Tuberculosis (TB) is an airborne disease transmitted by Mycobacterium tuberculosis (Mtb). Mtb infection may result in active tuberculosis or in latent infection. After Mtb exposure, macrophages and dendritic cells phagocytize bacteria. Mtb can resist intracellular destruction and establish infection leading to granuloma formation where the bacteria reside and persist for a long time, providing a safe shelter for Mtb, rather than being a host-protective containment. Mtb induces damages in lung tissue, taking advantage from its ability to survive inside macrophages. A quarter of the human population is latently infected by TB. Because of COVID-19, WHO reported a disruption of TB services. This situation raises concerns of future TB rebounds, increasing risk of multidrug- resistant (MDR) TB. So, new alternative strategies are needed to fight MDR-TB. For this aim, in this project we want to: face TB/MDR-TB by the eradication of intramacrophage Mtb by using an Antibody Drug Conjugates (ADC) approach; develop a robust method not limited to Mtb but using it as model for the identification of suitable antigens to obtain active ADCs. In fact, ADC technology has been efficiently applied in cancer therapy with 11 molecules actually approved. The PI’s group recently demonstrated that it is not mandatory to anchor a cytotoxic molecule to the monoclonal Antibodies (mAbs) to have an active ADC. Moreover, it is possible to have epigenetic modulation by using ADCs containing inhibitors of Hystone Deacetylase (HDAc) as drugs. These findings opened to the application of ADCs to different diseases beyond cancer. A single example is reported in the literature where an ADC contains an antibiotic for the treatment of S. aureus infections. The development of ADCs active for the treatment of Mtb infections, including MDR, is a very innovative powerful approach. The identification of suitable antigens, allowing the Mtb recognition by the ADC and facilitating the intracellular uptake of Mtb by macrophages, still remains challenging. Today, only a few, well characterized mAbs specific for cell-wall associated Mtb antigens are commercially available (i.e. anti-PstS1 and anti-Mpt83 mAbs), and they are not necessarily the best ones for the development of our ADCs. To overcome all issues related with the selection and production of new mAbs (an extremely time consuming and expensive procedure), we here propose a method to identify interesting Mtb antigens suitable for ADCs development relying on the expression (by recombinant Mtb strains) of modified selected Mtb surface antigens containing tag sequences (hemagglutinin HA epitope), which are recognized by commercial anti-HA-specific mAbs. The anti-PstS1, anti- Mpt83, and the anti-HA mAbs will be charged with different payloads (i.e. recently approved anti-TB drugs Bedaquiline and Pretonamid, or Linezolid). The anti-Mtb activity of the produced ADCs will be evaluated in in vitro, ex vivo and in vivo models.

The project is funded by European Union – Next-GenerationEU – National Recovery and Resilience PLAN (PNRR) – Mission 4, Component 2, Investiment 1.1 Fondo per il Programma Nazionale di Ricerca e Progetti di Rilevante Interesse Nazionale (PRIN). Project N. 2022JTPP53_001