaction, in which the cellular actors of the tumor microenvironment (TME) collaborate to mechanically condition the extracellular matrix (ECM) making it inhospitable to immune cells. TME is composed of several cell lines besides cancer cells, including tumor-associated macrophages, cancer-associated fibroblasts, CD4⁺ and CD8⁺ lymphocytes, and innate immunity cells. These populations interface with each other to prepare a conservative response, capable of evading the defense mechanisms implemented by the host’s immune system. The presence or absence, in particular, of cytotoxic CD8⁺ cells in the vicinity of the main tumor mass, is able to predict, respectively, the success or failure of drug therapy. Among various mechanisms of immunescaping, in this study, we characterized the modulation of the phenotypic profile of CD4⁺ and CD8⁺ cells in resting and activated states, in response to the mechanical pressure exerted by a three-dimensional in vitro system, able to recapitulate the rheological and stiffness properties of the tumor ECM.

Duchenne muscular dystrophy (DMD) is caused by the absence of the full form of the dystrophin protein, which is essential for maintaining the structural integrity of muscle cells, including those in the heart and respiratory system. Despite progress in understanding the molecular mechanisms associated with DMD, myocardial insufficiency persists as the primary cause of mortality, and existing therapeutic strategies remain limited. This study investigates the hypothesis that a dysregulation of the biological communication between infiltrating macrophages (MPs) and neurocardiac junctions exists in dystrophic cardiac tissue. In a mouse model of DMD ( mdx ), this phenomenon is influenced by the over‐release of chondroitin sulfate proteoglycan‐4 (CSPG4), a key inhibitor of nerve sprouting and a modulator of the neural function, by MPs infiltrating the cardiac tissue and associated with dilated cardiomyopathy, a hallmark of DMD. Givinostat, the histone deacetylase inhibitor under current development as a clinical treatment for DMD, is effective at both restoring a physiological microenvironment at the neuro‐cardiac junction and cardiac function in mdx mice in addition to a reduction in cardiac fibrosis, MP‐mediated inflammation, and tissue CSPG4 content. This study provides novel insight into the pathophysiology of DMD in the heart, identifying potential new biological targets. © 2024 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Tumor associated fibroblasts (TAFs) play a key role in tumor growth and metastatization. TAFs overexpress different biomarkers that are usually expressed at low levels in physiological conditions. Among them are the fibroblast growth factor receptors (FGFRs) that bind the fibroblast growth factors (FGFs). In particular, the overexpression of FGFR-2c in tumors has been associated with advanced clinical stages and increased metastatization. Here, we developed a non-invasive tool to evaluate, in vivo, the expression of FGFR-2c in metastatic cancer. This is based on ^99mTc-labelled FGF-2. Methods: ^99mTc-FGF-2 was tested in vitro and in vivo in mice bearing allografts of sarcoma cells. Images of ^99mTc-FGF-2 were acquired using a new portable high-resolution ultra-sensitive gamma camera for small animal imaging. Results: FGF-2 was labeled with high specific activity but low labelling efficiency, thus requiring post-labeling purification by gel-filtration chromatography. In vitro binding to 2C human keratinocytes showed a Kd of 3.36 × 10⁻⁹ M. In mice bearing J774A.1 cell allografts, we observed high and rapid tumor uptake of ^99mTc-FGF-2 with a high Tumor/Blood ratio at 24 h post-injection (26.1 %ID/g and 12.9 %ID) with low kidney activity and moderate liver activity. Conclusions: we labeled FGF-2 with ^99mTc and showed nanomolar Kd in vitro with human keratinocytes expressing FGF-2 receptors. In mice, ^99mTc-FGF-2 rapidly and efficiently accumulated in tumors expressing FGF-2 receptors. This new radiopharmaceutical could be used in humans to image TAFs.

Techniques like single photon emission computed tomography (SPECT) show high potential in small animals in vivo imaging framework. In this context, the spatial resolution represents an important parameter influencing scanner design, given the small size of the anatomical structures to be investigated. The purpose of the present work was to assess the performance of a scintigraphic system with an improved spatial resolution by applying the previously described Super Spatial Resolution (SSR) method. Our dual-head SPECT system is composed of gamma cameras based on the Hamamatsu H13700 position-sensitive photomultiplier tube (PSPMT). In each detector head, the PSPMT is coupled to a 28 × 28 array of CRY018 scintillation crystals. The pure Tungsten parallel square hole collimator ensures the position sensitivity, and a dedicated resistive chain readout so as an ADC board have been proprietary designed. To finalize the mechanical development of the SSR-SPECT system several tests were carried out. Based on the results obtained in the test phase, a partial review of the mechanical design was performed. Then a dedicated mechanical motion software was developed, and in particular, a kinematic software debugging and testing was assessed. Several experiments were carried out by using Derenzo phantoms and capillaries filled with radioactive sources. Finally, the performance of our system was evaluated by performing small animal imaging studies. The SPECT spatial resolution was experimentally determined to be about 1.6 mm. We reach a resolution of 1.18 mm by applying the SSR based on two images. The results of this study demonstrated the good capability of the system as a suitable tool for preclinical imaging especially in fields like neuroscience for the study of small brain structures.

Targeting fibrosis in Duchenne muscular dystrophy (DMD)-associated cardiomyopathy is a critical outstanding clinical issue, as cardiac failure remains a leading cause of death despite advances in supportive care. This study evaluates the therapeutic efficacy of CSPG4-targeted chimeric antigen receptor (CAR) T cells in reducing cardiac fibrosis and improving heart function in a preclinical model of the disease. DMD is a progressive genetic disorder characterized by degeneration of skeletal and cardiac muscle. Cardiomyopathy, driven by fibrosis and chronic inflammation, is a leading contributor to mortality in affected patients. Proteoglycans such as CSPG4, critical regulators of extracellular matrix dynamics, are markedly overexpressed in dystrophic hearts and promote pathological remodeling. Current treatments do not adequately target the fibrotic and inflammatory processes underlying cardiac dysfunction. CSPG4-specific CAR-T cells were engineered and administered to dystrophic mice. Therapeutic efficacy was assessed through histological, molecular, and echocardiographic analyses evaluating cardiac fibrosis, inflammation, innervation, and overall function. Treatment with CSPG4 CAR-T cells preserved myocardial integrity, improved cardiac performance, and reduced both fibrosis and inflammatory markers. The therapy also restored cardiac innervation, indicating a reversal of neural remodeling commonly seen in muscular dystrophy-related cardiomyopathy. CSPG4-targeted CAR-T therapy offers a novel, cell-based strategy to mitigate cardiac remodeling in dystrophic hearts. By addressing core fibrotic and inflammatory drivers of disease, this approach represents a significant advancement in the development of precision immune therapies for muscular dystrophies and cardiovascular conditions.