An inexpensive and much improved protocol has been developed for the synthesis of protected morpholino monomers from unprotected ribonucleosides in high overall yield, using oxidative glycol cleavage and reductive amination strategy. Unlike the previous methods, the present strategy allows installing the exocyclic amine protections at a later stage, and thus avoids the use of expensive, or commercially unavailable, exocyclic amine-protected ribonucleosides as starting materials. To demonstrate the flexibility of the present method in choosing protecting groups, the monomers have been protected with several such groups of different deblocking properties at the exocyclic amine position.

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Cationic guanidinium and phosphonium functionalized cytosine morpholino tetramer (G3) and trimer (P3) have been reported for the first time. They show efficient cellular uptake properties in full growth medium, quantified by fluorescence activated cell sorting (FACS) and visualized by fluorescence microscopy. An interesting feature of these transporters is their localization in the nucleolus or the rest of nucleus, determined by the nature of cationic moieties attached with cytosine base. Co-staining with MitoTracker Deep Red depicts their ability to penetrate mitochondrial membrane. Fluorescence imaging displays their uniform distribution throughout zebrafish larvae with no significant toxicity. FACS analysis at 4 8 C indicated that transfection of G3 transporter is energy dependent whereas P3 is energy independent. When covalently conjugated, G3 is capable of transporting a 25-mer phosphorodiamidate morpholino oligomer (PMO) into the cells. It exhibits antisense effect against Gli1 gene in hedgehog pathway, confirmed in preliminary b-glactosidase assay performed in Ptch1 -/cells. Further antisense effect was confirmed when G3-conjugated zebrafish Gli1-PMO was injected into embryo to obtain Gli1dependent phenotypes. To the best of our knowledge, herein, we first report that molecular transporters with a minimum number of three cationic groups are capable of efficiently passing through plasma, nuclear and mitochondrial membranes and show distribution in zebrafish larvae.[a] Dr. Figure 1. Chemical structures of (a) DNA, (b) PMO, transporters (c) G3 (1) and (d) P3 (2).

Morpholino‐modified nucleoside analogues have widespread applications in developmental biology. To achieve nucleobase‐functionalized forms of morpholino nucleosides, syntheses of 5‐substituted cytidine, 8‐substituted adenosine, and 8‐substituted guanosine morpholino nucleoside monomers are described for the first time. The syntheses are based on the use of 5‐iodocytidine, 8‐bromoadenosine, and 8‐bromoguanosine morpholino nucleosides as the key starting materials. These iodo or bromo derivatives have also been synthesized for the first time. Palladium‐mediated cross‐coupling reactions (Sonogashira, Suzuki, and Heck) were then employed with the halo derivatives to accomplish the substitutions. Different reaction conditions for C, A, and G were standardized to achieve the conversions. The strategy was devised in such a way that the useful N‐trityl protecting groups remain at the end. The catalyst combinations used for Sonogashira, Suzuki, and Heck reactions were Pd(PPh3)2Cl2·CuI, Pd(dppf)Cl2·CH2Cl2, and Pd(OAc)2, respectively. Heck coupling between 5‐iodocytidine monomer and methyl acrylate worked well, whereas with acrylonitrile the exocyclic amine of cytidine was found to form the aza‐Michael adduct. In this context, treatment of iodocytidine with methyl acrylate under two different sets of conditions was found to produce either the Michael addition product or the Heck coupling product. Four of the functionalized morpholino monomers have been further confirmed by single‐crystal X‐ray structural analysis. All of these functionalized monomers were obtained in good to excellent overall yields.

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Sonogashira coupling of 5‐iodouracil derivatives (I) with various terminal alkynes gives the title compounds (III) (18 examples) in good yields.

Alkylation of Acylmetalates with Alkyl Halides to Prepare Fischer Carbene Complexes: An Improved Protocol. -Alkoxy Fischer carbene complexes of chromium, tungsten and molybdenum are synthesized by alkylation of lithium acylmetalates with alkyl halides in the presence of catalytic amounts of tetrabutylammonium bromide under biphasic conditions. The new method can also be applied to synthesize optically active products, albeit in poor yield [cf. (VIII)]. -(NANDI, B.; SINHA*, S.; Tetrahedron 67 (2011) 1, 106-113, http://dx.