Mechanism of resistance to lysosomotropic treatments: inhibition of autophagy and metabolic adaptations – LYSORES

State of art
The physicochemical properties of drugs can lead to a therapeutic failure. Physicochemical properties can predictably influence intracellular distribution of a drug, an important consideration for its efficacy. In particular, drugs with LogP>2 and pKa>6 can be trapped in acidic intracellular compartments such as lysosomes away from their targets. Their protonation at acidic pH causes their sequestration and subsequent limitations of drug efficacy. These drugs are qualified as lysosomotropic.

Preliminary results
We demonstrated that lysosomal sequestration results in an increase of the lysosome pH, which inhibits lysosomal proteases and leads to an incomplete autophagic process [1]. The physio-pathological consequences of sequestration in lysosomes and the adaptation to autophagic defects need better understanding. Our recent results reveal that lysosomotropic-dependent inhibition of autophagy results in the activation of the serine biosynthetic pathway leading to a metabolic adaptation and sustained proliferation. The comprehensive investigations of metabolic alterations in response to lysosomotropic drugs resulting in acquired resistance have never been conducted.

Objectives
Mainly dedicated to sunitinib, the standard care of clear cell Renal Cell Carcinoma (ccRCC), our work will be extended to lysosomotropic drugs such as chloroquine (anti-malaria) or azithromycin (antibiotic, asthma).
The objectives of our project are:

(1) To detect the lysosomotropic potential of any drugs
To develop a "biotest" probe for the detection of lysosomotropic properties. In collaboration with the Institute of Chemistry of Nice, we will produce fluorescent probes for the accurate detection of lysosome pH variations. This biotest, which can be used routinely, will detect the lysosomotropic potential of any drugs and will therefore assess the efficacy of the drug before any drug development. This test will be patented and proposed to pharmaceutical companies.

(2) To understand the consequences of lysosome sequestration of drugs on metabolomic adaptations
We have identified the serine biosynthetic pathway as a potential mechanism of resistance (transcriptomic and metabolomic). We will characterize and validate the importance of this pathway in drug resistance (cell lines, patient primary cells and organoids). Also, we will test the therapeutic interest of targeting the actors of this metabolic pathway, an alternative strategy that we can propose to patients in therapeutic impasses.

Expected results
This project will evidence mechanistic and clinical concepts counteracting resistance to lysosomotropic drugs. The applications of this project are for diagnostic, prognostic and therapeutic purposes. A biotest, used to evaluate the lysosomotropic potential of drugs currently on the market or under development in preclinical studies, could anticipate failures of phase I/II clinical trials. A better knowledge of the mechanisms of resistance will promote; a) therapeutic strategies combining lysosomotropic drugs with inhibitors of metabolic enzymes; b) the identification of predictive markers of resistance.