Stromal content heavily impacts the transcriptional classification of colorectal cancer (CRC), with clinical and biological implications. Lineage-dependent stromal transcriptional components could therefore dominate over more subtle expression traits inherent to cancer cells. Since in patient-derived xenografts (PDXs) stromal cells of the human tumour are substituted by murine counterparts, here we deploy human-specific expression profiling of CRC PDXs to assess cancer-cell intrinsic transcriptional features. Through this approach, we identify five CRC intrinsic subtypes (CRIS) endowed with distinctive molecular, functional and phenotypic peculiarities: (i) CRIS-A: mucinous, glycolytic, enriched for microsatellite instability or KRAS mutations; (ii) CRIS-B: TGF-β pathway activity, epithelial–mesenchymal transition, poor prognosis; (iii) CRIS-C: elevated EGFR signalling, sensitivity to EGFR inhibitors; (iv) CRIS-D: WNT activation, IGF2 gene overexpression and amplification; and (v) CRIS-E: Paneth cell-like phenotype, TP53 mutations. CRIS subtypes successfully categorize independent sets of primary and metastatic CRCs, with limited overlap on existing transcriptional classes and unprecedented predictive and prognostic performances.

Doxycycline was found to act synergistically with the antifungal fluconazole against Candida albicans. Combination with doxycycline converts fluconazole from fungistatic to fungicidal, prevents the onset of drug resistance, and is also effective against a clinical isolate characterized by elevated resistance to fluconazole. Investigation of the interactions between the two drugs by way of checkerboard assays indicated that doxycycline had an influence on the MIC for fluconazole, as defined by CLSI standards, only at high concentrations (200 g/ml). However, lower concentrations were effective at eliminating residual cell growth at supra-MICs of fluconazole. Using MIC-0, defined as a drug combination resulting in optically clear wells, as an endpoint, doxycycline was found to be synergistic with fluconazole at a concentration as low as 25 g/ml, with a fractional inhibitory concentration index of <0.5. Doxycycline-mediated growth inhibition can be reversed by externally added iron, indicating that iron depletion may account for the synergism. Consistently, we confirmed old literature data about iron-chelating activity of doxycycline. Synergism of fluconazole with doxycycline does not appear to be mediated by calcineurin, since doxycycline further aggravates the susceptibility to fluconazole of mutants lacking the catalytic or the regulatory subunits of calcineurin. Growth in the presence of fluconazole and doxycycline is restored by an elevated dosage of ERG11 in Saccharomyces cerevisiae but not in C. albicans, despite the full competence of the pathogen's protein to act as a suppressor in baker's yeast. Infections by Candida albicans have been on the rise in the past decades, mostly due to changes in the clinical practice, such as the increased use of immunosuppressants and broad-spectrum antibacterial agents (26,29,47). Fungal infections represent a challenge for clinicians, because of the scarcity and sometimes the limited efficacy of antifungal drugs (1, 44). Fluconazole and other azole antifungals, targeting ergosterol biosynthesis (23), have been extensively used in the recent clinical practice thanks to their reduced toxicity and lower cost, compared to amphotericin B in its conventional and lipidic forms, respectively (44). The major disadvantage of azole antifungals, however, is their fungistatic nature, an aspect that favors the onset of drug resistance. The development of isogenic strains of C. albicans characterized by stepwise increased tolerance to fluconazole in patients undergoing continued treatment with this drug has been documented in several cases (34,56,57). For these reasons, the conversion of fluconazole from fungistatic to fungicidal via combinations with other drugs is highly desirable.In a different perspective, tools for the detailed study of the molecular genetics of C. albicans have been greatly improved in recent times (6,7,40). Regulatable promoters are available to the research community, to induce or repress gene expression in C. albicans. Most of them, such as the PCK1 and the MAL2...

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