Processes for the continuous production of biomolecules

Abstract

The present invention relates to a system (1) for the continuous production of a biomolecule, comprising a feed tank (2) suitable for storing a product and a reaction chamber (4) designed to receive a product from the feed tank that forms a reaction phase in the reaction chamber, and the reaction chamber designed to produce the biomolecule from the reaction phase. The system also comprises a scale tank (6) in fluid communication with the reaction chamber and suitable for storing an immiscible phase, and a monitoring and control unit (2) configured to control the injection flow rate of the immiscible phase into the reaction chamber in order to maintain a certain constant filling level of the reaction chamber according to the amount of reaction phase injected into the reaction chamber to produce a certain amount of the biomolecule.

Description

[Technical Field]

[0001] The present invention is directed generally to the field of in vitro production of biomolecules, specifically production of mRNA into lipid nanoparticles for use in therapeutic applications such as vaccines. [Background technology]

[0002] Recent scientific and technological advances have made biomolecules promising candidates for a variety of uses, including diagnostic applications and therapeutic products such as vaccines.

[0003] Driven by major scientific, technological, and medical advances, the emergence of new therapies targeting genetic mechanisms, such as gene replacement, correction, or modulation of protein expression, is enabling innovative treatments as well as new opportunities for emergency response in epidemic crisis situations. In this context, various approaches have been developed for large-scale mRNA production. The most recent process utilizes in vitro enzymatic reactions to synthesize mRNA from self-replicating DNA templates, followed by encapsulation of the total RNA in lipid nanoparticles as an immiscible delivery vehicle.

[0004] These processes are expensive, require highly skilled personnel, and generate hazardous waste streams that must be mediated, while production rates depend heavily on the performance of the enzymatic reactions, the ability to remove process and product-related impurities, and the control and efficiency of encapsulation into lipid nanoparticles.

[0005] Additionally, current practice is to produce mRNA in batch mode, which provides little flexibility in terms of scaling up production; for each scale, the process must be significantly adapted and the changes must be approved by regulatory authorities.

[0006] The recent pandemic has demonstrated that there is a need to accelerate the development of current vaccines and their availability to the population.

[0007] In this regard, WO2021212034 describes a novel process for producing mRNA in an in vitro system using continuous flow production.

[0008] The system described in WO2021212034 comprises a reaction chamber in which in vitro transcription takes place to continuously produce mRNA.

[0009] The reaction chamber operates continuously so that it can be infused with new input material while mRNA is being produced.

[0010] This system is particularly adapted for producing large amounts of mRNA, such as millions of doses in a pandemic situation, but not for producing small amounts of mRNA, e.g., thousands of doses for early clinical trials, without wasting large amounts of enzymes and buffers.

[0011] In practice, in relation to scale-up modifications in mRNA production, small-scale production is generally carried out in smaller batch reactors with volumes adapted to the desired production amount.

[0012] Therefore, the system described in WO2021212034 is not adapted to quickly synthesize and produce different amounts of mRNA at low cost. Summary of the Invention

[0013] SUMMARY OF THE INVENTION It is therefore an object of the present invention to solve the above-mentioned problems.

[0014] To this end, the present invention relates to a system for the continuous production of biomolecules, comprising a supply tank suitable for storing a product, and a reaction chamber, the reaction chamber being designed to be supplied with a product from the supply tank which forms a reaction phase in the reaction chamber, the reaction chamber being designed to produce the biomolecule from the reaction phase, the system comprising a scale tank in fluid communication with the reaction chamber and suitable for storing an immiscible phase which is not miscible with the reaction phase, and a monitoring and control unit for controlling the injection flow rate of the immiscible phase into the reaction chamber in order to maintain a certain filling level of the reaction chamber according to the amount of reaction phase injected into the reaction chamber to produce a certain amount of biomolecule, such that the smaller the amount of reaction phase injected into the reaction chamber, the greater the amount of immiscible phase injected.

[0015] The idea behind the present invention is to use a single system adapted to produce biomolecules from small amounts to large amounts, or vice versa.

[0016] The idea consists more specifically in dimensioning the system for the production of biomolecules on a large scale while allowing their production on a small scale by keeping the same reaction conditions and therefore the same quality results (concentration, purity, production time, etc.), i.e. the reaction phase must always have the same reaction conditions.

[0017] For this purpose, when small quantities are produced, even at a scale lower than that for which the reaction chamber is normally designed, an immiscible phase is injected to fill the volume not used by the reaction phase in the reaction chamber.

[0018] The two phases are immiscible, and the reaction phase flowing within the reaction chamber is always contained within a volume adapted for a stable and efficient reaction.

[0019] Thus, the system of the present invention is advantageously adapted to produce different amounts of a biomolecule, for example at different stages of vaccine clinical development.

[0020] The idea is also to use a single system to produce a selected amount of a biomolecule at a specific residence time, which means that the system according to the invention is designed to produce small or large amounts of a biomolecule at the same residence time.

[0021] The system according to the invention may also have the following features: - the system comprises a valve at an outlet of the reaction chamber and the monitoring and control unit, the monitoring and control unit being configured to receive data representative of the presence of a reaction phase at the outlet of the reaction chamber and then control the valve to separate the immiscible phase from the reaction phase; - the data representative of the presence of a reactive phase at the outlet of the reaction chamber is calculated based on the duration of the residence time of the reactive phase in the reaction chamber; - the system comprises a mixing chamber in fluid communication between the supply tanks and the reaction chamber, the mixing chamber being designed to receive products from the supply tanks, mix the products to form the reaction phase, and then supply the reaction phase to the reaction chamber, and the monitoring and control unit is configured to control in real time the injection flow rates of the products from each supply tank to maintain a constant ratio between the products in the mixing chamber; - the system comprises a first chromatography device comprising a first column in fluid communication between a reaction chamber and a collection tank and a second column in fluid communication between the first column and the collection tank, the first chromatography device comprising a first temporary storage tank in fluid communication between the first column and the second column, the first chromatography device further comprising a first outlet valve at the outlet of the first column designed to switch fluid communication from the first column to the first temporary storage tank or from the first column to the collection tank, and the first chromatography device comprising a first inlet valve at the inlet of the second column designed to open or close fluid communication between the first temporary storage tank and the second column; - the first chromatography device comprises a second temporary storage tank in fluid communication between the second column and the first column, the first chromatography device comprises a second outlet valve at the outlet of the second column designed to switch fluid communication from the second column to the second temporary storage tank or from the second column to the collection tank, and the first chromatography device further comprises a second inlet valve at the inlet of the first column designed to open or close fluid communication from the second temporary storage tank to the first column; - the first chromatography device comprises means for measuring the concentration of the biomolecule at the outlet of the first column or the second column, and the monitoring and control unit is configured to control the outlet valve at the outlet of the first column or the second column to feed the first or second temporary storage tank, respectively, when the concentration of the biomolecule in the pre-fraction and the post-fraction of the eluted product fraction from the first column or the second column is above a certain predetermined concentration threshold, or to feed the waste tank, when the concentration of the biomolecule is below the certain predetermined concentration threshold; - the system comprises a scale buffer tank suitable for storing a completing buffer, the scale buffer tank being fluidly connected to the first column, and the monitoring and control unit is configured to control the injection flow rate of the completing buffer from the scale buffer tank to the first column in order to maintain a certain filling level of the column according to the amount of reaction phase injected into the column, such that the smaller the amount of reaction phase injected into the first column, the greater the amount of buffer injected into the first column; - the system comprises an in-line diafiltration system in fluid communication between the purification device and the lipid nanoparticle formulation system, the diafiltration system including several stages designed to pass a reaction phase with an exchange buffer, the monitoring and control unit being configured to receive data representing a certain concentration of a biomolecule used by the lipid nanoparticle formulation system to form the lipid nanoparticles, and the monitoring and control unit being configured to control the injection of the exchange buffer in the final stage to dilute the biomolecule to the certain concentration of the biomolecule; - the system comprises a purification system arranged at the outlet of the reaction chamber designed to separate the produced biomolecule from the rest of the reaction phase; the system comprises means for measuring a certain concentration of each product in the remainder of the reaction phase, and the monitoring and control unit is configured to control the injection of the remainder of the reaction phase into the mixing chamber.

[0022] The invention also extends to a process for the continuous production of biomolecules using the system of the invention, wherein the biomolecule produced is RNA such as mRNA, or protein, or DNA, and the biomolecule produced may be a therapeutic agent such as a vaccine.

[0023] More specifically, the process of the present invention for the continuous production of biomolecules comprises: - supplying a product from a supply tank to a reaction chamber, wherein the product forms a reaction phase in the reaction chamber; producing the biomolecule from the reaction phase in the reaction chamber; storing an immiscible phase that is not miscible with the reaction phase in a scale tank in fluid communication with the reaction chamber; and controlling, using a monitoring and control unit, the injection flow rate of the immiscible phase into the reaction chamber to maintain a certain filling level of the reaction chamber according to the amount of reaction phase injected into the reaction chamber to produce a certain amount of biomolecule, such that the less the amount of reaction phase injected into the reaction chamber, the greater the amount of immiscible phase injected.

[0024] The process of the present invention also comprises: - receiving, at the monitoring and control unit, data representative of the presence of the reaction phase at the outlet of the reaction chamber, and then using the monitoring and control unit to control a valve at the outlet of the reaction chamber to separate the immiscible phase from the reaction phase, - the data representative of the presence of a reaction phase at the outlet of the reaction chamber is calculated based on the duration of the residence time of the reaction phase in the reaction chamber; - feeding products from the feeding tanks into a mixing chamber in fluid communication between the feeding tanks and the reaction chamber, mixing the products to form the reaction phase, and then feeding the reaction phase into the reaction chamber, and using the monitoring and control unit to control in real time the injection flow rate of the products from each feeding tank to maintain a constant ratio between the products in the mixing chamber; - separating the product fraction eluted from the first column into a pre-fraction and a post-fraction of the product fraction eluted from the first column; directing the pre-fraction and the post-fraction from the first column to a first temporary storage tank, and once both the pre-fraction and the post-fraction are stored in the temporary storage tank, directing the pre-fraction and the post-fraction from the first temporary storage tank to a second column; - separating the product fraction eluted from the second column into a pre-fraction and a post-fraction of the product fraction eluted from the second column, directing the pre-fraction and the post-fraction from the second column to a second temporary storage tank, and once both the pre-fraction and the post-fraction have been stored in the second temporary storage tank, directing the pre-fraction and the post-fraction from the second temporary storage tank to the first column; - measuring the concentration of the biomolecule at the outlet of the first column or the second column and controlling, using the monitoring and control unit, the outlet valve at the outlet of the first column or the second column to feed the first or second temporary storage tank, respectively, when the concentration of the biomolecule in the pre- and post-fractions of the eluted product fraction from the first or second column is above a certain predetermined concentration threshold, or to feed the waste tank, when the concentration of the biomolecule is below the certain predetermined concentration threshold; - storing a buffer in a scale buffer tank fluidly connected to the first column and controlling, using said monitoring and control unit, the injection flow rate of the buffer from the scale buffer tank to the first column in order to maintain a certain filling level of the column according to the amount of reaction phase injected into the column, such that the smaller the amount of reaction phase injected into the first column, the greater the amount of buffer injected into the first column; - passing a reaction phase with an exchange buffer in several stages of an in-line diafiltration system in fluid communication between the purification device and the lipid nanoparticle formulation system, receiving in a monitoring and control unit data representing a certain concentration of a biomolecule to be used by the lipid nanoparticle formulation system to form lipid nanoparticles, and controlling, using the monitoring and control unit, the injection of the exchange buffer in the final stage to dilute the biomolecule to the certain concentration of the biomolecule; - separating the produced biomolecules from the remainder of the reaction phase using a purification system arranged at the outlet of the reaction chamber; - measuring a certain concentration of each product in said remainder of the reaction phase and controlling the injection of said remainder of the reaction phase into the mixing chamber using said monitoring and control unit. The invention will be better understood and other advantages will become apparent from the detailed description of embodiments given by way of non-limiting example and illustrated by the accompanying drawings, in which: [Brief explanation of the drawings]

[0025] [Figure 1] 1 is a schematic diagram of a system of the present invention. [Figure 2] 1 is a schematic diagram of a first portion of a system of the present invention used for the production of biomolecules. [Figure 3] FIG. 1 is a schematic diagram of a second part of the system of the invention used for the purification of molecules. [Figure 4] FIG. 1 is a schematic diagram of a third part of the system of the present invention used for filtration of biomolecules. DETAILED DESCRIPTION OF THE INVENTION

[0026] The system 1 for continuous production of biomolecules of the present invention, as shown in FIG. 1, is particularly adapted for producing different amounts and types of biomolecules, such as RNA (ribonucleic acid), DNA (deoxyribonucleic acid) or proteins.

[0027] These manufactured biomolecules can be used as therapeutic agents, such as vaccines.

[0028] The system 1 of the present invention operates continuously thanks to, inter alia, a monitoring and control unit 2 configured to enforce reaction conditions based on input parameterizations and real-time monitoring of said reaction conditions via multiple sensors distributed throughout the system for feedback control of the reaction conditions according to the input parameterizations.

[0029] The reaction conditions that can be controlled in a non-exhaustive manner to obtain the required quantity and quality of biomolecules are temperature, pH, flow rate, residence time, and amount of product injected for the reaction.

[0030] In the embodiment shown in FIG. 1, the system of the present invention can be divided into several parts, each having a specific function in the production of a biomolecule.

[0031] The first part 1P is dedicated to the production of biomolecules.

[0032] The first part 1P comprises two supply tanks 3 suitable for storing products and a reaction chamber 4 designed to receive products from the supply tanks 3, forming a reaction phase in the reaction chamber 4.

[0033] The reaction chamber 4, also called reactor or bioreactor, is here designed to produce the biomolecules of interest from a reaction phase.

[0034] More specifically, the system 1 may include a mixing chamber 5 in fluid communication between the supply tanks 3 and the reaction chambers 4. In this case, the mixing chamber 5 is designed to receive products from the supply tanks 3, mix the products to form a reaction phase, and then deliver the reaction phase to the reaction chambers 4. The product injection flow rates from each supply tank are controlled in real time by the monitoring and control unit 2, which is configured to maintain a constant ratio between the products in the mixing chambers 4. This makes it possible to maintain a constant reaction despite changes in the amount of biomolecule produced. Indeed, the goal is to maintain a stable and reproducible reaction regardless of the amount of biomolecule produced.

[0035] As shown in FIG. 1, the system 1 includes a scale tank 6 in fluid communication with the reaction chamber 4 and suitable for storing an immiscible phase that is not miscible with the reaction phase.

[0036] The immiscible phase used in the system of the present invention can be a liquid, such as an organic phase, a solid, or a gas.

[0037] Therefore, the monitoring and control unit 2 is configured to control the injection flow rate of the immiscible phase into the reaction chamber 4 so that the less the amount of reaction phase injected into the reaction chamber 4, the greater the amount of immiscible phase injected, in order to keep a certain filling level of the reaction chamber 4 constant according to the amount of reaction phase injected into the reaction chamber 4 to produce a certain amount of biomolecule.

[0038] Furthermore, in order to separate the immiscible phase from the reaction phase, the system may comprise a valve 7 at the outlet 4a of the reaction chamber 4. The monitoring and control unit 2 is therefore configured to receive data representative of the presence of the reaction phase at the outlet 4a of the reaction chamber 4 and to then control said valve 7.

[0039] For example, the data representative of the presence of a reactive phase at the outlet 4 a of the reaction chamber 4 is calculated based on the duration of a particular residence time of the reactive phase within the reaction chamber 4 .

[0040] Thus, the reactive phase is injected into the intermediate tank IT and the immiscible phase is injected into the waste tank WT.

[0041] Without limiting the scope of the invention, the data representative of the presence of the reaction phase at the outlet 4a of the reaction chamber 4 can also be determined by a UV sensor or Raman.

[0042] In order to maintain the reaction phase in the reaction chamber 4 at a substantially predetermined temperature according to the input parameterization, the system 1 may comprise, as shown in FIG. 2, a temperature sensor 8 designed to measure the temperature of the reaction phase and a thermostat 9 placed in contact with the reaction chamber 4 and controlled by the monitoring and control unit 2 as a function of the measured temperature.

[0043] The system may also comprise a pH sensor 10 designed to measure the pH of the reaction phase. In that case, the system comprises at least one buffer tank 11 suitable for storing a pH adjuster product, fluidly connected to the reaction chamber 4. The monitoring and control unit 2 is therefore configured to control the injection flow rate of the pH adjuster product as a function of the measured pH level, so as to maintain the reaction phase substantially at a predetermined pH level in the reaction chamber 4.

[0044] The system may also comprise mass flow meters 12 designed to measure the flow rate of each product injected into the reaction chamber 4. In that case, the system 1 comprises mass flow controllers 13 designed to control the injection flow rates of these products in the reaction chamber 4. The monitoring and control unit 2 is therefore configured to control the mass flow controllers 13 to control the injection flow rates of the reaction phases as a function of the measured flow rates so as to maintain a specific flow rate of the reaction phases in the reaction chamber 4.

[0045] The system 1 of the present invention may also comprise means MC for measuring the concentration of biomolecules at the outlet 4a of the reaction chamber 4 obtained after a residence time preset in the input parameterization and for controlling instead the injection of products into the reaction chamber 4 in order to increase the amount of biomolecules produced to obtain the desired amount of biomolecules.

[0046] The system 1 of the invention may also comprise a purification system 14, here a filtration system as an example, arranged at the outlet 4a of the reaction chamber 4, designed to separate the produced biomolecules from the rest of the reaction phase. In that case, the system may comprise means for measuring a certain concentration of each product in said rest of the reaction phase, and said monitoring and control unit is configured to control the injection of said rest of the reaction phase in the mixing chamber 5 accordingly.

[0047] The second part 2P of the system 1 is dedicated to the purification of biomolecules, as shown in FIG.

[0048] For that purpose, the system of the invention comprises a first chromatography device 15 comprising a first column 16 in fluid communication between the reaction chamber 4, more specifically an intermediate tank IT, and a collection tank 17, and a second column 18 in fluid communication between the first column 16 and the collection tank 17. The first chromatography device 15 further comprises a first temporary storage tank 19 in fluid communication between the first column 16 and the second column 18. The first chromatography device 15 further comprises a first outlet valve 20 at the outlet 16a of the first column 16, designed to switch the fluid communication from the first column 16 to the first temporary storage tank 19 or from the first column 16 to the collection tank 17, and a first inlet valve 21 at the inlet 18b of the second column 18, designed to open or close the fluid communication between the first temporary storage tank 19 and the second column 18.

[0049] More specifically, the first chromatography device 15 may comprise a second temporary storage tank 22 in fluid communication between the second column 18 and the first column 16. In that case, the first chromatography device 15 comprises a second outlet valve 23 at the outlet 18a of the second column 18, which is designed to switch fluid communication from the second column 18 to the second temporary storage tank 22 or from the second column 18 to the recovery tank 17, and a second inlet valve 24 at the inlet 16b of the first column 16, which is designed to open or close fluid communication from the second temporary storage tank 22 to the first column 16.

[0050] The first chromatography device 1 may further comprise means for measuring the concentration of biomolecules at the outlets 16a, 18a of the first column 16 or the second column 18, and the monitoring and control unit 2 is configured to control the outlet valves 20, 23 at the outlets 16a, 18a of the first column 16 or the second column 18 to supply the biomolecules in the pre-fraction and post-fraction of the eluted product fraction from the first column 16 or the second column 18 to the first temporary storage tank 19 or the second temporary storage tank 22, respectively, when the concentration of the biomolecules in the pre-fraction and post-fraction of the eluted product fraction from the first column 16 or the second column 18 is above a certain predetermined concentration threshold, or to supply the biomolecules to the waste tank 25 when the concentration of the biomolecules is below the certain predetermined concentration threshold.

[0051] The second chromatographic device 26 can be used to improve the purification of the first chromatographic device 1. For example, reverse phase chromatography can be used.

[0052] The third part 3P is dedicated to the filtration of purified biomolecules, and the fourth part 4P is dedicated to lipid nanoparticle formation, as shown in Figure 4.

[0053] In this regard, the system 1 comprises an in-line diafiltration system 27 in fluid communication between the first purification device 1 or the second purification device 26 and the lipid nanoparticle formulation system 28, the diafiltration system 27 comprising several stages 29 designed to pass the reaction phase using an exchange buffer stored in a buffer tank 30.

[0054] Accordingly, the monitoring and control unit 2 is configured to receive data representing a certain concentration of the biomolecule to be used by the lipid nanoparticle formulation system 28 to form lipid nanoparticles, and data on the real-time concentration of the biomolecule measured by the concentration sensor 31 at the outlet of the final stage 29. Accordingly, the monitoring and control unit 2 is configured to control the injection of an exchange buffer in the final stage 29 to dilute the biomolecule at said certain concentration of the biomolecule, and to inject said biomolecule at said certain concentration into the intermediate tank 32 before directing said biomolecule to the lipid nanoparticle formulation system 28.

[0055] The invention further relates to a process for producing a biomolecule, here for example for the production of mRNA (Messenger ribonucleic acid) for use as a vaccine.

[0056] The first step consists of in vitro transcription of mRNA. Thus, a first master mix containing linearized specific DNA encoding at least part of a protein is stored in a first supply tank 3, and a second master mix containing RNA polymerase and nucleotides is stored in a second supply tank 3. The first and second master mixes are mixed in a specific ratio in a chamber 5 to form a reaction phase.

[0057] In order to maintain a constant ratio between both master mixes in the mixing chamber 5, the injection flow rates of the products from each supply tank 3 are controlled in real time by the monitoring and control unit 2.

[0058] The reaction phase is then injected into the reaction chamber 4 to realize the in vitro transcription of mRNA according to the reaction conditions imposed by the input parameterization of the monitoring and control unit 2.

[0059] This means that the reaction chamber 4 contains all the necessary products and is able to provide all the reaction conditions required for the production of mRNA.

[0060] The reaction chamber is also provided with an immiscible phase that is not miscible with the reaction phase since the reaction phase is an aqueous phase.

[0061] Therefore, the monitoring and control unit 2 is configured to control the injection flow rate of the immiscible phase into the reaction chamber 4 to maintain a certain filling level of the reaction chamber 4 according to the amount of reaction phase injected into the reaction chamber 4 to produce a certain amount of biomolecule.

[0062] More specifically, the smaller the amount of reaction phase injected into the reaction chamber 4, the greater the amount of immiscible phase injected.

[0063] Therefore, it is necessary to separate the immiscible phase from the reaction phase to allow rapid and reliable purification of the produced mRNA.

[0064] In this way, the process of the present invention consists of determining by the monitoring and control unit 2 the presence or absence of a reactive phase at the outlet 4a of the reaction chamber 4 based on a specific duration of a specific residence time of the reactive phase in the reaction chamber 4, and controlling by the monitoring and control unit 2 the separation of the reactive phase from the immiscible phase.

[0065] Also, in order to limit production costs, there is a need to recycle the products used in the reaction steps, such as enzymes and nucleotides. To this end, the process consists of separating the produced biomolecules from the rest of the reaction phase at the outlet 4a of the reaction chamber 4 by diafiltration.

[0066] An additional step consists of determining the concentration of each product of the remainder of the reaction phase and then controlling the injection of the remainder of the reaction phase into the mixing chamber 5. The monitoring and control unit 2 is therefore set to adjust the concentration of each master mix added from the supply tank 2 after the remainder of the reaction stage in order to maintain a constant ratio.

[0067] The separated reaction phase is then purified in a chromatographic process that consists of separating the product fraction eluted from the first column 16 into a pre-fraction and a post-fraction of the product fraction eluted from the first column 16, directing the pre-fraction and the post-fraction from the first column 16 to a temporary storage tank 19, and once both the pre-fraction and the post-fraction have been stored in the temporary storage tank 19, directing the pre-fraction and the post-fraction from the temporary storage tank 19 to the second column 18.

[0068] Thus, the product fraction eluted from the second column 18 can be directed to a collection tank or can be purified again.

[0069] If the product fraction needs to be purified again, the process includes the additional steps of separating the product fraction eluted from the second column 18 into pre- and post-product fractions eluted from the second column 18, directing the pre- and post-product fractions from the second column 18 to a second temporary storage tank 22, and once both the pre- and post-product fractions have been stored in the second temporary storage tank 22, directing the pre- and post-product fractions from the second other temporary storage tank 22 to the first column 16.

[0070] The idea is to elute the maximum amount of mRNA from the column by collecting the post- and pre-fractions without wasting time.

[0071] In that case, it is possible to measure the concentration of biomolecules at the outlet of the first column 16 or the second column 18 by means of a measuring means MM, such as a UV sensor, in order to direct the pre-fraction and post-fraction from the first column or the second column, respectively, to a first or second temporary storage tank 19, 22 if the concentration is above a certain predetermined concentration threshold, or to a waste tank 30 if the concentration is below said certain predetermined concentration threshold.

[0072] Also, depending on the amount of biomolecule to be produced, there is a need to purify different amounts of biomolecule using the purification device 15 of the system 1 according to the present invention.

[0073] Therefore, the idea of ​​the present invention is to control the injection flow rate of the supplementary buffer in the first column 16 in order to maintain a certain constant filling level in the first column 16 as the reaction phase passes through the first column 16.

[0074] In this situation, one single first column 16 may be used and sized for the large amount of biomolecules to be produced.

[0075] This allows for a continuous process by limiting the use of consumables and human intervention at the point of change in production scale.

[0076] Changing the consumables should be understood to mean changing the column depending on the amount of biomolecules to be produced.

[0077] Therefore, the monitoring and control unit 2 is configured to control the injection flow rate of the supplementary buffer from the scale buffer tank BT of the chromatography device 15 into the first column 16 in order to maintain this certain filling level of the first column 16 according to the amount of reaction phase injected into the first column 16.

[0078] More specifically, this means that the smaller the volume of reaction phase injected into the first column 16, the greater the volume of buffer injected into the first column 16.

[0079] By way of example, the complementary buffer used can be a neutral solution that does not modify the pH or the integrity and quality of the biomolecules produced.

[0080] It will be appreciated that the monitoring and control unit may be set to control the injection flow rate of the supplemental buffer from the scale buffer tank BT into the second column 18 in order to maintain a certain fill level.

[0081] Reverse phase chromatography can then be used to improve the quality of the purification.

[0082] The third step consists of filtering the mRNA purified by a chromatographic process.

[0083] This step consists of filtering the mRNA in the reaction phase by passing it through several stages 29 of the diafiltration system 27 with an exchange buffer and by injecting a certain amount of exchange buffer in the final stage 29 to dilute the mRNA at a specific concentration ready to be used for lipid nanoparticle formulation.

[0084] The fourth step consists of the formation of lipid nanoparticles.

[0085] This step is known in its entirety, but the aim of the present invention is to be able to use a certain concentration of mRNA that is particularly well suited for the formation of nanoparticles, this concentration being determined by the input parameters and the dilution carried out during the diafiltration process.

Claims

1. A process (1) for the continuous production of biomolecules, comprising: - feeding a product from a feed tank into a reaction chamber (4), said product forming a reaction phase in said reaction chamber; - producing said biomolecule from said reaction phase in said reaction chamber, said process comprising: - storing an immiscible phase that is not miscible with the reaction phase in a scale tank (6) in fluid communication with the reaction chamber; - controlling the injection flow rate of the immiscible phase into the reaction chamber using a monitoring and control unit (2) in order to maintain a certain filling level of the reaction chamber according to the amount of the reaction phase injected into the reaction chamber to produce a certain amount of biomolecule, so that the less the amount of the reaction phase injected into the reaction chamber, the greater the amount of the immiscible phase injected.

2. The process comprises: receiving, in said monitoring and control unit, data representative of the presence of said reaction phase at the outlet of said reaction chamber; - then using said monitoring and control unit, controlling a valve (7) at said outlet (4a) of said reaction chamber (4) in order to separate said immiscible phase from said reaction phase.

3. 3. The process for continuous production of biomolecules according to claim 2, characterized in that the data representing the presence of the reaction phase at the outlet of the reaction chamber is calculated based on the duration of the residence time of the reaction phase in the reaction chamber.

4. The process comprises: - feeding the product from said supply tank into a mixing chamber (5) in fluid communication between said supply tank and said reaction chamber; mixing said products to form said reaction phase and then feeding said reaction phase into said reaction chamber; - controlling in real time, by means of the monitoring and control unit, the injection flow rates of the products from each supply tank in order to maintain a constant ratio between the products in the mixing chamber.

5. The process comprises: - separating the product fraction eluted from the first column (16) into a fraction before and a fraction after the product fraction eluted from said first column; - directing said pre-fraction and said post-fraction from said first column to a first temporary storage tank (19); - directing the front and rear fractions from the first temporary storage tank to a second column (18) once both the front and rear fractions have been stored in the temporary storage tank.

6. The process comprises: - separating the product fraction eluted from the second column into a fraction before and a fraction after the product fraction eluted from the second column; - directing said pre-fraction and said post-fraction from said second column to a second temporary storage tank (22); - directing the front and rear fractions from the second temporary storage tank to the first column once both the front and rear fractions have accumulated in the second temporary storage tank.

7. The process comprises: - measuring the concentration of a biomolecule at the outlet of the first column or the second column; - controlling, using the monitoring and control unit, an outlet valve at the outlet of the first column or the second column to feed the first or second temporary storage tank, respectively, when the concentration of the biomolecule in the front and back fractions of the eluted product fraction from the first column or the second column is above a certain predetermined concentration threshold, or to feed the biomolecule in the back and back fractions of the eluted product fraction from the first column or the second column to a waste tank (25), when the concentration of the biomolecule is below the certain predetermined concentration threshold.

8. The process comprises: storing a buffer in a scale buffer tank (BT) fluidly connected to said first column; - controlling the injection flow rate of the buffer from the scale buffer tank into the first column using the monitoring and control unit in such a way that the less the amount of reaction phase injected into the first column, the greater the amount of buffer injected into the first column, in order to maintain a certain filling level of the column according to the amount of reaction phase injected into the column.

9. The process comprises: - passing said reaction phase with an exchange buffer in several stages (29) of an in-line diafiltration (27) system in fluid communication between the purification device and a lipid nanoparticle formulation system (28); - receiving, at said monitoring and control unit, data representing certain concentrations of biomolecules used by said lipid nanoparticle formulation system to form said lipid nanoparticles; - controlling the injection of the exchange buffer in a final stage using the monitoring and control unit in order to dilute the biomolecule at a certain concentration of the biomolecule.

10. 10. A process for the continuous production of biomolecules according to any one of claims 1 to 9, characterized in that the process comprises a step of separating the produced biomolecules from the remainder of the reaction phase using a purification system (14) arranged at the outlet of the reaction chamber.

11. The process comprises: - measuring a certain concentration of each product in the remainder of the reaction phase; - controlling the injection of the remainder of the reaction phase into the mixing chamber using the monitoring and control unit.

12. 12. The process for continuous production of biomolecules according to any one of claims 1 to 11, wherein the produced biomolecule is RNA.

13. 13. The process for continuous production of biomolecules according to claim 12, wherein the RNA produced is mRNA.

14. 12. The process for continuous production of biomolecules according to any one of claims 1 to 11, wherein the produced biomolecule is a protein.

15. 12. The process for continuous production of biomolecules according to any one of claims 1 to 11, wherein the produced biomolecule is DNA.

16. 16. The process for continuous production of biomolecules according to any one of claims 11 to 15, wherein the produced biomolecule is a therapeutic agent.

17. 17. The process for continuous production of biomolecules of claim 16, wherein the therapeutic agent is a vaccine.

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