Poor efficiency of chemotherapeutics in the eradication of Cancer Stem Cells (CSCs) has been driving the search for more active and specific compounds. In this work, we show how cell density-dependent stage culture profiles can be used in drug development workflows to achieve more robust drug activity (IC₅₀ and EC₅₀) results. Using flow cytometry and light microscopy, we characterized the cytological stage profiles of the HL-60-, A-549-, and HEK-293-derived sublines with a focus on their primitive cell content. We then used a range of cytotoxic substances—C-123, bortezomib, idarubicin, C-1305, doxorubicin, DMSO, and ethanol—to highlight typical density-related issues accompanying drug activity determination. We also showed that drug EC₅₀ and selectivity indices normalized to primitive cell content are more accurate activity measurements. We tested our approach by calculating the corrected selectivity index of a novel chemotherapeutic candidate, C-123. Overall, our study highlights the usefulness of accounting for primitive cell fractions in the assessment of drug efficiency.

Seven conjugates composed of well-known fluoroquinolone antibacterial agents, ciprofloxacin (CIP) or levofloxacin (LVX), and a cell-penetrating peptide transportan 10 (TP10-NH₂) were synthesised. The drugs were covalently bound to the peptide via an amide bond, methylenecarbonyl moiety, or a disulfide bridge. Conjugation of fluoroquinolones to TP10-NH₂ resulted in congeners demonstrating antifungal in vitro activity against human pathogenic yeasts of the Candida genus (MICs in the 6.25–100 µM range), whereas the components were poorly active. The antibacterial in vitro activity of most of the conjugates was lower than the activity of CIP or LVX, but the antibacterial effect of CIP-S-S-TP10-NH₂ was similar to the mother fluoroquinolone. Additionally, for two representative CIP and LVX conjugates, a rapid bactericidal effect was shown. Compared to fluoroquinolones, TP10-NH₂ and the majority of its conjugates generated a relatively low level of reactive oxygen species (ROS) in human embryonic kidney cells (HEK293) and human myeloid leukemia cells (HL-60). The conjugates exhibited cytotoxicity against three cell lines, HEK293, HepG2 (human liver cancer cell line), and LLC-PK1 (old male pig kidney cells), with IC₅₀ values in the 10–100 µM range and hemolytic activity. The mammalian toxicity was due to the intrinsic cytoplasmic membrane disruption activity of TP10-NH₂ since fluoroquinolones themselves were not cytotoxic. Nevertheless, the selectivity index values of the conjugates, both for the bacteria and human pathogenic yeasts, remained favourable.

In this review we explore innovative approaches in the treatment of hematologic cancers by combining various therapeutic modalities. We discuss the synergistic potential of combining inhibitors targeting different cellular pathways with immunotherapies, molecular therapies, and hormonal therapies. Examples include combining PI3K inhibitors with proteasome inhibitors, NF-κB inhibitors with immunotherapy checkpoint inhibitors, and neddylation inhibitors with therapies targeting the tumor microenvironment. Additionally, we discuss the potential use of small molecules and peptide inhibitors in hematologic cancer treatment. These multidimensional therapeutic combinations present promising strategies for enhancing treatment efficacy and overcoming resistance mechanisms. However, further clinical research is required to validate their effectiveness and safety profiles in hematologic cancer patients.

Infections of Candida spp. etiology are frequently treated with azole drugs. Among azoles, the most widely used in the clinical scenario remains fluconazole (FLC). Promising results in treatment of dangerous, systemic Candida infections demonstrate the advantages of combined therapies carried out with combinations of at least two different antifungal agents. Here, we report five conjugates composed of covalently linked FLC and cell penetrating or antimicrobial peptide: TP10-7-NH₂, TP10-NH₂, LFcinB(2-11)-NH₂, LFcinB[Nle^1,11]-NH₂, and HLopt2-NH₂, with aspects of design, chemical synthesis and their biological activities. Two of these compounds, namely FLCpOH-TP10-NH₂ and FLCpOH-TP10-7-NH₂, exhibit high activity against reference strains and fluconazole-resistant clinical isolates of C. albicans, including strains overproducing drug transporters. Moreover, both of them demonstrate higher fungicidal effects compared to fluconazole. Analysis performed with fluorescence and scanning electron microscopy as well as flow cytometry indicated the cell membrane as a molecular target of synthesized conjugates. An important advantage of FLCpOH-TP10-NH₂ and FLCpOH-TP10-7-NH₂ is their low cytotoxicity. The IC₉₀ value for the human cells after 72 h treatment was comparable to the MIC₅₀ value after 24 h treatment for most strains of C. albicans. In reported conjugates, FLC was linked to the peptide by its hydroxyl group. It is worth noting that conjugation of FLC by the nitrogen atom of the triazole ring led to practically inactive compounds. Two compounds produced by us and reported herein appear to be potential candidates for novel antifungal agents.