Characterization of the bisintercalative DNA binding mode of a bifunctional platinum–acridine agent

The DNA interactions of PT-BIS(ACRAMTU) ([Pt(en)(ACRAMTU)(2)](NO(3))(4); ACRAMTU = 1-[2-(acridin-9-ylamino)ethyl]-1,3-dimethylthiourea, en = ethylenediamine), a bifunctional platinum–acridine conjugate, have been studied in native and synthetic double-stranded DNAs and model duplexes using various biophysical techniques. These include ethidium-DNA fluorescence quenching and thermal melting experiments, circular dichroism (CD) spectroscopy and plasmid unwinding assays. In addition, the binding mode was studied in a short octamer by NMR spectroscopy in conjunction with molecular modeling. In alternating copolymers, PT-BIS(ACRAMTU) shows a distinct preference for poly(dA-dT)(2), which is ∼3-fold higher than that of ACRAMTU. In the ligand-oligomer complex, d(GCTATAGC)(2)·PT-BIS(ACRAMTU) (complex I*), PT-BIS(ACRAMTU) increases the thermal stability of the B-form host duplex by ΔT(m) > 30 K (CD and UV melting experiments). The agent unwinds pSP73 plasmid DNA by 44(±2)° per bound molecule, indicating bisintercalative binding. A 2-D NMR study unequivocally demonstrates that PT-BIS(ACRAMTU)'s chromophores deeply bisintercalate into the 5′-TA/TA base pair steps in I*, while the platinum linker lies in the minor groove. An AMBER model reflecting the NMR results shows that bracketing of the central AT base pairs in a classical nearest neighbor excluded fashion is feasible. PT-BIS(ACRAMTU) inhibits DNA hydrolysis by BstZ17 I at the enzyme's restriction site, GTA↓TAC. Possible consequences for other relevant DNA–protein interactions, such as those involved in TATA-box-mediated transcription initiation and the utility of the platinum-intercalator technology for the design of sequence-specific agents are discussed

Osteoarthritis of the knee – clinical assessments and inflammatory markers11Supported by a grant from the Robert Bosch Stiftung, Stuttgart, Germany.

AbstractObjective: The present cross sectional study was performed to test the hypothesis that in osteoarthritis (OA) of the knee severity of this disease is related to local levels of inflammatory metabolites and their corresponding enzymes.Methods: From 41 patients with OA of the knee (age range 45–79 years) undergoing arthroscopy blood, synovial fluid (SF) and synovial membrane (SM) were collected. Clinical conditions were primarily assessed by the WOMAC-index and radiographic grading (K&L-grade). Concentrations of PGE2, TxB2and NO2/3and that of IL-6, IL-1α, IL-1β, TNFα, COX-2 and iNOS were determined in SF and SM, respectively.Results: With advancing age K&L-grade and COX-2 in SM increased significantly (P=0.005 and P=0.01, respectively). TNFα and IL-1α were not detectable in SM samples. Apart from a correlation between PGE2and WOMAC-index (r=0.36, P=0.035) no significant relationships could be found between the various inflammatory parameters and any of the assessed clinical signs.Conclusions: Apparently no direct relationships exist between the measured markers of inflammation (e.g. PGE2, NO2/3) or the involved enzymes (e.g. COX-2, iNOS) and the severity of OA of the knee. The degenerative condition of this disease might be due to the more local, mainly mechanical injury with little systemic upset. However, further longitudinal studies are needed to clarify whether the assessed biochemical markers could serve as predictors for the progression of OA

Zirconium tetraazamacrocycle complexes display extraordinary stability and provide a new strategy for zirconium-89-based radiopharmaceutical development

89 Zr–Tetraazamacrocycle complexes display extraordinary stability

Chemosensitivity–Gene Expression Correlations and Functional Enrichment Analysis Provide Insight into the Mechanism of Action of a Platinum–Acridine Anticancer Agent

Growth inhibition (GI) and gene expression profiles across the 60 cell lines of the NCI-60 screen were analyzed to elucidate mechanistic differences between a nucleolus-targeting platinum-acridine (PA) hybrid anticancer agent (compound 1) and the standard-of-care chemotherapies cisplatin, oxaliplatin, mitomycin c, doxorubicin, and topotecan. The study utilized Pearson correlation and functional enrichment analysis tools (including gene set enrichment analysis, GSEA) in combination with the gene ontology (GO) resource to identify cellular processes that might contribute to the unique spectrum of activity and high potency of compound 1. The GSEA results are consistent with DNA being a major target of compound 1 based on the negative correlation observed between its potency and expression levels of genes implicated in DNA double-strand break (DSB) repair. GO terms related to RNA processing, including ribosome biogenesis and RNA splicing, are also negatively enriched in GSEA for compound 1, suggesting a mechanism by which these processes render cells more resistant to the hybrid agent. The opposite trend is observed for the other DNA-targeted drugs. Significant functional interactions (STRING) exist between genes/gene products involved in ribosome biogenesis and DSB repair in rDNA, including BRCA1, BABAM1, RPA2, MDM2, RAD21, RAD54L, RPL5, and PAK1IP1. The results suggest that high levels of ribosome biogenesis may buffer compound 1’s cytotoxicity by triggering the ribosomal protein (RPL5)-MDM2-p53 surveillance pathway to facilitate downstream repair of DSBs in rDNA as a nucleolar stress response

A Membrane Transporter Determines the Spectrum of Activity of a Potent DNA-Targeted Hybrid Anticancer Agent

Cytotoxic drugs that are mechanistically distinct from current chemotherapies are attractive components of personalized combination regimens for combating aggressive forms of cancer. To gain insight into the cellular mechanism of a highly potent platinum–acridine hybrid agent, we performed a correlation analysis of NCI-60 compound screening results and gene expression profiles. We discovered a plasma membrane transporter, human multidrug and toxin extrusion protein 1 (hMATE1, SLC47A1), as the dominant pan-cancer predictor for cancer cell chemosensitivity to the hybrid agent. We have validated the role of hMATE1 using transporter inhibition, gene knockdown, and chemical sensitization assays. The results suggest that hMATE1 may have applications as a molecular marker to identify and target tumors that are likely to respond to platinum–acridines. Furthermore, enhancement of hMATE1 expression by epigenetic drugs emerges as a potential co-treatment strategy to sensitize tumor tissue to platinum–acridines and other anticancer drugs transported by hMATE1.</i