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AbstractDNA damage occurs throughout life from a variety of sources, and it is imperative to repair damage in a timely manner to maintain genome stability. Thus, DNA repair mechanisms are a fundamental part of life. Nucleotide excision repair (NER) plays an important role in the removal of bulky DNA adducts, such as cyclobutane pyrimidine dimers from ultraviolet light or DNA crosslinking damage from platinum‐based chemotherapeutics, such as cisplatin. A main component for the NER pathway is transcription factor IIH (TFIIH), a multifunctional, 10‐subunit protein complex with crucial roles in both transcription and NER. In transcription, TFIIH is a component of the pre‐initiation complex and is important for promoter opening and the phosphorylation of RNA Polymerase II (RNA Pol II). During repair, TFIIH is important for DNA unwinding, recruitment of downstream repair factors, and verification of the bulky lesion. Several different disease states can arise from mutations within subunits of the TFIIH complex. Most strikingly are xeroderma pigmentosum (XP), XP combined with Cockayne syndrome (CS), and trichothiodystrophy (TTD). Here, we summarize the recruitment and functions of TFIIH in the two NER subpathways, global genomic (GG‐NER) and transcription‐coupled NER (TC‐NER). We will also discuss how TFIIH's roles in the two subpathways lead to different genetic disorders. Show less

Title: Mitochondrial Viscosity Probes: Iridium(III) Complexes Induce Apoptosis in HeLa Cells. Abstract: Mitochondrial viscosity has emerged as a promising biomarker for diseases such as cancer and neurodegenerative disorders, yet accurately measuring viscosity at the subcellular level remains a significant challenge. In this study, we synthesized and characterized three cyclometalated iridium(III) complexes (Ir1-Ir3) containing 5-fluorouracil derivatives as ligands. Among these, Ir1 selectively induced apoptosis in HeLa cells by increasing mitochondrial production of reactive oxygen species (ROS), which triggered a cascade of events leading to mitochondrial dysfunction. Additionally, the fluorescence lifetime of Ir1 demonstrated high sensitivity to intracellular viscosity changes, enabling real-time fluorescence lifetime imaging microscopy (FLIM) of cellular micro-viscosity during apoptosis. These findings underscore the potential of cyclometalated Ir(III) complexes for both therapeutic and diagnostic applications at the subcellular level. Show less

In this contribution we report the synthesis, characterization and in vitro anticancer activity of novel cyclometalated 4-phenylthiazole-derived ruthenium(II) (2a-e) and osmium(II) (3a-e) complexes. Formation and sufficient purity of the complexes were unambigiously confirmed by ¹H-, ¹³C- and 2D-NMR techniques, X-ray diffractometry, HRMS and elemental analysis. The binding preferences of these cyclometalates to selected amino acids and to DNA models including G-quadruplex structures were analyzed. Additionally, their stability and behaviour in aqueous solutions was determined by UV-Vis spectroscopy. Their cellular accumulation, their ability of inducing apoptosis, as well as their interference in the cell cycle were studied in SW480 colon cancer cells. The anticancer potencies were investigated in three human cancer cell lines and revealed IC₅₀ values in the low micromolar range, in contrast to the biologically inactive ligands. Show less

A series of eight N‐heterocyclic carbenes (NHC) gold(I) complexes, involving 1,3‐bis(2,6‐diisopropylphenyl)imidazol‐2‐ylidene (iPr) ligand in combination with 6‐mercaptopurine derivatives (HL1–8), has been prepared and thoroughly characterized, including elemental analysis, mass spectrometry, infrared and multinuclear NMR spectroscopy, and single crystal X‐ray analysis. The complexes, showing general composition of [Au (iPr)(Ln)] 1–8, were evaluated for their in vitro cytotoxicity against four human cancer cell lines including A2780 (ovarian) and A2780R (ovarian Cisplatin resistant), PC3 (prostate) and MCF‐7 (breast), and normal human MRC‐5 cells (lung fibroblasts). The complexes revealed significant cytotoxicity, with the best IC50 values ≈ 3.4–6.4 μM against A2780 and reasonable selectivity. Cellular effects of the selected complexes on the A2780 cells were evaluated using various flow cytometry assays. Complexes 1, 3, and 4 showed a strong pro‐apoptotic effect and a strong effect on the loss of mitochondrial membrane potential. These findings indicate that their major mechanism of action is based on the collapse of the mitochondrial metabolism and activation of the intrinsic signaling pathway of apoptosis, consequently resulting in cell death. The complexes 1–8 revealed only negligible effect on the production of inflammatory‐related cytokine (TNF‐α), as well as the activation of nuclear factor kappa‐light‐chain‐enhancer of activated B cells (NF‐κB) or peroxisome proliferator‐activated receptor gamma (PPARγ). Moreover, the shotgun proteomic analysis was performed, and the obtained results suggest that the mechanism of action of complexes 1, 3, and 4 differs somewhat from that of Auranofin. Show less

Abstract The review is devoted to copper coordination compounds based on 2,2′-bipyridine / 1,10-phenanthroline and diverse N-, O-, S-donor ligands exhibiting cytotoxic properties. Therefore, they can be a starting platform for developing antitumor drugs. The review covers the structural aspects of the complexes, the features of their cytotoxic activity and its mechanism, as well as in vivo studies. Show less

Human Replication Protein A (RPA) was historically discovered as one of the six components needed to reconstitute simian virus 40 DNA replication from purified components. RPA is now known to be involved in all DNA metabolism pathways that involve single-stranded DNA (ssDNA). Heterotrimeric RPA comprises several domains connected by flexible linkers and is heavily regulated by post-translational modifications (PTMs). The structure of RPA has been challenging to obtain. Various structural methods have been applied, but a complete understanding of RPA's flexible structure, its function, and how it is regulated by PTMs has yet to be obtained. This review will summarize recent literature concerning how RPA is phosphorylated in the cell cycle, the structural analysis of RPA, DNA and protein interactions involving RPA, and how PTMs regulate RPA activity and complex formation in double-strand break repair. There are many holes in our understanding of this research area. We will conclude with perspectives for future research on how RPA PTMs control double-strand break repair in the cell cycle. Show less

Ovarian cancer represents a leading cause of cancer-related deaths in women worldwide. Chemotherapeutic agents are usually employed to treat the patients, and Ruthenium(II)-based compounds have been investigated as possible substitutes for platinum drugs. In this work, we studied three different Ru(II)-phosphine-mercapto complexes (1-3) as potential cytotoxic agents against A2780 and A2780-cisR ovarian cancer cells. A time-dependent cytotoxicity was observed for 2, which also exhibited better selectivity than cisplatin control. A similar cytotoxic behavior was observed on 3D tumor spheroids. Although no changes were observed in cell cycle distribution, compound 2 affected the mitochondrial membrane potential on A2780 cells, and caused cell death via apoptotic pathway, which was confirmed by flow cytometry assay. Western blotting experiments revealed that 2 affected the expression of p53, PCNA, γH2AX and cleaved caspase-3, making it a promising anticancer agent for ovarian cancer. Show less

Immunogenic cell death (ICD), which results from insufficient cellular adaptation to specific stressors, occupies a central position in the development of novel anticancer treatments. Several therapeutic strategies to elicit ICD - either as standalone approaches or as means to convert immunologically cold tumours that are insensitive to immunotherapy into hot and immunotherapy-sensitive lesions - are being actively pursued. However, the development of ICD-inducing treatments is hindered by various obstacles. Some of these relate to the intrinsic complexity of cancer cell biology, whereas others arise from the use of conventional therapeutic strategies that were developed according to immune-agnostic principles. Moreover, current discovery platforms for the development of novel ICD inducers suffer from limitations that must be addressed to improve bench-to-bedside translational efforts. An improved appreciation of the conceptual difference between key factors that discriminate distinct forms of cell death will assist the design of clinically viable ICD inducers. Show less

Title: Cyrene™ as a green alternative to Abstract: Ruthenium(II) polypyridyl complexes (RPCs) that emit from triplet metal-to-ligand charge transfer (MLCT) states find a wide variety of uses ranging from luminophores to potential anti-cancer or anti-bacterial therapeutics. Herein we describe a greener, microwave-assisted synthetic pathway for the preparation of homoleptic [Ru(N^N)3]2+ and bis-heteroleptic [Ru(N^N)2(N'^N')]2+ type complexes. This employs the bio-renewable solvent Cyrene™, dihydrolevoglucosenone, as a green alternative to N,N'-dimethylformamide (DMF) in the synthesis of Ru(N^N)2Cl2 intermediate complexes, obtaining comparable yields for N^N = 2,2'-bipyridine, 1,10-phenanthroline and methylated derivatives. Employing these intermediates, a range of RPCs were prepared and we verify that the ubiquitous luminophore [Ru(bpy)3]2+ (bpy = 2,2'-bipyridine) can be prepared by this two-step green pathway where it is virtually indistinguishable from a commercial reference. Furthermore, the novel complexes [Ru(bpy)2(10,11-dmdppz)]2+ (10,11-dmdppz = 10,11-dimethyl-dipyridophenazine) and [Ru(5,5'-dmbpy)2(10,11-dmdppz)]2+ (5,5'-dmbpy = 5,5'-dimethyl-bpy) intercalate duplex DNA with high affinity (DNA binding constants, Kb = 5.7 × 107 and 1.0 × 107 M-1, respectively) and function as plasma membrane and nuclear DNA dyes for confocal and STED microscopies courtesy of their long-lived MLCT luminescence. Show less

Cisplatin (cDDP) resistance is a matter of concern in triple-negative breast cancer therapeutics. We measured the metabolic response of cDDP-sensitive (S) and -resistant (R) MDA-MB-231 cells to Pd2Spermine(Spm) (a possible alternative to cDDP) compared to cDDP to investigate (i) intrinsic response/resistance mechanisms and (ii) the potential cytotoxic role of Pd2Spm. Cell extracts were analyzed by untargeted nuclear magnetic resonance metabolomics, and cell media were analyzed for particular metabolites. CDDP-exposed S cells experienced enhanced antioxidant protection and small deviations in the tricarboxylic acid cycle (TCA), pyrimidine metabolism, and lipid oxidation (proposed cytotoxicity signature). R cells responded more strongly to cDDP, suggesting a resistance signature of activated TCA cycle, altered AMP/ADP/ATP and adenine/uracil fingerprints, and phospholipid biosynthesis (without significant antioxidant protection). Pd2Spm impacted more markedly on R/S cell metabolisms, inducing similarities to cDDP/S cells (probably reflecting high cytotoxicity) and strong additional effects indicative of amino acid depletion, membrane degradation, energy/nucleotide adaptations, and a possible beneficial intracellular γ-aminobutyrate/glutathione-mediated antioxidant mechanism. Show less

This study investigates the activity of novel gold(I) and copper(I)/zinc(II) heteronuclear complexes against colon cancer. The synthesised heteronuclear Au(I)-Cu(I) and Au(I)-Zn(II) complexes were characterised and evaluated for their anticancer activity using human colon cancer cell lines (Caco-2). The complexes exhibited potent cytotoxicity, with IC₅₀ values in the low micromolar range, and effectively induced apoptosis in cancer cells. In the case of complex [Cu{Au(Spy)(PTA)}₂]PF₆ (2), its cytotoxicity is ×10 higher than its mononuclear precursor, while showing low cytotoxicity towards differentiated healthy cells. Mechanistic studies revealed that complex 2 inhibits the activity of thioredoxin reductase, a key enzyme involved in redox regulation, leading to an increase in reactive oxygen species (ROS) levels and oxidative stress, in addition to an alteration in DNA's tertiary structure. Furthermore, the complexes demonstrated a strong binding affinity to bovine serum albumin (BSA), suggesting the potential for effective drug delivery and bioavailability. Collectively, these findings highlight the potential of the investigated heteronuclear Au(I)-Cu(I) and Au(I)-Zn(II) complexes as promising anticancer agents, particularly against colon cancer, through their ability to disrupt redox homeostasis and induce oxidative stress-mediated cell death. Show less

Purpose

Platinum-based drugs are cytotoxic drugs commonly used in cancer treatment. They cause DNA damage, effects of which on chromatin and cellular responses are relatively well described. Yet, the nuclear stress responses related to RNA processing are incompletely known and may be relevant for the heterogeneity with which cancer cells respond to these drugs. Here, we determine the type and extent of nuclear stress responses of prostate cancer cells to clinically relevant platinum drugs.

Methods

We study nucleolar and Cajal body (CB) responses to cisplatin, carboplatin, and oxaliplatin with immunofluorescence-based methods in prostate cancer cells. We utilize organelle-specific markers NPM, Fibrillarin, Coilin, and SMN1, and study CB-regulatory proteins FUS and TDP-43 using siRNA-mediated downregulation.

Results

Different types of prostate cancer cells have different sensitivities to platinum drugs. With equally cytotoxic doses, cisplatin, and oxaliplatin induce prominent nucleolar and CB stress responses while the nuclear stress phenotypes to carboplatin are milder. We find that Coilin is a stress-specific marker for platinum drug response heterogeneity. We also find that CB-associated, stress-responsive RNA binding proteins FUS and TDP-43 control Coilin and CB biology in prostate cancer cells and, further, that TDP-43 is associated with stress-responsive CBs in prostate cancer cells.

Conclusion

Our findings provide insight into the heterologous responses of prostate cancer cells to different platinum drug treatments and indicate Coilin and TDP-43 as stress mediators in the varied outcomes. These results help understand cancer drug responses at a cellular level and have implications in tackling heterogeneity in cancer treatment outcomes. Show less

Abstract Solute Carrier Family 25 (SLC25) is the largest family of mitochondrial membrane proteins in the human body, consisting of 53 members. Mitochondrial phosphate carriers (MPiC), cellular iron metabolism, voltage-dependent anion channels (VDAC), and oxidative phosphorylation in the SLC25 family play dominant roles in material transport, energy metabolism, etc. SLC25 family-related proteins are involved in the regulation of the progression of a variety of cancers, including colon, gastric, and lung cancers. In addition, the SLC25 family has been implicated in endoplasmic reticulum stress (ERS) and immunity. Since SLC25 family proteins are involved in cancer progression and are associated with endoplasmic reticulum stress and immunity, exploring inhibitors of SLC25 family-related proteins is essential. However, the exact mechanism of SLC25 family-related proteins involved in cancer, as well as potential targets and SLC25 inhibitors have not been reported in the literature. This article focuses on summarizing the relevance of the SLC25 family to cancer, ERS, and immunity. This review also provides a comprehensive overview of SLC25 family-related inhibitors. Show less

PandaOmics is a cloud-based software platform that applies artificial intelligence and bioinformatics techniques to multimodal omics and biomedical text data for therapeutic target and biomarker discovery. PandaOmics generates novel and repurposed therapeutic target and biomarker hypotheses with the desired properties and is available through licensing or collaboration. Targets and biomarkers generated by the platform were previously validated in both in vitro and in vivo studies. PandaOmics is a core component of Insilico Medicine's Pharma.ai drug discovery suite, which also includes Chemistry42 for the de novo generation of novel small molecules, and inClinico─a data-driven multimodal platform that forecasts a clinical trial's probability of successful transition from phase 2 to phase 3. In this paper, we demonstrate how the PandaOmics platform can efficiently identify novel molecular targets and biomarkers for various diseases. Show less

Nucleophosmin (NPM1) is a key nucleolar protein released from the nucleolus in response to stress stimuli. NPM1 functions as a stress regulator with nucleic acid and protein chaperone activities, rapidly shuttling between the nucleus and cytoplasm. NPM1 is ubiquitously expressed in tissues and can be found in the nucleolus, nucleoplasm, cytoplasm, and extracellular environment. It plays a central role in various biological processes such as ribosome biogenesis, cell cycle regulation, cell proliferation, DNA damage repair, and apoptosis. In addition, it is highly expressed in cancer cells and solid tumors, and its mutation is a major cause of acute myeloid leukemia (AML). This review focuses on NPM1's structural features, functional diversity, subcellular distribution, and role in stress modulation. Show less

Lung cancer is a leading cause of cancer-related death worldwide that needs updated therapies to contrast both the serious side effects and the occurrence of drug resistance. A panel of non-small cell lung cancer (NSCLC) cells were herein employed as cancer models. Eight structurally related gold(I) and gold(III) complexes with NHC and halides or triphenylphosphane ligands were investigated as lung cancer cell growth inhibitors. As expected, gold compounds with PPh3 were found to be more cytotoxic than homoleptic [(NHC)2-Au(I)]X or heteroleptic NHC-Au(I)X or NHC-Au(III)X3 complexes. Mixed ligand gold(I) compounds exhibiting the linear NHC-AuPPh3 (compound 7) or the trigonal NHC-Au(Cl)PPh3 (compound 8) arrangements at the central metal were found to be the best lung cancer cytotoxic compounds. Analysis of the TrxR residual activity of the treated cells revealed that these compounds efficiently inhibit the most accredited molecular target for gold compounds, the TrxR, with compound 8 reaching more than 80% activity reduction in lung cells. Some of the current cancer lung therapy protocols consist of specific lung cancer cell cytotoxic agents combined with antifolate drugs; interestingly, the herein gold compounds are both TrxR and antifolate inhibitors. The human DHFR was inhibited with IC50 ranging between 10–21 µM, depending on substrate concentrations, proceeding by a likely allosteric mechanism only for compound 8. Show less

Title: Analysis of antiproliferative activity of new half-sandwich arene Ru(II) thiophene based aroylhydrazone complexes. Abstract: Efforts in researching the efficient anti-tumor properties of three novel arene ruthenium(II) complexes incorporating thiophene-based aroylhydrazone ligands have been undertaken. The complexes' elemental composition was [(η6-p-cymene)Ru(L)Cl]. They were comprehensively characterized through elemental and spectroscopic analyses (FT-IR, UV-vis, NMR, and HR-MS). Single crystal X-ray diffraction studies revealed a pseudo-octahedral geometry with bidentate coordination of the ligands in a representative complex. The in vitro assessment of the complexes' cancer cell growth inhibition was conducted using the MTT assay against A549 (human lung carcinoma), HeLa (human cervical carcinoma), HuH-7 (hepatocellular carcinoma), and NIH-3T3 (mouse fibroblast non-cancerous cell line). Results indicated significant cytotoxicity across all cancer cell lines, with IC50 concentrations of complex 2 being 6.8 μM for A549, 11.6 μM for HeLa, and 9.4 μM for HuH-7, compared to cisplatin with IC50 values of 18.9 μM, 17.68 μM, and 24 μM respectively. Notably, complex 2 demonstrated particularly promising cytotoxicity against all tested cancerous cell lines. Fluorescent staining analysis such as acridine orange/ethidium bromide (AO-EB) and HOECHST 33342 revealed cell death mechanisms involving membrane disintegration and nuclear condensation following treatment with complex 2. Further studies were conducted to measure reactive oxygen species (ROS) levels using the dichlorodihydrofluorescein diacetate (DCFH-DA) assay, and mitochondrial membrane potential (MMP) was assessed using the JC-1 dye assay. These studies demonstrated that complex 2 increased ROS levels, decreased membrane potential, and promoted mitochondrial dysfunction-mediated cell death pathways. Additionally, flow cytometry analysis, utilizing dual staining of Annexin V-FITC and propidium iodide (PI), was employed to quantitatively study apoptosis induction. Show less

Iridium(III) complexes nowadays became rising stars in various health-related applications. Thus, there is a necessity to assess cytotoxicity of the synthesized molecules against cancer/normal cell lines. In this report, we present a dataset of 2694 experimental cytotoxicity values of 803 iridium complexes against 127 different cell lines. We specify the experimental conditions and provide representation of the complexes molecules in machine-readable format. The dataset provides a starting point for exploration of new iridium-based cellular probes and opens new possibilities for predictions of toxicities and data-driven generation of new organometallic anticancer agents. Show less

Oxidation-reduction (redox) reactions are central to the existence of life. Reactive species of oxygen, nitrogen and sulfur mediate redox control of a wide range of essential cellular processes. Yet, excessive levels of oxidants are associated with ageing and many diseases, including cardiological and neurodegenerative diseases, and cancer. Hence, maintaining the fine-tuned steady-state balance of reactive species production and removal is essential. Here, we discuss new insights into the dynamic maintenance of redox homeostasis (that is, redox homeodynamics) and the principles underlying biological redox organization, termed the 'redox code'. We survey how redox changes result in stress responses by hormesis mechanisms, and how the lifelong cumulative exposure to environmental agents, termed the 'exposome', is communicated to cells through redox signals. Better understanding of the molecular and cellular basis of redox biology will guide novel redox medicine approaches aimed at preventing and treating diseases associated with disturbed redox regulation. Show less

Cancer stem cells (CSCs) are defined as a rare population of cancer cells related to tumor initiation and maintenance. These cells are primarily responsible for tumor growth, invasion, metastasis, recurrence, and resistance to chemotherapy. In this paper, we demonstrated the ability of Ru(II)-based complexes containing 2-thiouracil derivatives with the chemical formulas trans-[Ru(2TU)(PPh₃)₂(bipy)]PF₆ (1) and trans-[Ru(6m2TU)(PPh₃)₂(bipy)]PF₆ (2) (where 2TU = 2-thiouracil and 6m2TU = 6-methyl-2-thiouracil) to suppress liver CSCs by targeting NF-κB and Akt/mTOR signaling. Complexes 1 and 2 displayed potent cytotoxic effects on cancer cell lines and suppressed liver CSCs from HepG2 cells. Increased phosphatidylserine exposure, loss of mitochondrial transmembrane potential, increased PARP (Asp214) cleavage, DNA fragmentation, chromatin condensation and cytoplasmic shrinkage were detected in HepG2 cells treated with these complexes. Mechanistically, complexes 1 and 2 target NF-κB and Akt/mTOR signaling in HepG2 cells. Cell motility inhibition was also detected in HepG2 cells treated with these complexes. Complexes 1 and 2 also inhibited tumor progression in mice with HepG2 cell xenografts and exhibited tolerable systemic toxicity. Taken together, these results indicate that these complexes are new anti-HCC drug candidates that can suppress liver CSCs. Show less