Design of macrophages expressing the fourth generation of Chimeric Antigen Receptors for the treatment of solid tumors.

Abstract

Recently, there has been a rapid expansion in the development of oncology therapeutics products. With significant commercial potential, cell and gene therapies have emerged as a promising advancement in the healthcare area. Immunotherapy with CAR-T cells has shown promising results in the treatment of refractory hematological malignancies. However, the successful application of CAR-T therapy in solid tumors remains limited due to the physical peculiarities and physiological characteristics of these tumors. Given the ability to infiltrate solid tumors and interact with the cellular components of the tumor microenvironment, Carisma Therapeutics stood out for pioneering the application of macrophages as a potential candidate for immunotherapy with CAR receptors. However, the use of primary macrophages for immunotherapy is limited, mainly: (1) the presence of cellular mechanisms that difficult their genetic manipulation; (2) the limited capacity for in vitro expansion and, consequently, the difficulty of application on a large clinical scale; (3) its leading role in cytokine release syndrome; (4) the variability among donors, with the cells of some donors being able to respond better or not than others, which hinders the reproducibility of the data and, finally; (6) the definition of the minimum effective dose. Thus, to overcome these limitations and develop for the first time in Brazil immunotherapy with CAR-macrophages (CAR-M), we propose in our technological solution the use of CD34+ cells purified from the placental umbilical cord as a source for CAR-M immunotherapy. The differential of the use of CD34+ hematopoietic stem cells as a source for CAR-M therapy focuses mainly on ease of acquisition and storage, high expansion capacity, and permissibility to gene manipulation. In this way, our solution not only suggests a viable and reliable "off-the-shelf CAR-M", but also a national strategy to reduce the cost of production and increase accessibility to the public health care system and developing countries. In this initial phase, we seek, as a proof of concept, to validate, in vitro, the transduction of CD34+ cells with our 3rd and 4th generation CAR vectors and to evaluate the antitumor activity of CAR+ macrophages differentiated from CD34+ cells. We believe that these generated macrophages will be able to phagocytize the antigen-specific target in addition to contributing to antitumor activity through the generation of ROS and the release of pro-inflammatory cytokines. (AU)