There are provided compounds of Formula (A) and pharmaceutically acceptable salts and esters thereof, and pharmaceutical compositions thereof, used for the prevention or treatment in a mammal of joint and bone disorders such as arthritis and osteoporosis.

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There are provided compounds of Formula (A) and pharmaceutically acceptable salts and esters thereof, and pharmaceutical compositions thereof, used for the prevention or treatment in a mammal of joint and bone disorders such as arthritis and osteoporosis.

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This specification discloses a novel methodology for labelling RNA via enzymatic incorporation of a minimally perturbing fluorescent tricyclic cytosine analogue. This analogue is shown to be 100% incorporated in example transcripts and is fully compatible with both in vitro and in cell transcription. Spectroscopic characterization shows that the incorporation rate of the cytosine analogue is on par with its natural counterpart. Using live cell imaging and flow cytometry, labelled mRNAs are efficiently and correctly translated upon transfection into living cells and cell-free systems. The spectral properties of the modified transcripts and their correct translation product allow for their straightforward and simultaneous visualization. This technology therefore offers a general route to understanding the biological behaviour of RNA of interest, including RNA based drugs. The fluorescent tricyclic cytosine analogue has formula (I):

##STR00001##

This specification discloses a novel methodology for labelling RNA via enzymatic incorporation of a minimally perturbing fluorescent tricyclic cytosine analogue. This analogue is shown to be 100% incorporated in example transcripts and is fully compatible with both in vitro and in cell transcription. Spectroscopic characterization shows that the incorporation rate of the cytosine analogue is on par with its natural counterpart. Using live cell imaging and flow cytometry, labelled mRNAs are efficiently and correctly translated upon transfection into living cells and cell-free systems. The spectral properties of the modified transcripts and their correct translation product allow for their straightforward and simultaneous visualization. This technology therefore offers a general route to understanding the biological behaviour of RNA of interest, including RNA based drugs. The fluorescent tricyclic cytosine analogue has formula (I):

##STR00001##

In the present invention, a method for producing a compound of formula I comprises a step for causing a compound of formula III to undergo a Pechmann condensation reaction with respect to a compound of formula II in the presence of an organic solvent and an acid catalyst to obtain a compound of formula IV, and due to such method, provided are a novel photoreactive compound and a method for producing same that can be used for nucleic acid photoreaction technology.

In the present invention, a method for producing a compound of formula I comprises a step for causing a compound of formula III to undergo a Pechmann condensation reaction with respect to a compound of formula II in the presence of an organic solvent and an acid catalyst to obtain a compound of formula IV, and due to such method, provided are a novel photoreactive compound and a method for producing same that can be used for nucleic acid photoreaction technology.

There are provided compounds of Formula (A) and pharmaceutically acceptable salts and esters thereof, and pharmaceutical compositions thereof, used for the prevention or treatment in a mammal of joint and bone disorders such as arthritis and osteoporosis. ##STR00001##

There are provided compounds of Formula (A) and pharmaceutically acceptable salts and esters thereof, and pharmaceutical compositions thereof, used for the prevention or treatment in a mammal of joint and bone disorders such as arthritis and osteoporosis. ##STR00001##

Herein reported are new tricyclic cytidine compounds, such as 8-diethylamino-tC (8-DEA-tC), that respond to DNA and/or RNA duplex formation with up to a 20-fold increase in fluorescent quantum yield as compared with the free nucleoside, depending on neighboring bases. This turn-on response to duplex formation is by far the greatest of any reported nucleoside analogue that can participate in Watson-Crick base pairing. Measurements of the quantum yield of 8-DEA-tC mispaired with adenosine and, separately, opposite an abasic site show that there is almost no fluorescence increase without the formation of correct Watson-Crick hydrogen bonds. Kinetic isotope effects from the use of deuterated buffer show that the duplex protects 8-DEA-tC against quenching by excited state proton transfer. DFT calculations provide a rationale for the observed photophysical properties that is dependent on duplex integrity and the electronic structure of the analogue.

Herein reported are new tricyclic cytidine compounds, such as 8-diethylamino-tC (8-DEA-tC), that respond to DNA and/or RNA duplex formation with up to a 20-fold increase in fluorescent quantum yield as compared with the free nucleoside, depending on neighboring bases. This turn-on response to duplex formation is by far the greatest of any reported nucleoside analogue that can participate in Watson-Crick base pairing. Measurements of the quantum yield of 8-DEA-tC mispaired with adenosine and, separately, opposite an abasic site show that there is almost no fluorescence increase without the formation of correct Watson-Crick hydrogen bonds. Kinetic isotope effects from the use of deuterated buffer show that the duplex protects 8-DEA-tC against quenching by excited state proton transfer. DFT calculations provide a rationale for the observed photophysical properties that is dependent on duplex integrity and the electronic structure of the analogue.