Sunday, May 18, 2025
5/18/2025
Monitoring Autophagy in the Heart and in Tumors Treated with Potentially Cardiotoxic Chemotherapy - The treatment of cancer has been revolutionized by the development of targeted and immune-based therapies. However, many of these new therapies, as well as established ones such as doxorubicin, can damage the heart and cause severe heart failure. A growing body of evidence suggests that perturbations in cardiomyocyte autophagy may play a central role in chemotherapy-induced heart failure, and the ability to image this process in vivo could provide important insights. We have recently developed an autophagy-detecting nanoparticle (ADN) and have used this agent to image autophagy in the heart during chemotherapy with doxorubicin and dasatanib. The core of the agent consists of ferumoxytol, an MRI-detectable nanoparticle, to which several polyarginine- Cy5.5 moieties are attached. The polyarginine peptides facilitate the translocation of the agent into the cell, where it is taken up by autophagosomes and trafficked to the lysosomes. Folding of the polyarginine peptides also results in stacking and quenching of the Cy5.5 fluorochromes until the peptides are cleaved by cathepsins in the autophagolysosomes. We have shown using chemical inhibitors/stimulators of autophagy, and cells with genetic deletion of the key autophagy proteins ATG5 and ATG7, that the activation of ADN is specific for autophagy. Likewise, using a transgenic mouse with overexpression of the DDiT4L transcription factor in the heart and the canonical autophagy model of intermittent fasting, we have shown that ADN can detect autophagy in vivo with a high degree of sensitivity and specificity. In this proposal we will use ADN to interrogate changes in autophagy in both the heart and cancer. While the upregulation of autophagy in the heart is protective, it has the potential to either protect or harm tumors undergoing chemotherapy. We hypothesize that the imaging of autophagy with ADN will allow conditions and strategies to be identified where the upregulation of autophagy is cardioprotective but does not attenuate the effect of the chemotherapy on the tumor. In aim 1 of the proposal, we will make a small chemical modification to the probe, enabling it to be detected before and after its activation, and use confocal/super-resolution microscopy of live cells to develop a detailed kinetic model of autophagy. IPSC-derived cardiomyocytes and relevant tumor cell lines will be studied. In aim 2 we will assess the impact of autophagy upregulation on a library of tumor cell lines exposed to a large panel of chemotherapies, many known to cause heart failure. In aim 3 we will use a multispectral fluorescence approach to image autophagy, apoptosis and inflammation in the heart and in tumors in mice in vivo. The impact of autophagy upregulation will be assessed and a detailed kinetic model of autophagy in the heart and in tumors will be derived. Execution of the proposed aims will provide important insights into the role of autophagy in the heart and cancer, and provide a platform to identify safe and effective autophagy regulating strategies to protect the heart during chemotherapy.
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