Several Fungal species have the potential to cause a broad spectrum of diseases in humans, ranging from mild superficial to disseminated invasive infections that involve the bloodstream and vital organs. Invasive fungal infections are severe, life-threatening diseases that result in the deaths of 1.5 million patients each year. The most common fungal species responsible for the majority of invasive fungal infections include Candida, Cryptococcus, and Aspergillus. The current treatment options for invasive fungal infections are restricted to three classes of antifungals: Azoles, polyenes, and echinocandins. The emergence of new fungal species, especially C. auris, marked by high resistance profiles and increased mortality rates (30-60%), has further exacerbated the limitations in its therapeutic options. This emphasizes the urgent need for effective alternatives to combat these deadly pathogens. C. auris isolates exhibited high resistance capability especially against azole (fluconazole) and polyene (amphotericin B) antifungals. Here, we utilized the combinatorial strategy to screen ~3400 FDA-approved drugs and clinical compounds to identify hits that can enhance/restore the antifungal activity of azoles and amphotericin B against resistant C. auris. The HIV protease inhibitors (lopinavir and ritonavir) were identified as potent enhancers to the antifungal activity of azole drugs (fluconazole, voriconazole and itraconazole). We confirmed that lopinavir and ritonavir have the capability to interfere with fungal efflux pump machinery. The in vivo efficacy of the combination of azole antifungals and HIV protease inhibitors was also evaluated to discover the best combination of itraconazole, lopinavir and ritonavir. Three drugs (lansoprazole, rolapitant and idebenone) were identified to effectively enhance the antifungal effects of amphotericin B and overcome its resistance in C. auris. Furthermore, the synergistic interactions of these combinations were applied on other medically important Candida, Cryptococcus, and Aspergillus species. In a comprehensive mechanistic study, we discovered that lansoprazole interferes with an essential target in the fungal mitochondrial cytochrome system, cytochrome bc1. This interference induces oxidative stress in fungal cells and subsequently enhances the antifungal activity of amphotericin B. For rolapitant, a transcriptomic analysis along with ATP luminescence assays confirmed that rolapitant at sub-inhibitory concentrations significantly interferes with ATP production in C. auris. For idebenone, checkerboard assays confirmed the synergistic interactions between amphotericin B and idebenone against a diversity of medically important fungal species. This combination exhibited a rapid fungicidal activity within 4 hours. Additionally, the cytotoxicity of this combination was assessed in a cell line model of kidney cells. Based on the potent in vitro and in vivo synergistic relationships observed for the identified combinations, it can be concluded that our approach offers a new hope to restore the antifungal activity of the existing antifungal drugs, even against resistant fungal infections. Additionally, it provides valuable insights into identifying novel targets to overcome resistance in multidrug-resistant fungal pathogens. / Doctor of Philosophy / Fungi comprise a diverse group of organisms that interact with humans in many good and bad aspects. Candida auris, a recently identified fungus, poses a significant threat to patients with weak immune systems. Infections with C. auris can be associated with mortality rates of up to 60%. Notably, this fungus is characterized by its powerful spreading capability and displays extraordinary resistance to antifungal agents, rendering many existing antifungal drugs ineffective. As a result, there is an unmet need to find efficient treatments for such deadly fungal infections. In this study, several drugs were identified with the potential to restore the activity of traditional antifungal drugs. The study identified four promising drugs (lopinavir, lansoprazole, rolapitant, and idebenone) with the potential to enhance the activity of the antifungal drugs against C. auris. lopinavir showed great potential to enhance the activity of azole antifungals, including fluconazole, voriconazole, and itraconazole. Furthermore, three other drugs (lansoprazole, rolapitant, and idebenone) were identified for their potential to enhance the activity of amphotericin B, which is considered a last-line antifungal therapy. We clarified the mechanisms by which these drugs could restore the activity of antifungal agents. Finally, we confirmed the effectiveness of these combinations in animal models, providing valuable insights into their potential for clinical applications. In summary, our research has opened promising avenues to overcome resistance and develop new treatments for hard-to-treat fungal infections.