The present disclosure provides for base editors which satisfy a need in the art for installation of targeted transversions of guanine (G) to thymine (T), or correspondingly, transversions of adenine (A) to cytosine (C). The domains of the disclosed base editors include a nucleic acid programmable DNA binding protein and a guanine oxidase or a guanine methyltransferase. The base editors may be engineered through the use of continuous or non-continuous evolution systems. In particular, the present disclosure provides for guanine-to-thymine (or cytosine-to-adenine) base editors that can install single-base trans version mutations. In addition, methods for targeted nucleic acid editing are provided. Further provided are pharmaceutical compositions comprising, and vectors and kits useful for the generation of, guanine-to-thymine base editors. Cells containing such vectors and cells containing base editors and guide RNAs are also provided. Further provided are methods of treatment comprising administering the base editors to a subject in need thereof.

The invention provides a method of determining a predisposition to a condition associated with ATP depletion, such as an ischaemic event, in a subject comprising: a. measuring the concentration of one or more purines in a body fluid of the subject, the purines being selected from adenosine, inosine, hypoxanthine, xanthine and ATP, and b. comparing the measured concentration with a threshold concentration of the one or more purines, wherein the threshold concentration is preferably in the range 2 [micro]M to 8 [micro]M and wherein a measured concentration higher than the threshold concentration indicates the presence of ischaemia.

Methods and therapeutics for treating nitric oxide (NO) mediated conditions comprising administering a therapeutically effective amount of a pharmaceutical composition containing a therapeutic, wherein the therapeutic one of increases xanthine oxidase (XO) associated nitrite conversion to nitric oxide and increases nitrite conversion to nitric oxide via diallyl disulfide (DADS), and the NO mediated condition includes one of inflammation, wound healing, and an infection.

Methods and therapeutics for treating nitric oxide (NO) mediated condition comprising administering a therapeutically effective amount of a pharmaceutical composition containing a therapeutic, wherein the therapeutic one of increases xanthine oxidase (XO) associated nitrite conversion to nitric oxide and increases nitrite conversion to nitric oxide via DADS. Method and devices for measuring an amount of nitrite in a sample comprising obtaining a sample, adding diallyl disulfide (DADS) to the sample, measuring a post DADS amount of nitric oxide (NO) in the sample, and using the post DADS amount of NO in the sample to determine the amount of nitrite in the sample. Methods and therapeutics for diagnosing and treating critical limb ischemia (CLI) versus non-critical limb ischemic peripheral artery disease (PAD) in a patient comprising determining a patient indicator value, where the patient indicator is value one of a total hydrogen sulfide metabolite plasma level, a total nitric oxide, nitrite, and nitrite (NOx) plasma level, a ratio of free hydrogen sulfide plasma level to total NOx plasma level, and a ratio of total hydrogen sulfide metabolite plasma level and total NOx plasma level, diagnosing the patient with as having CLI based on the patient indicator value, and administering to the patient a therapeutic for CLI.

The present invention relates to compositions comprising a polymeric matrix or a gel containing functional enzymes capable of re-creating under culture conditions the cell microenvironment existing in vivo. The present invention also relates to devices for cell cultures comprising such compositions, in particular hydrogel and the use thereof to control the chemical microenvironment of a cell culture or mimic physiological or pathological conditions of the in vivo cells. The compositions and the devices described herein could be also used in vitro for evaluating the therapeutic effect of a compound on a determined cell line or on primary cells.

The present invention relates to compositions comprising a polymeric matrix or a gel containing functional enzymes capable of re-creating under culture conditions the cell microenvironment existing in vivo. The present invention also relates to devices for cell cultures comprising such compositions, in particular hydrogel and the use thereof to control the chemical microenvironment of a cell culture or mimic physiological or pathological conditions of the in vivo cells. The compositions and the devices described herein could be also used in vitro for evaluating the therapeutic effect of a compound on a determined cell line or on primary cells.

The present invention relates to processes for the formation of furandicarboxylic acid (FDCA), in particular 2,5-furandicarboxylic acid (2,5-FDCA), and mono- and diester derivatives thereof, via a multistep biocatalytic oxidation reaction of 5-hydroxymethylfurfural (HMF) using, for example, an enzyme selected from the group consisting of xanthine oxidoreductase (XOR), galactose oxidase variant M.sub.3-5, aldehyde dehydrogenase, and/or ketoreductase. The invention also relates to copolymers that comprise the furandicarboxylic acid monomers and derivatives thereof, processes for the formation of the copolymers and uses for the copolymers.

The present invention relates to compositions comprising a polymeric matrix or a gel containing functional enzymes capable of re-creating under culture conditions the cell microenvironment existing in vivo. The present invention also relates to devices for cell cultures comprising such compositions, in particular hydrogel and the use thereof to control the chemical microenvironment of a cell culture or mimic physiological or pathological conditions of the in vivo cells. The compositions and the devices described herein could be also used in vitro for evaluating the therapeutic effect of a compound on a determined cell line or on primary cells.

Methods and therapeutics for treating nitric oxide (NO) mediated condition comprising administering a therapeutically effective amount of a pharmaceutical composition containing a therapeutic, wherein the therapeutic one of increases xanthine oxidase (XO) associated nitrite conversion to nitric oxide and increases nitrite conversion to nitric oxide via DADS. Method and devices for measuring an amount of nitrite in a sample comprising obtaining a sample, adding diallyl disulfide (DADS) to the sample, measuring a post DADS amount of nitric oxide (NO) in the sample, and using the post DADS amount of NO in the sample to determine the amount of nitrite in the sample. Methods and therapeutics for diagnosing and treating critical limb ischemia (CLI) versus non-critical limb ischemic peripheral artery disease (PAD) in a patient comprising determining a patient indicator value, where the patient indicator is value one of a total hydrogen sulfide metabolite plasma level, a total nitric oxide, nitrite, and nitrite (NOx) plasma level, a ratio of free hydrogen sulfide plasma level to total NOx plasma level, and a ratio of total hydrogen sulfide metabolite plasma level and total NOx plasma level, diagnosing the patient with as having CLI based on the patient indicator value, and administering to the patient a therapeutic for CLI.

The present invention relates to processes for the formation of furandicarboxylic acid (FDCA), in particular 2,5-furandicarboxylic acid (2,5-FDCA), and mono- and diester derivatives thereof, via a multistep biocatalytic oxidation reaction of 5-hydroxymethylfurfural (HMF) using, for example, an enzyme selected from the group consisting of xanthine oxidoreductase (XOR), galactose oxidase variant M.sub.3-5, aldehyde dehydrogenase, and/or ketoreductase. The invention also relates to copolymers that comprise the furandicarboxylic acid monomers and derivatives thereof, processes for the formation of the copolymers and uses for the copolymers.