| CPC A61K 9/1277 (2013.01) [A61K 9/145 (2013.01); A61K 47/60 (2017.08); A61K 47/69 (2017.08); B82Y 5/00 (2013.01)] | 20 Claims | 

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               1. An apparatus for producing a lipid-encapsulated RNA nanoparticle, comprising: 
              (a) a 1st tube having an ID of between about 0.1″ and 0.132″; connected at one end to a 1st HPLC pump and at the other end to a mixing module, wherein the 1st HPLC pump is configured to pump an aqueous solution comprising RNA through the 1st tube at a flow rate of at least 150 mL/min; 
                (b) a 1st reservoir connected to the 1st HPLC pump, wherein the 1st reservoir contains the aqueous solution; 
                (c) a 2nd tube having an ID of between about 0.005″ and 0.02″ connected at one end to a 2nd HPLC pump and at the other end to the mixing module, wherein the 2nd HPLC pump is configured to pump an ethanol solution comprising one or more lipids through the 2nd tube at a flow rate greater than 50 mL/min, 
                wherein the 2nd tube is perpendicularly joined to the 1st tube at the mixing module; 
                (d) a 2nd reservoir connected to the 2nd HPLC pump, wherein the 2nd reservoir contains the ethanol solution, 
                wherein the apparatus is configured to mix the ethanol solution with the aqueous solution by introducing the ethanol solution into the aqueous solution in a region within the mixing module to produce an output solution having a flow that produces turbulence; 
                wherein the one or more lipids comprise a cationic lipid having a pKa of about 6 to about 7 and a structure of Formula I: 
              ![]() or a pharmaceutically acceptable salt or solvate thereof, wherein 
              R5 and R6 are each independently selected from the group consisting of a linear or branched C1-C31 alkyl, C2-C31 alkenyl or C2-C31 alkynyl and cholesteryl; 
                  L5 and L6 are each independently selected from the group consisting of a linear C1-C20 alkyl and C2-C20 alkenyl; 
                  X5 is —C(O)O— or —OC(O)—; 
                  X6 is —C(O)O— or —OC(O)—; 
                  X7 is S or O; 
                  L7 is absent or lower alkyl; 
                  R4 is a linear or branched C1-C6 alkyl; and 
                  R7 and R8 are each independently selected from the group consisting of a hydrogen and a linear or branched C1-C6 alkyl. 
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