Anna Messere

A new ferrocenemethyl-thymidine nucleoside: Synthesis, incorporation into oligonucleotides and optical spectroscopic studies on the resulting single strand, duplex and triplex structures

Abstract A new thymidine analogue, bearing a ferrocenemethyl residue at the N-3 position of the base, was synthesized in high yields via Mitsunobu reaction of ferrocenemethanol with sugar protected thymidine, converted into the corresponding 3′-phosphoramidite and incorporated into oligonucleotides. Duplex and triplex formation experiments, evaluated by UV and CD spectroscopy, showed a dramatic decrease of the affinity towards complementary single strands, while for triplexes, the introduction of a ferrocene residue in the third strand resulted in higher melting temperatures, associated with a reduced content of triplex structure.

Long non-coding RNA containing ultraconserved genomic region 8 promotes bladder cancer tumorigenesis.

Ultraconserved regions (UCRs) have been shown to originate non-coding RNA transcripts (T-UCRs) that have different expression profiles and play functional roles in the pathophysiology of multiple cancers. The relevance of these functions to the pathogenesis of bladder cancer (BlCa) is speculative. To elucidate this relevance, we first used genome-wide profiling to evaluate the expression of T-UCRs in BlCa tissues. Analysis of two datasets comprising normal bladder tissues and BlCa specimens with a custom T-UCR microarray identified ultraconserved RNA (uc.) 8+ as the most upregulated T-UCR in BlCa tissues, although its expression was lower than in pericancerous bladder tissues. These results were confirmed on BlCa tissues by real-time PCR and by in situ hybridization. Although uc.8+ is located within intron 1 of CASZ1, a zinc-finger transcription factor, the transcribed non-coding RNA encoding uc.8+ is expressed independently of CASZ1. In vitro experiments evaluating the effects of uc.8+ silencing, showed significantly decreased capacities for cancer cell invasion, migration, and proliferation. From this, we proposed and validated a model of interaction in which uc.8+ shuttles from the nucleus to the cytoplasm of BlCa cells, interacts with microRNA (miR)-596, and cooperates in the promotion and development of BlCa. Using computational analysis, we investigated the miR-binding domain accessibility, as determined by base-pairing interactions within the uc.8+ predicted secondary structure, RNA binding affinity, and RNA species abundance in bladder tissues and showed that uc.8+ is a natural decoy for miR-596. Thus uc.8+ upregulation results in increased expression of MMP9, increasing the invasive potential of BlCa cells. These interactions between evolutionarily conserved regions of DNA suggest that natural selection has preserved this potentially regulatory layer that uses RNA to modulate miR levels, opening up the possibility for development of useful markers for early diagnosis and prognosis as well as for development of new RNA-based cancer therapies.

Tuning RNA Interference by Enhancing siRNA/PAZ Recognition.

Chemically modified siRNAs were synthesized to enhance the corresponding silencing activities. The introduced modifications endowed siRNAs with high silencing effect, long RNAi persistence, and better serum resistance. Theoretical data allowed us to correlate the observed siRNAs interfering performance with the peculiar interactions with PAZ.

Nitration of Cinnamic Acids Using Cerium (IV) Ammonium Nitrate Immobilized on Silica

Abstract Treatment of cinnamic acids with cerium (IV) ammonium nitrate supported on silica (CAN/SiO2) was used to synthesize nitro derivative and ipso substitution products. The position of the substituents defines the type and the yields of the products. This is the first example of an ipso substitution reaction by solid‐phase approach.

A novel synthetic strategy for monosubstituted cyclodextrin derivatives

A first solid phase approach to obtain monosubstituted CD conjugates to different labels has been developed. A new solid support has been designed to get a variety of C-6 monofunctionalized CDs (α, β, MeβCD and HPβCD) covalently linked through a phosphodiester bridge to different labels, in highly pure form and under very mild detachment conditions.

SYNTHESIS AND CHARACTERIZATION OF NEW 3'-3' LINKED OLIGODEOXYRIBONUCLEOTIDES FOR ALTERNATE STRAND TRIPLE HELIX FORMATION

Abstract Protected forms of 1,2,3-propanetriol andcis, cis-1,3,5-cyclohexanetriol were incorporated onto solid supports which were exploited in the solid phase synthesis of 3′-3′ linked oligodeoxyribonucleotides (ODNs), involving only nucleoside 3′-phosphoramidites as building blocks, UV thermal denaturation analysis showed the ability of ODNs with this inversion of polarity motif to cooperatively hybridize with duplexes of the type 5′-(Pu)m(Py)n-3′ in an alternate strand recognition approach. Download : Download full-size image

A Facile Solid-Phase Strategy for the Synthesis of Oligonucleotide-Tetraphenylporphyrin Conjugates

A convenient solid-phase synthesis of oligonucleotides conjugated at the 3′-end with a tetraphenylporphyrin residue, by means of a new polymeric support bearing as a linker a lysine derivative, has been developed. A porphyrin linked 17-mer, designed for antisense experiments, has been prepared in good yields, and its hybridization properties with a complementary DNA fragment evaluated by UV thermal analysis.

1-Substituted 2′-deoxyinosine analogues

The base 2-carbon of 2′,3′-di-O-acetyl-2′-deoxyinosine is strongly activated towards nucleophilic attack when either the 4-nitrophenyl or 2,4-dinitrophenyl group is attached to its N-1 position (product 1 or 2). 1-(ω-Aminoalkyl)- and 1-(ω-hydroxyalkyl)-2′-deoxyinosine derivatives 5, 8–10 have been efficiently synthesized by a rearrangement of the purine ring upon treatment of compound 1 or 2 with the appropriate α,ω-diamine or α,ω-hydroxyamine. Moreover 1-amino-2′-deoxyinosine 11 and 1-hydroxy-2′-deoxyinosine 13 have been easily prepared in high yields by reaction of substrate 1 or 2, respectively, with hydrazine or hydroxylamine.

PNA-DNA Chimeras Forming Quadruplex Structures

Abstract 1H-NMR, CD, and UV spectroscopy have been used to investigate the structure of PNA/DNA chimeras forming quadruplex structures. In particular, we synthesized 5′TGGG3′-t (1) and 5′TGG3′-gt (2), where lower and upper case letters indicate PNA and DNA residues, respectively. CD spectrum and all NMR data of (1) are typical of quadruplexes involving four parallel strands. UV melting profile of (1) indicates that its thermal stability is quite similar to that observed for the reference structure [d(TGGGT)]4. 1H-NMR spectrum for 5′TGG3′-gt (2) shows that this oligonucleotide is not able to fold into a single, well-defined species.

Synthesis and Biological Properties of Caffeic Acid-PNA Dimers Containing Guanine

Caffeic acid (CA; 3,4-dihydroxycinnamic acid) is endowed with high antioxidant activity. CA derivatives (such as amides) have gained a lot of attention due to their antioxidative, antitumor and antimicrobial properties as well as stable characteristics. Caffeoyl-peptide derivatives showed different antioxidant activity depending on the type and the sequence of amino acid used. For these reasons, we decided to combine CA with Peptide Nucleic Acid (PNA) to test whether the new PNA-CA amide derivatives would result in an improvement or gain of CA’s biological (i.e., antioxidant, cytotoxic, cytoprotective) properties. We performed the synthesis and characterization of seven dimer conjugates with various combinations of nucleic acid bases and focused NMR studies on the model compound ga-CA dimer. We demonstrate that PNA dimers containing guanine conjugated to CA exhibited different biological activities depending on composition and sequence of the nucleobases. The dimer ag-CA protected HepG2, SK-B-NE(2), and C6 cells from a cytotoxic dose of hydrogen peroxide (H2O2).