The present invention provides, among other things, multimeric coding nucleic acids that exhibit superior stability for in vivo and in vitro use. In some embodiments, a multimeric coding nucleic acid (MCNA) comprises two or more encoding polynucleotides linked via 3 ends such that the multimeric coding nucleic acid compound comprises two or more 5 ends.

This invention provides a range of translatable polynucleotide and oligomer molecules for expressing a human phenylalanine hydroxylase (PAH), or a fragment thereof having PAH activity. The polynucleotide and oligomer molecules are expressible to provide the human PAH or a fragment thereof having PAH activity. The molecules can be used as active agents to express an active polypeptide or protein in cells or subjects. The agents can be used in methods for ameliorating, preventing, delaying onset, or treating a disease or condition associated with phenylketonuria, decreased metabolism of phenylalanine, or increased levels of phenylalanine in a subject.

A lentiviral vector system for expressing a lentiviral particle is disclosed. The lentiviral vector system includes a therapeutic vector. The therapeutic vector comprises a phenylalanine hydroxylase (PAH) sequence for expressing at least one of PAH or a variant thereof, wherein the PAH sequence is truncated.

A method of attaching a cell or a membrane-coated particle-of-interest to a microtube is provided. The method comprising: co-electrospinning two polymeric solutions through co-axial capillaries, wherein a first polymeric solution of the two polymeric solutions is for forming a shell of the microtube and a second polymeric solution of the two polymeric solutions is for forming a coat over an internal surface of the shell, the first polymeric solution is selected solidifying faster than the second polymeric solution and a solvent of the second polymeric solution is selected incapable of dissolving the first polymeric solution and wherein the second polymeric solution comprises the cell or the membrane-coated particle-of-interest, thereby attaching the cell or the membrane-coated particle-of-interest to the microtube. Also provided are microtubes with attached, entrapped or encapsulated cells or membrane-coated particles and methods of using same

A method of attaching a molecule-of-interest to a microtube, by co-electrospinning two polymeric solutions through co-axial capillaries, wherein a first polymeric solution of the two polymeric solutions is for forming a shell of the microtube and a second polymeric solution of the two polymeric solutions is for forming a coat over an internal surface of the shell, the first polymeric solution is selected solidifying faster than the second polymeric solution and a solvent of the second polymeric solution is selected incapable of dissolving the first polymeric solution and the second polymeric solution comprises the molecule-of-interest, thereby attaching the molecule-of-interest to the microtube. An electrospun microtube comprising an electrospun shell, an electrospun coat over an internal surface of the shell and a molecule-of-interest attached to the microtube.

Described herein are ceDNA vectors having linear and continuous structure for delivery and expression of a transgene. ceDNA vectors comprise an expression cassette flanked by two ITR sequences, where the expression cassette comprises a codon optimized nucleic acid sequence encoding a PAH protein, in combination with particular promoter sequences and cis-regulatory elements. Further provided herein are methods and cell lines for reliable gene expression of PAH protein in vitro, ex vivo and in vivo using the ceDNA vectors. Also provided herein are methods and compositions comprising ceDNA vectors useful for the expression of PAH protein in a cell, tissue or subject, and methods of treatment of diseases with said ceDNA vectors expressing PAH protein. Such PAH protein can be expressed for treating disease, e.g., Phenylketonuria (PKU).

The invention relates to the use of adeno-associated virus (AAV) vectors to achieve long term expression of a transgene in the liver of a juvenile subject. The invention includes the stable long-term amelioration of disease symptoms of the subjection following a single administration of an AAV vector to a juvenile subject, wherein the AAV vector delivers the transgene to the subject's liver.

Provided herein is a tyrosine selection marker system, and uses thereof. In some embodiments, nucleic acid constructs, vectors, host cells and related compositions and methods for generating and selecting tyrosine prototroph cells are provided.

This invention provides a range of translatable polynucleotide and oligomer molecules for expressing a human phenylalanine hydroxylase (PAH), or a fragment thereof having PAH activity. The polynucleotide and oligomer molecules are expressible to provide the human PAH or a fragment thereof having PAH activity. The molecules can be used as active agents to express an active polypeptide or protein in cells or subjects. The agents can be used in methods for ameliorating, preventing, delaying onset, or treating a disease or condition associated with phenylketonuria, decreased metabolism of phenylalanine, or increased levels of phenylalanine in a subject.

This disclosure relates to mRNA therapy for the treatment of hyperphenylalaninemias such as phenylketonuria (PKU). mRNAs for use in the invention, when administered in vivo, encode human phenylalanine hydroxylase (PAH), functional fragments thereof (e.g., those comprising the catalytic domain or the catalytic domain and the tetramerization domains), and fusion proteins comprising PAH. mRNAs of the invention are preferably encapsulated in lipid nanoparticles (LNPs) to effect efficient delivery to cells and/or tissues in subjects, when administered thereto. mRNA therapies of the invention increase and/or restore deficient levels of PAH expression and/or activity in subjects. mRNA therapies of the invention further decrease abnormal accumulation of phenylalanine associated with deficient PAH activity in subjects.