METHODS AND APPARATUSES FOR MANUFACTURING FOR REMOVING MATERIAL FROM A THERAPEUTIC COMPOSITION
Methods and apparatuses for making and using therapeutics, including in particular mRNA therapeutics, that separate double-stranded RNA from single-stranded RNA as part of a continuous flow. These methods and apparatuses may include formulation of an RNA therapeutic using a permeable insert integrated into a microfluidic path device. In particular, these methods and apparatuses may include formulation of an RNA therapeutic by removing dsRNA from a solution of RNA by within a microfluidic path device including a cellulose material.
1 . A method of manufacturing a therapeutic polynucleotide using a system comprising a plurality of fluid depots configured to be secured in sealed fluid communication with one or more microfluidic path devices, the method comprising: transporting reagents between one or more fluid depots of the plurality of fluid depots and a plurality of reactors on the one or more microfluidic path devices in a sealed and closed fluidic path that is protected from atmospheric contact to perform the steps of: forming a synthetic template, performing in vitro transcription from the template to produce a therapeutic polynucleotide, and purifying the therapeutic polynucleotide.
2 . The method of claim 1 , wherein purifying the therapeutic polynucleotide comprises two-dimensional (2D) purification of the therapeutic polynucleotide within one or more of the plurality of reactors.
3 . The method of claim 1 , further comprising formulating the therapeutic polynucleotide with a delivery vehicle in one or more reactors on the one or more microfluidic path devices to form a therapeutic polynucleotide composition.
4 . The method of claim 3 , further comprising concentrating the therapeutic polynucleotide composition in the one or more microfluidic path devices.
5 . The method of claim 3 , wherein the delivery vehicle comprises an amphipathic nanoparticle.
6 . The method of claim 5 , wherein the amphipathic nanoparticle comprises an amino-lipidated peptoid.
7 . The method of claim 1 , wherein purifying the therapeutic polynucleotide comprises removing double-stranded mRNA using a cellulose material within the one or more reactors.
8 . The method of claim 1 , wherein the system automatically and continuously performs the steps of forming the synthetic template, performing in vitro transcription from the template, and purifying the therapeutic polynucleotide with optical feedback from one or more sensors of system.
9 . The method of claim 1 , wherein the therapeutic polynucleotide is an mRNA.
10 . The method of claim 1 , wherein the therapeutic polynucleotide is an mRNA, a circular RNA or a self-replicating RNA.
11 . The method of claim 1 , wherein the system transports the reagents between the one or more fluid depots and the plurality of reactors by fluid power.
12 . The method of claim 11 , wherein the system pneumatically transports the reagents between the one or more fluid depots and the plurality of reactors.
13 . The method of claim 1 , wherein the system transports the reagents between the one or more fluid depots and the plurality of reactors by deflecting one or more elastic layers within the one or more microfluidic path devices.
14 . The method of claim 1 , wherein the method is performed at a site of care.
15 . The method of claim 1 , wherein the system automatically and continuously performs the steps of forming a synthetic template, performing in vitro transcription from the template to produce a therapeutic polynucleotide, and purifying the therapeutic polynucleotide in less than 3 days.
16 . The method of claim 1 , further comprising sealing the fluid depots to the one or more microfluidic path devices and pressurizing the fluid depots before transporting the reagents between the fluid depots and the plurality of reactors on the one or more microfluidic path devices.
17 . The method of claim 1 , further comprising recording movement of fluid within the one or more microfluidic path devices during the performance of the steps in a file associated with the therapeutic polynucleotide that is manufactured.
18 . A therapeutic polynucleotide made using the method of claim 1 .
19 . A method of manufacturing a therapeutic mRNA using a system comprising a plurality of fluid depots configured to be secured in sealed fluid communication with one or more microfluidic path plate devices, wherein the one or more microfluidic path plate devices comprise a plurality of reactors, the method comprising:
delivering a template precursor material from one or more fluid depots to a first one or more reactor regions of the plurality of reactors and processing the template precursor material to prepare a template from the template precursor material;
transferring the template to a second one or more reactor regions of the plurality of reactors and processing the template by in vitro transcription to form a therapeutic mRNA; and
transferring the therapeutic mRNA to a third one or more reactor regions of the plurality of reactors and purifying the therapeutic mRNA by two-dimensional (2D) purification within the third one or more reactor regions;
wherein all of the method steps are performed without exposing the template and therapeutic mRNA to atmospheric contact.
20 . A method of manufacturing a therapeutic mRNA using a system comprising a plurality of fluid depots in sealed fluid communication with one or more microfluidic path plate devices, wherein the one or more microfluidic path plate devices comprise a plurality of reactors, the method comprising:
delivering, using fluid power, a template precursor material from one or more fluid depots to a first one or more reactor regions of the plurality of reactors and processing the template precursor material to prepare a template from the template precursor material;
transferring, using fluid power, the template to a second one or more reactor regions of the plurality of reactors and processing the template by in vitro transcription to form a therapeutic mRNA;
transferring, using fluid power, the therapeutic mRNA to a third one or more reactor regions of the plurality of reactors and purifying the therapeutic mRNA by two-dimensional (2D) purification within the third one or more reactor regions;
transferring, using fluid power, the therapeutic mRNA to a fourth one or more reactor regions of the plurality of reactors and encapsulating the therapeutic mRNA with a delivery vehicle to form a therapeutic mRNA composition; and
concentrating, using fluid power, the therapeutic mRNA composition in a fifth one or more fluid depots,
wherein all of the method steps are performed without exposing the template and the therapeutic mRNA to atmospheric contact.
21 - 99 . (canceled)