This project with a strong translational character, aims to identify and validate novel therapeutic targets present in the tumor immune microenvironment (TIME) and which is responsible for tumor progression and resistance to current immunotherapeutic approaches with immune checkpoint inhibitors. To this end, we will use modern 'omics' technologies together with an in-depth bioinformatics analysis on the characterization of TIME from patients with adenocarcinoma of the lung (LUAD). By using malignant pleural effusions (MPEs), we will recreate ex vivo systems comparable to the patients' tumors. The experimental program will allow us to devise new and more effective therapeutic combinations. The interdisciplinary team proposing the project consists of 4 Operational Units (OUs) which is composed of medical oncologists, pathologists, biologists, and immunologists that belong two different institutes such as the Department of Clinical and Molecular Medicine (DMCM) at the Azienda Ospedaliera Sant'Andrea Università La Sapienza in Rome and IRCCS Regina Elena National Cancer Institute (IRE). The OUs led by Prof. Rita Mancini and Prof. Paolo Marchetti at the DMCM and those led by Dr. Maurizio Fanciulli and Dr. Paola Nisticò at the IRE will contribute synergistically to the realization of the following 3 main Tasks:

• Expanding on the biobank obtained from MPEs of LUAD patients
• Identifying therapeutic targets in the immune microenvironment
• Validating targets in ex vivo models (organoids) by immuno-functional assays

The goal of our project is to validate potential new targets using either gene silencing techniques, or commercial monoclonal antibodies specific for the identified targets. However, it is well known that the latter are generally not the best reagents from a therapeutic point of view because they are monoclonals of murine origin that cannot be used for clinical purposes. Therefore, it is necessary either to generate murine antibodies and then subject them to the humanization processes, or to generate human antibodies directly or through the use of transgenic mice humanized for human immunoglobulins, or phage libraries. In this context, both multinational companies and also Biotechs in the biopharmaceutical sector have adequate experience to pursue this avenue. This project is valuable as it represents the synergy between our laboratories (with the development of organoid models directly generated by patients) and companies that produce human or humanized antibodies with their monoclonal antibodies, within the Lazio Bioscience Technological District (DTB).

The therapeutic targets identified and developed during the course of the project will undergo patent protection pathways through the Technology Transfer Offices (TTOs) of the participating institutes, where a plan will be set in place to disseminate and exploit the potential clinical and market spin-offs in case of industrial development.