Evolution of drug resistance and associated trade-offs in non-albicans Candida pathogens

Juan Carlos Núñez Rodriguez

Dr. Toni Gabaldón

Genómica comparativa

According to the World Health Organization, antimicrobial resistance is one of the most urgent global health challenges of the 21st century. In this context, resistance to antifungals is particularly concerning, as there are only three families of antifungal drugs available for clinical use. Invasive fungal infections affect 6.5 million people each year and cause more than 2.5 million deaths directly attributable to fungal diseases, figures comparable to those of diseases such as tuberculosis or malaria. Among fungal pathogens, Candida non-albicans (NAC) species have experienced a remarkable increase in recent decades, to the point of currently representing more than half of all Candida infections. These species show a great capacity to adapt to antifungal treatments thanks to their high genomic plasticity and their intrinsic resistance to certain drugs. However, unlike C. albicans, the molecular mechanisms that explain this resistance in many NACs remain poorly understood, often reduced to the study of a small set of key genes. This doctoral thesis aimed to deepen our knowledge of the mechanisms of resistance and evolutionary adaptation in NAC species, especially in Nakaseomyces (Candida) glabratus and in the three species of the Candida parapsilosis complex. To achieve this purpose, a new methodology called Q-PHAST has been developed, which allows large-scale phenotypic studies to be performed in a quantitative and accessible manner. Combined with in vitro evolution experiments and complete genome sequencing, this tool has provided an unprecedented level of resolution in the study of antifungal resistance processes. Through this integrated approach, the thesis has described the genomic mechanisms underlying resistance to azoles and echinocandins in the C. parapsilosis complex, identifying mutations, aneuploidies, copy number variations and haploidization processes that contribute to resistance. In N. glabratus, it has been shown that the acquisition of resistance can lead to associated vulnerabilities (trade-offs) that can be therapeutically exploited to eliminate multiresistant strains. As a direct consequence of this work, the CandiRes project has been initiated, a platform for the discovery of new antifungals based on the knowledge obtained during the thesis. This platform is optimized to design first-in-class drugs with specific mechanisms of action for NAC species active against multiresistant strains. This approach allows the development of a new generation of precision antifungal drugs, specifically targeted against difficult-to-treat species with a high potential for resistance. The CandiRes project has the potential to transform the treatment of invasive fungal infections, contributing to reducing mortality and improving patient prognosis, making all the basic knowledge acquired in the field of resistance directly applicable to solving real clinical problems. Overall, this work establishes a bridge between fundamental research and biomedical innovation, opening the way to new precision antifungal therapies that could mark a before and after in the fight against fungal infections.

The thesis by Juan Carlos Núñez Rodríguez addresses the urgent need for new antifungal therapies due to the rise of multidrug-resistant non-albicans Candida (NAC) species, which cause severe invasive fungal diseases (IFDs) with high mortality and limited treatment options. Key points: Clinical Context: IFDs cause over 3.8 million deaths annually, with candidiasis invasive (CI) being a major contributor. Mortality rates reach 40–70%, and resistance to azoles, echinocandins, and polyenes is increasing. Research Achievements: Developed Q-PHAST (Quantitative Phenotyping and Antifungal Susceptibility Testing), a high-throughput phenotyping platform published in Nature Protocols. Discovered polygenic and dynamic resistance mechanisms in NAC species, including mutations, aneuploidies, and haploidization. Identified collateral vulnerabilities in resistant strains, enabling selective elimination using drugs like cyclosporin A. Translational Impact: Founded CandiRes, a spin-off project focused on discovering first-in-class precision antifungal drugs targeting NAC species. CandiRes Strategy: Screens compounds directly on multiresistant clinical isolates using a proprietary biobank (>500 sequenced strains) and a chemical library (>150,000 compounds). Employs high-throughput screening (HTS), genomic analysis, transcriptomics, and haploinsufficiency profiling to identify new mechanisms of action. Current candidates outperform fluconazole, show no mammalian toxicity, and are in hit-to-lead development. Commercialization: Plans include patent protection, spin-off creation, and strategic partnerships with biotech and pharma companies. Impact: Potential to reduce mortality, shorten hospital stays, and address one of the most urgent unmet medical needs globally.

Invasive fungal diseases, IFDs, Candidiasis invasive, CI, Candida, Non-albicans Candida, NAC, Candida auris, Candida parapsilosis, Nakaseomyces glabratus, Drug resistance, Multidrug resistance, MDR, Antifungal therapy, Azoles, Echinocandins, Polyenes, Q-PHAST, Quantitative phenotyping, High-throughput screening, HTS, Evolutionary trade-offs, Collateral sensitivity, Cyclosporin A, Calcineurin pathway, FKS mutations, Haploidization, Genomic analysis, Transcriptomics, RNA-seq, Haploinsufficiency profiling, HIP, Biobank, Chemical library, New chemical entities, NCE, Mechanism of action, MoA, Structure-activity relationship, SAR, ADME-Tox, Drug discovery, Precision medicine, Spin-off, CandiRes, IRB Barcelona, Drug screening platform, Patent protection, Markush structures, Tech transfer, Translational research, Antifungal innovation, Global health, High mortality infections, Neglected diseases, Pharmaceutical partnerships, Clinical unmet needs.