Method and system for orthogonal bioorthogonal release of species

Abstract

The present disclosure relates to a system for release of a releasable species, the system (100) comprises a device (110) comprising a reservoirs (111) arranged to contain trigger species (121,122), transport means (112) and an outlet (113). At least two types of target species (133,135) are comprised in an interaction region (130) outside the device (110), each comprising a releasable species (134,136). The device (110) is arranged to transport each trigger species (121,122) from the reservoir (111) into said interaction region (130) via said outlet (113) utilizing said transport means (112). The target species (133,135) are arranged to be fluidly connected to said outlet (113). The device (110) comprises a computer (115) arranged to control said transport means (112). Each type of trigger species (121,122) is configured to react with a corresponding target species (133,135), and wherein said reaction comprises releasing the corresponding releasable species (134,136).

Description

[0001] Title

[0002] Method and system for orthogonal bioorthogonal release of species

[0003] TECHNICAL FIELD

[0004] The present disclosure relates to molecular release systems, drug release systems, orthogonal bioorthogonal chemistry, and chronotherapy drug delivery.

[0005] BACKGROUND ART

[0006] Bioorthogonal chemistry has emerged as an enabling method for the construction and use of molecules in contexts that were previously unthinkable, such as inside a living organism. The impact of bioorthogonal reactivity has been far-reaching with applications spanning protein synthesis, drug discovery, in vivo chemistry and polymer science. Bioorthogonal chemistry is expected to develop into a tool capable of delivering new drugs, creating new materials and answering biological questions that cannot be addressed by modem molecular biology tools.

[0007] In some situations, administrating two or more drugs in a specific region may provide significant benefits, especially if good temporal control over the concentration of said drugs can be achieved. Choreographed release of a combination of different drugs can solve this problem, but their independent temporal release is often challenging, and in many cases impossible, to realize using existing technologies. Moreover, such a choreographed release of multiple drugs with distinct release profiles opens new ground by providing novel chronotherapeutic treatment scenarios.

[0008] There is a need for methods and systems for an improved spatial and temporal control of release and activation of species, such as releasing two or more different drugs / compounds.

[0009] SUMMARY OF THE INVENTION

[0010] This invention describes a drug delivery system with electronic precision that allows multiple- yet independent drug release with distinct concentration time profiles. One object of the invention is to provide tools for improved controlled release of compounds in time, space and concentration.

[0011] This has, in accordance with the present disclosure, been achieved by means of a system for orthogonal bioorthogonal release of at least two types of releasable species. The system comprises a device comprising one or more reservoirs, transport means and an outlet, wherein said one or more reservoirs are arranged to contain at least two types of trigger species. The system further comprises at least two types of target species comprised in an interaction region outside the device, wherein each of said target species comprise a releasable species or a precursor thereof. Said outlet is arranged in and / or at the interaction region, and wherein the device is arranged to transport each trigger species from said one or more reservoirs into said interaction region via said outlet utilizing said transport means. Said target species are arranged to be fluidly connected to said outlet, thereby being accessible for said trigger species transported into the interaction region via said outlet. The device comprises a computer arranged to control said transport means and being arranged to control transporting said trigger species into said interaction region via said outlet. Each type of trigger species is configured to react with a corresponding target species, and said reaction comprises releasing the corresponding releasable species. Said reactions between trigger species and corresponding target species are bioorthogonal reactions, and wherein two or more of said reactions are mutually orthogonal to each other.

[0012] This has the advantage of allowing two- or more parallel systems each using separate bioorthogonal click and release systems, such as trigger and activatable drug, that are mutually orthogonal to each other, thereby allowing for individual release and thus administrated completely separately from each other at the same target area using individual timing, positioning and released concentration. This further has the advantage of allowing for parallel independent chronotherapies using a combination of drugs with distinct timing of drugs (‘clocking the drug’) that can significantly improve drug tolerability and efficacy in patients.

[0013] In some embodiments, the computer is arranged to control the timing of transport and the amount transported of each trigger species into the interaction region based on predetermined instructions comprising a schedule for transport rates of two or more trigger species over a period of time. In some embodiments, the system comprises a sensor arranged in and / or at the interaction region, wherein said sensor is connected to said computer, and wherein said computer is arranged to control the timing of transport and the amount transported of each trigger species into the interaction region based on a sensor signal from said sensor and a set of criteria.

[0014] This has the advantage of allowing the system to create and / or execute schedules for transport of trigger species, thereby releasing releasable species, based on sensor values and / or predetermined instructions. This further has the advantage of allowing for choreographed release of a plurality of releasable species over time.

[0015] In some embodiments, said computer is arranged to store data indicative of amounts of transported trigger species and / or sensor signals from the sensor, and is arranged to control transport of each trigger species based on said stored data and a set of criteria.

[0016] This has the advantage of allowing the system to create and execute schedules for transport of trigger species, thereby releasing releasable species, based on previous sensor signals and transporting of trigger species. This further has the advantage of allowing the system to measure a concentration of releasable species or the trigger species with a sensor and adjusting the transport rate of corresponding trigger species based on the measured concentration and a corresponding target concentration, thereby allowing for chronotherapy and synchronized drug release.

[0017] In some embodiments, the system comprises a scaffold and / or hydrogel arranged in said interaction region, and wherein said at least two types of target species are covalently bound to said scaffold and / or hydrogel.

[0018] This has the advantage of allowing the target species to be kept in a defined location and being accessible to objects fluidly connected to the scaffold or hydrogel, such as trigger species exiting the outlet of the device.

[0019] In some embodiments, each of said at least two types of target species is covalently bound to a corresponding separate region of said scaffold and / or hydrogel, thereby allowing targeted release of releasable species in a specific region in said interaction region functionalized with the corresponding target species. This has the advantage of allowing control over release of releasable species in both the type of releasable species and the release location in the interaction region.

[0020] In some embodiments, a region of said scaffold and / or hydrogel comprises at least one type of capture species each configured to, upon reacting with a corresponding trigger species, covalently bind and / or modify said trigger species, wherein a modified trigger species is unable to chemically activate the corresponding target species so as to cause release of said releasable species.

[0021] This has the advantage of allowing improved control over the transported trigger species and the interaction region, by capturing or inactivating trigger species attempting to move between the interaction region and the environment the risk is reduced that transported trigger species exit into the environment or that trigger species in the environment enter the interaction region. This may further have the advantage of allowing a plurality of systems to operate in the same vicinity with the same or similar trigger species with reduced risk of trigger species from one system impacting other systems.

[0022] In some embodiments, the transport means comprise electrokinetic transport means arranged to provide electrokinetic transport of trigger species from said one or more reservoirs into said interaction region via said outlet.

[0023] The present disclosure further relates to a method for orthogonal bioorthogonal release of at least two types of releasable species. The method comprising

[0024] - providing at least two types of target species comprised in an interaction region, wherein said target species each comprise a corresponding releasable species or a precursor thereof, wherein each target species is configured to, upon reacting with a corresponding trigger species, release the corresponding release species;

[0025] - determining a target release of said release species utilizing computer; and

[0026] - providing trigger species into said interaction region based on the determined target release of said releasable species; wherein said reactions between trigger species and corresponding target species are bioorthogonal reactions, and wherein two or more of said reactions are mutually orthogonal to each other. In some embodiments, the method comprises measuring the state of said interaction region utilizing a sensor arranged to determine sensor data, and wherein determining the target release of said releasable species is based on obtained sensor data indicative of a measured state in said interaction region.

[0027] BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Fig. 1a-b shows schematically a system for orthogonal bioorthogonal release of species.

[0029] Fig. 2 shows schematically a system for orthogonal bioorthogonal release of species with capture species.

[0030] Fig. 3 shows a method for orthogonal bioorthogonal release of species.

[0031] DETAILED DESCRIPTION

[0032] Throughout the figures, same reference numerals refer to same parts, concepts, and / or elements. Consequently, what will be said regarding a reference numeral in one figure applies equally well to the same reference numeral in other figures unless not explicitly stated otherwise.

[0033] It is to be understood that the figures are schematic representations aimed at highlighting the novel features of the examples. Shown parts may not to scale and some parts may not be depicted for readability, such as the polymer backbone of hydrogels. Typically, the examples shown are in an aqueous and / or biological environment.

[0034] The term target species relates to a chemical substance or ensemble composed of chemically identical molecular entities.

[0035] The term trigger species relates to an ion or molecule. Typically, the trigger species is arranged to react with a corresponding target species in a bioorthogonal manner. Typically, the trigger species is introduced or transported to react with the corresponding target species at a specified location. The term target species relates to an ion or molecule. Typically, the target species is arranged to react with a corresponding trigger species in a bioorthogonal manner, whereby the target species releases a releasable species. The target species comprises the releasable species or a precursor for the releasable species. Typically, the target species is arranged at a specified location, such as a being bound to a scaffold or a hydrogel in a desired interaction region.

[0036] The term releasable species relates to an ion or molecule released as a result of a reaction between a trigger species and a corresponding target species. Typically, the releasable species is a drug, a toxin, or a bioactive compound arranged to interact with something inside and / or outside the interaction region. Typically, the releasable species is less readily transportable by the techniques used to transport the trigger species, such as being large or uncharged.

[0037] The term bioorthogonal chemistry relates to a class of reactions that are intrinsically selective transformations not commonly found in biology. Bioorthogonal reactions are intrinsically chemoselective. For example, bioorthogonal reactions typically proceed rapidly and selectively in biological environments with little or no reactivity towards endogenous functional groups. Typically, bioorthogonal chemistry is configured to readily proceed in aqueous environments at biocompatible pH and temperature.

[0038] The terms bioorthogonal bond-cleavage reaction, BBCR, click-to-release, C2R, and bioorthogonal uncaging relate to a bioorthogonal reaction that initiates a subsequent elimination process, leading to the liberation of a molecule, such as a drug, protein, toxin, or bioactive compound.

[0039] The term orthogonal bioorthogonal relates to a plurality of sets of interacting species reacting in a bioorthogonal manner, wherein each set of interacting species reacts mutually orthogonal to other sets. For example, a first trigger species and a corresponding first target species react independently of a second trigger species and a corresponding second target species, despite being in proximity.

[0040] The term hydrogel relates to a polymer network that may be functionalized, or loaded, with molecules. For example, the hydrogel may be functionalized with covalently bound activatable and / or releasable species, such as drugs, toxin or bioactive molecules, thus functioning as a reservoir of said species. The term scaffold relates to support structures for attaching and functionalizing molecules.

[0041] The term transport means relates to a system or device for controlled delivery of a trigger species from a reservoir into a desired volume. For example, the transport means may be controlled by a computer arranged to provide concentration, temporal, and spatial control of the trigger species. For example, the transport means may be of electrokinetic nature, such as an ion pump, or a fluidic pump. It is to be understood that the term computer may be simple control circuitry comprised in the device and / or integrated within the transport means that is arranged to control the operation of the transport means based on a received signal or from an internal schedule for transport.

[0042] The expression fluidly connected relates to the ability to flow or diffuse from one component to another, but the two components need not be physically connected to one another.

[0043] Fig. 1a-b shows schematically a system for orthogonal bioorthogonal release of species. Fig. 1a shows an example system with a device 110 arranged to transport a first trigger species 121 and a second trigger species 122 from one reservoir 111 into an interaction region 130 outside the device 110. Fig. 1 b shows an example system arranged to transport a first trigger species 121 and a second trigger species 122 from a reservoir 111 into an interaction region 130 outside the device 110. In the figures, the first trigger species 121 is represented by a “V” shaped reactive site and the second trigger species 122 is represented by a “U” shaped reactive site, and the corresponding target species 133,135 have reactive sites of a matching shape. It is to be understood that the depicted example systems are typically arranged to function in aqueous environments.

[0044] Fig. 1a shows schematically an example system for orthogonal bioorthogonal release of at least two types of releasable species. The example system 100 comprises a device 110 comprising a reservoir 111 , transport means 112 and an outlet 113, wherein said reservoir 111 is arranged to contain at least two types of trigger species 121 , 122. The system 100 further comprises at least two types of target species 133,135 comprised in an interaction region 130 outside the device 110, wherein said target species 133,135 each comprise a releasable species 134,136 and / or a precursor thereof. The outlet 113 is arranged in and / or at the interaction region 130, and the device 110 is arranged to transport each trigger species 121 ,122 from said reservoir 111 into said interaction region 130 via said outlet 113 utilizing said transport means 112. The target species 133,135 are arranged to be fluidly connected to said outlet 113, thereby being accessible for said trigger species 121 ,122 transported into the interaction region 130 via said outlet 113, such as the target species 133,135 being covalently bound to a hydrogel and / or a scaffold in the interaction region 130 that said trigger species 121 ,122 can readily diffuse through.

[0045] The device 110 comprises a computer 115 arranged to control said transport means 112 and being arranged to control transporting said trigger species 121 ,122 into said interaction region 130 via said outlet 113. Each type of trigger species 121 ,122 is configured to react with a corresponding target species 133,135, wherein said reaction comprises releasing the corresponding releasable species 134,136. Said reactions between trigger species 121 ,122 and corresponding target species 133,135, are bioorthogonal reactions, and wherein two or more of said reactions being mutually orthogonal to each other. In some of these examples, all the reactions of corresponding trigger species 121 ,122 and target species 133,135 are all mutually orthogonal to each other.

[0046] In some examples, the interaction region 130 is comprised in a vessel. In some examples, the interaction region 130 is a region within 100 mm of the outlet 113. In some of these examples, the interaction region 130 is a region within 50 mm, 30 mm, 20 mm, 10 mm, 5 mm, or 2 mm of the outlet 113.

[0047] In the example system in fig. 1a, the device 110 comprises a sensor 116 arranged to measure outside the device 111 connected to the computer 115, wherein the computer 115 is arranged to control the transport means based on sensor signals obtained from said sensor. In some of these examples, the sensor 116 is arranged at the outlet 113. In some examples, the sensor 116 is arranged in the inner hydrogel region and / or in the environment outside the hydrogel or the device 110.

[0048] In some examples, the sensor 116 is arranged to measure a concentration of trigger species 121 ,122. In some examples, the sensor 116 is arranged to measure a concentration of releasable species 134,136. In some examples, the computer 115 is arranged to calculate a schedule for transport rates of two or more trigger species 121 , 122 over a period of time based on sensor signals and / or predetermined instructions, and to control the transport means 112 to transport trigger species 121 ,122 based on said calculated schedule. In some examples, the predetermined instructions comprise a set of criteria. In some examples, the predetermined instructions comprise a schedule for transport rates of two or more trigger species 121 , 122 over a period of time. In some examples, the computer 115 is arranged to control the transport means 112 to transport trigger species 121 ,122 based on predetermined instructions. In some examples, the device 110 comprises a memory storage (not shown) connected to the computer 115 and arranged to store said predetermined instructions and / or sensor signals.

[0049] In some of these examples, the computer 115 is arranged to store transport data indicative of the amount of transported two or more trigger species 121 ,122, and calculate the schedule for transport rates of two or more trigger species 121 , 122 over a period of time based on said stored transport data. The amount of transported trigger species 121 , 122 may, for example, correspond to an amount of depletion of the target species 133,135 in the interaction region 130, such that after transporting a large amount of trigger species 121 ,122 the required amount of transported trigger species 121 ,122 increases for a certain amount of interaction with the target species 133,135. Alternatively, the amount of transported trigger species 121 ,122 may indicate the amount of trigger species 121 , 122 remaining in the one or more reservoirs 111. In some examples, the computer 115 is arranged to store sensor data, and calculate the transport rates of two or more trigger species 121 ,122 based on said stored sensor data. In some examples, the computer 115 is arranged to continuously calculate transport rates of the two or more trigger species 121 , 122 based on sensor signals and / or predetermined instructions, and control the transport means 112 based on said calculate transport rates.

[0050] In fig. 1a is shown a sequence of each trigger species 121 ,122 reacting with a corresponding target species 133,135 in the interaction region 130 to release their corresponding releasable species 134,136, wherein depicted reaction intermediates 133’, 135’ represent non-thermodynamically stable compounds. In some examples, the at least one trigger species 121 ,122 is arranged to react with a corresponding target species 133,135 in the interaction region 130, wherein said reaction comprises bioorthogonal bond-cleavage reaction (BBCR), click-to-release (C2R), and / or bioorthogonal uncaging.

[0051] In some examples, the at least one trigger species 121 ,122 comprises tetrazine, Tz, bicyclo[6.1 ,0]non-4-yn-9-ylmethanol, cyclooctyne, bicyclo[6.1 ,0]non-4-yn-9-ylmethanol, BCN, cycloalkynes, fluoride, and / or trans-cyclooctene, TCO, or derivatives thereof.

[0052] In some examples, the at least one target species 133,135 comprises trans-cyclooctene, TCO, iminosydnone, mesoionic compounds, ortho-carbamoylmethylene silyl ethers, tetrazine, Tz, or derivatives thereof.

[0053] Target species 133,135 comprising ortho-carbamoylmethylene silyl ethers may allow for desilylation induced release using silyl ether groups.

[0054] In some examples, each trigger species 121 ,122 and corresponding target species 133,135 comprises respectively tetrazine, Tz, and trans-cyclooctene, TCO; bicyclo[6.1 ,0]non-4-yn-9-ylmethanol, BCN and iminosydnone; bicyclo[6.1 ,0]non-4-yn-9-ylmethanol, BCN and chlorinated iminosydnone; bicyclo[6.1 ,0]non-4-yn-9-ylmethanol, BCN and mesoionic compounds; fluoride, and ortho-carbamoylmethylene silyl ethers; tetrazines and vinyl ethers; tetrazines and vinylboronic acids; trans-cyclooctene, TCO, and tetrazine, Tz; or derivatives thereof. In some examples, the at least one target species 133,135 comprises chlorinated iminosydnone.

[0055] In some examples, the at least two types of trigger species 121 , 122 each comprise at least one of the following moieties: tetrazine (Tz), fluoride, trans-cyclooctene, TCO, and bicyclononyne, cyclooctyne, BCN. In some of these examples, the at least two types of trigger species 121 ,122 comprise different moieties.

[0056] It is to be understood that in order to transport trigger species, such as BCN or Tz, with an ion pump it may be necessary to form charged derivatives thereof.

[0057] It is to be understood that several combinations of trigger species and target species are possible to use, and the releasable species may typically be selected rather freely and without many restrictions due to the reacting parts of the trigger species and target species.

[0058] In some of these examples, the releasable species 134,136 is arranged to causes a biological response. In some examples, the releasable species 134,136 is arranged to interact with cells, microorganisms, tissue and / or parts thereof to cause a biological response. In some of these examples, the interaction region comprising the target species 133,135 is arranged at and / or around said cells and / or tissue. In some examples, the interaction region comprising the target species 133,135 is arranged to be inside a non-human animal and / or a human.

[0059] In some examples, the system 100 is configured to be arranged at and / or around cells, microorganisms and / or tissue, such as in being configured to be in fluid contact with a cell culture via a culture medium.

[0060] In some examples, the system 100 is configured to be arranged inside a non-human animal and / or a human, such as the hydrogel 130 being injectable and the device 110 being implantable.

[0061] In some examples, the device 110 comprises an energy source (not shown) arranged to power the transport means 112. In some of these examples, the energy source is a battery. In some examples, the battery is connected to at least two electrodes (not shown) of the transport means 112 arranged to generate electric fields in the transport path of the trigger species 121 between the reservoir 111 and the outlet 113. In some examples, the transport means 112 may be arranged to transport each trigger species 121 , 122 individually from the reservoir 111 to a corresponding outlet 113. In some of these examples, the computer 115 may be arranged to provide individually controlled release of each releasable species 134,136 by transporting each trigger species 121 ,122 individually from the reservoir 111. In some examples, the transport means 112 are arranged to transport each of said at least two trigger species 121 ,122 independently from the corresponding reservoir 111 to the corresponding outlet 113. In some of these examples, the transport means 112 are arranged to simultaneously transport each trigger species 121 ,122 independently from the corresponding reservoir 111 to the corresponding outlet 113. In some examples, the transport means 112 are arranged to transport trigger species from the corresponding reservoir 111 to the corresponding outlet 113 for at least 1 second, at least 10 seconds, or at least 1 minute. In some examples, the transport means 112 are arranged to simultaneously transport at least two trigger species 121 ,122 from the corresponding reservoir 111 to the corresponding outlet 113.

[0062] In some examples, the transport means 112 comprise a mechanical pump, and / or a non-mechanical pump. In some examples, the transport means 112 comprise a micropump. In some examples, the transport means 112 comprise an electroosmotic pump, and / or an ion pump.

[0063] In some examples, the device 110 is arranged to independently transport a first 121 and a second trigger species 122 utilizing said transport means 112. In some examples, the device 110 is arranged to transport the first 121 and the second trigger species 122 at at least two different relative rates, such as transporting more of the first trigger species 121 during a first mode of operation, and more of the second trigger species 122 during a second mode of operation.

[0064] In an example scenario, the transport means 112 are arranged to transport Tetrazine at a rate of 5 pmol / min. Depending on transported molecule and choice of transport means 112 the rate may be in a range between attomol per minute to micromol per minute. In some examples, the transport means 112 are arranged to transport at least the first species 112 at 1 femtomol per minute to 1 nanomol per minute. It is to be understood that the term transport means 112 may relate to one or more individual species transport solutions, such as two separate ion pumps each arranged to transport species from a different reservoir 111.

[0065] In some examples, each target species 133,135 comprises a stoichiometric distribution of a set of releasable species 134, 136. For an example scenario, a large number of the first target species 133 each have one of three possible releasable species 134, 50% have A, 30% have B, and 20% have C, such that transporting a large number of copies of the first trigger species 121 that react randomly with a large number of said first target species 133, may upon reacting, release a distribution of releasable species 134 consisting of 50% of releasable species A, 30% of releasable species B, and 20% of releasable species C. Thus, the system 100 may be designed such that transporting multiple copies of one type of trigger species 121 ,122 arranged to trigger the corresponding target species 133,135 may cause release of a plurality of different releasable species 134,136 in a known distribution. In some examples, at least one target species 133,135 comprises a stoichiometric distribution of at least two different releasable species 134,136, or precursors thereof. For these examples, it is to be understood that if the first target species comprises a stoichiometric distribution of a set of releasable species it would herein still be referred to as the first target species 133 by its ability to react with the first species 121 even though its releasable payload can vary.

[0066] In some examples, the device 110 is arranged to independently transport at least a first trigger species 121 and a second trigger species 122 utilizing said transport means 112. In some of these examples, the computer is arranged to transport the at least first trigger species 121 and the second trigger species 122 at at least two different relative rates, such as transporting more of the first trigger species 121 during a first mode and more of the second trigger species 122 during a second mode.

[0067] In some examples, the device 110 is arranged to hold a first and second trigger species 121 , 122 of opposite charge in the reservoir 110, and the transport means 112 is arranged to select the trigger species 121 ,122 to be transported by utilizing electrokinetic transport means.

[0068] It is to be understood that the example in fig. 1a of a system 100 further comprising first trigger species 121 and a second trigger species 122, with corresponding target species 133,135, may be extended to example systems with a third trigger species or more, with corresponding third target species or more. It is to be understood that each type of trigger species is a different type of trigger species, such that an example system arranged for a first, second and third trigger species each with corresponding target species would typically allow for at least three pairs of reacting species.

[0069] Fig. 1 b shows a schematic example system 100 according to the system 100 described in fig. 1a, wherein the device 100 comprises two reservoirs 111. For each reservoir 111 , transport means are arranged to transport trigger species 121 ,122 from said reservoir to an outlet. In the example shown in fig. 1 b, the first trigger species 121 is held in a first reservoir 111 with a first transport means 112 arranged to transport said trigger species 121 into the interaction region 130 via a first outlet 113, and the second trigger species 122 is held in a second reservoir 111 with a second transport means 112 arranged to transport said trigger species 122 into the interaction region 130 via a second outlet 113.

[0070] In some examples, the first species 121 is cationic and the second species 122 is anionic. In some examples, a first part of the transport means 112 is arranged to transport cations and a second part of the transport means 112 is arranged to transport anions. In some examples, the transport means 112 comprise at least one ion selective material, such as an ion selective membrane material.

[0071] In some examples, the device comprises at least three reservoirs 111 arranged to hold at least three trigger species 121 ,122, and wherein the transport means 112 are arranged to transport said trigger species 121 ,122 into the interaction region 130 via at least one outlet 113. It is to be understood that a plurality of reservoirs arranged to hold a plurality of trigger species 121 , 122 means that for each trigger species 121 , 122 at least one of the reservoirs 111 is arranged to hold said trigger species 121 , 122.

[0072] In some examples, each reservoir 111 is arranged to hold one trigger species 121 ,122, In some of these examples, the transport means 112 are arranged to transport each trigger species 121 , 122 from the corresponding reservoir 111 into the interaction region 130 via at least one outlet 113. In some of these examples, the transport means 112 are arranged to transport each trigger species 121 ,122 from the corresponding reservoir 111 into the interaction region 130 via a separate outlet 113, such that the number of reservoirs 111 , trigger species 121 , 122, and outlets 113 are equal.

[0073] Fig. 2 shows schematically a system for orthogonal bioorthogonal release of species with capture species. In the example shown in fig. 2, the interaction region 130 comprises a first capture species 137 and a second capture species 138, wherein said capture species 137,138 are arranged to, upon reacting with the corresponding trigger species 121 ,122, covalently bind said trigger species, thereby stopping said trigger species 121’, 122’ from diffusing out into the environment or reacting with its corresponding target species 133,135. Fig. 2 shows an example where the interaction region 130 has an inner region 132 in which target species 133,135 are covalently bound to a scaffold and / or a hydrogel, and an outer region 131 in which capture species 137,138 are covalently bound to a scaffold and / or a hydrogel, such that the outer region 131 is in contact with the environment, and that any path from the inner region 132 to the environment goes through a minimum distance of the outer region 131 and vice versa. A maximum probability of trigger species diffusing between the inner region 132 and the environment may be controlled by adjusting the concentration of each capture species 137, 138 in the outer region 131 , and said minimum travel path distance through the outer region 131.

[0074] It is to be understood that the examples are not limited to having all capture species 137,138 separated from all target species 133,135. In some examples, one region of a scaffold and / or a hydrogel in the interaction region 130 may be functionalized with both the first target species 133 and the second capture species 138. The use of sets of trigger species 121 , 122, target species 133, 135 and capture species 137,138 with orthogonal reactivity between sets allows for freely functionalizing zones in the scaffold and / or the hydrogel for capture and release reactions to occur at different positions for each set.

[0075] In some examples, at least one capture species 137,138 is covalently bound to at least one region of the scaffold and / or the hydrogel. It is to be understood that said region may be the region of the whole scaffold and / or hydrogel, or any part(s) thereof. In some examples, at least one capture species 137,138 is covalently bound to at least one region of the scaffold and / or the hydrogel, wherein said region with the at least one capture species 137,138 bound is arranged such that any path between the outlet 113 and / or the interaction region 130 to the environment outside the scaffold and / or hydrogel must pass through said region of the scaffold and / or the hydrogel.

[0076] In some examples, said at least one region of the scaffold and / or the hydrogel is functionalized with a concentration of capture species 137,138 of at least 10 pM . In some of these examples, the concentration of capture species 141 is at least 100 pM, at least 1 mM, or at least 10 mM.

[0077] In some examples, each of said at least two types of target species 133,135 is covalently bound to a corresponding region of said scaffold and / or hydrogel, thus allowing one type of trigger species 121 ,122 to cause release of releasable species 134,136 in a specific region in said interaction region 130 functionalized with the corresponding target species 133,135. In some of these examples, each of said at least two types of target species 133,135 is covalently bound to a corresponding separate region of said scaffold and / or hydrogel, thereby allowing targeted release of releasable species 134,136 in a specific region in said interaction region 130 functionalized with the corresponding target species 133,135.

[0078] In some examples, the scaffold and / or the hydrogel is arranged at the device 110. In some examples, the outlet 113 is at the scaffold and / or the hydrogel. In some examples, the scaffold and / or the hydrogel is arranged to surround and / or encapsulate the interaction region 130. In some examples, the scaffold and / or the hydrogel is arranged to surround and / or encapsulate the device 110.

[0079] In some examples, the scaffold and / or the hydrogel is comprised in a vessel. In some examples, the scaffold and / or the hydrogel is in a region within 100 mm of the outlet 113. In some of these examples, the scaffold and / or the hydrogel is in a region within 50 mm, 30 mm, 20 mm, 10 mm, 5 mm, or 2 mm of the outlet 113.

[0080] In some examples, the device 110 is surrounded and / or encapsulated by the scaffold and / or the hydrogel comprising at least one capture species 137,138. In some of these examples, the device 110 is surrounded and / or encapsulated by a first scaffold and / or hydrogel comprising the at least one capture species 137,138, and the outlet 113 and / or inner region 132 is surrounded and / or encapsulated by a second scaffold and / or hydrogel comprising the at least one capture species 137,138. By surrounding the device 110 or the inner region 132 with hydrogel comprising capture species 137,138 may provide protection from corresponding trigger species 121 ,122 escaping into the environment, and from trigger species 121 ,122 in the environment entering the inner region 132.

[0081] In some examples, the interaction region 130 is comprised in a vessel, and the scaffold and / or the hydrogel is arranged in said vessel.

[0082] In some examples, the scaffold and / or the hydrogel is a biocompatible scaffold and / or a biocompatible hydrogel.

[0083] In some examples, the scaffold comprises ureido-pyrimidinones, polyethylene glycol, poly(lactic-co-glycolic acid), polyethylene oxide), poly(vinyl alcohol), alginate, chitosan, and / or hyaluronic acid. In some examples, the scaffold comprises a porous structure.

[0084] In some examples, the hydrogel and / or scaffold comprises linear, branched or hyperbranched hydrogels, self-assembling peptide hydrogels, UV curable crosslinked, or non-crosslinked hydrogels, RGD-modified hydrogels, poly(ethylene glycol), polyethylene oxide (PEO), polyethylene glycol) diacrylate, alginate, cellulose, poly(acrylic acid), hyaluronic acid (HA), Carrageenan, Agarose, Collagen, or derivatives thereof.

[0085] In some examples, the interaction region 130 comprises a capture species 137,138 for each of the trigger species 121 ,122 being arranged to bind said trigger species 121 ,122.

[0086] In some examples, the trigger species 121 ,122, the target species 133,135, and the capture species 137,138 react via a bioorthogonal reaction. In some of these examples, the target species 133,135 is a click-activatable species, such that said releasable species 134,136 is released and becomes biologically active upon reaction of the corresponding trigger species 121 , 122 with target species 133, 135.

[0087] In some examples, the interaction region 130 comprises a modifying species (not shown) arranged to chemically react with at least one trigger species 121 ,122 to modify said trigger species 121 ,121 into a modified inactivated first species (not shown) unable to perform said reaction with the corresponding target species 133,135. In some of these examples, the modifying species is arranged to repeatedly chemically react with multiple copies of a trigger species 121 , 122, and thus allowing a plurality of trigger species 121 ,122 to be inactivated by one modifying species. In some of these examples, the scaffold and / or the hydrogel comprised in the interaction region 130 is functionalized with said modifying species.

[0088] In some examples, the interaction region 130 comprises a repelling species and / or an attracting species (not shown), wherein the repelling species and / or the attracting species are arranged to repel or attract at least one trigger species 121 ,122 to reduce movement of said first species 121 between the environment and an inner region 132 of the interaction region 130. In some of these examples, the repelling species and / or the attracting species are permanently charged moieties comprising sulfonate groups, phosphonium moieties, quaternary ammonium groups, and / or carboxylic acid groups.

[0089] In some examples, the reactions between each trigger species 121 ,122 and the corresponding target species 133,135, capture species 137,138, and modifying species are bioorthogonal.

[0090] In some examples, the device 110 comprises a sensor (not shown) connected to the computer, wherein the computer 115 is arranged to control the transport means based on sensor signals obtained from said sensor. In some of these examples, the sensor is arranged at the outlet 113. In some examples, the sensor is arranged in and / or at a functionalized scaffold and / or hydrogel.

[0091] Fig. 2 illustrates the first and second trigger species 121 , 122 reacting with their corresponding capture species 137,138 and corresponding target species 133,135 via the same reactive site. It is to be understood that the reaction with the corresponding capture species 137,138 may occur at a functional group that is different from the functional group for reacting with the corresponding target species 133,135. In some examples, one capture species 137,138 is arranged to bind two or more trigger species 121 ,122. In some of these examples, one capture species 137,138 is arranged to bind three or more trigger species 121 ,122. In some examples, the first and / or second trigger species 121 ,122 each react with the corresponding capture species 137,138 and corresponding target species 133,135 via the same reactive site. In some examples, for at least one trigger species 121 ,122, the corresponding capture species 137,138 is the corresponding target species 133,135 without its releasable species 134,136, or is a precursor thereof. Utilizing a capture species 137,138 with substantially the same reactive site as the corresponding target species 133,135 may allow for a more predictable system performance as variations in conditions may be assumed to impact the capture species 137,138 and target species 133,135 similarly.

[0092] In some examples, at least one releasable species 134,136 is a drug, toxin and / or bioactive compound. In some examples, the at least one releasable species 134,136 and any scaffolds and / or hydrogel in the interaction region 130 is configured to allow said releasable species 134,136 to exit the interaction region 130. In some examples, the at least one releasable species 134,136 and any scaffolds and / or hydrogel in the interaction region 130 is configured to retain said releasable species 134,136 within the interaction region 130.

[0093] In some examples, the first and / or second capture species 137,138 comprises the active site of the corresponding target species 133,135. In some examples, the first and / or second target species 133,135 comprises, or is a derivative of, the corresponding capture species 137,138.

[0094] In some examples, the first and / or second capture species 137,138 each comprise trans-cyclooctene, TCO, iminosydnone, mesoionic compounds, ortho-carbamoylmethylene silyl-phenolic ethers, tetrazine, Tz, or derivatives thereof.

[0095] Fig. 3 shows a method for orthogonal bioorthogonal release of at least two types of release species. The method 300 comprising

[0096] - providing 310 at least two types of target species comprised in an interaction region, wherein said target species each comprise a corresponding releasable species and / or a precursor thereof, wherein each target species is configured to, upon reacting with a corresponding trigger species, release the corresponding release species;

[0097] - determining 320 a target release of said release species; and

[0098] - providing 330 trigger species into said interaction region based on the determined target release of said release species; wherein said reactions between trigger species and corresponding target species are bioorthogonal reactions, and wherein two or more of said reactions are mutually orthogonal to each other.

[0099] In some examples, providing 310 at least two types of target species comprised in an interaction region comprises providing said at least two types of target species bound to scaffolds and / or a hydrogel comprised in said interaction region.

[0100] In some examples, determining 320 a target release of said release species comprises calculating an amount of trigger species for the target release of release species utilizing computer. In some examples, determining 320 a target release of said release species comprises calculating an amount of trigger species over time utilizing computer. In some of these examples, the target release of said release species comprises instructions for transporting two or more trigger species to the interaction region over a period of time, such as may be depicted by a transport rate vs time plot.

[0101] It is to be understood that the expression “target release of said release species” relates to a desired amount, or rate over time, of release species being released. It is to be understood that “providing 330 trigger species into said interaction region based on the determined target release of said release species” relates to transporting the amount, or rate over time, of corresponding trigger species expected to result in the target release.

[0102] In some examples, determining 320 a target release of said release species comprises calculating a cascaded release over time for releasing two or more release species. In some of these examples, the cascaded release over time comprises instructions for transporting two or more trigger species to the interaction region over a period of time.

[0103] In some examples, providing 330 trigger species into said interaction region comprises transporting said trigger species utilizing transportation means, such as an ionic pump.

[0104] In some examples, the method 300 comprises measuring 340 the state of said interaction region utilizing a sensor arranged to determine sensor data, and wherein determining 320 the target release of said release species is based on obtained sensor data indicative of a measured state in said interaction region. In some examples, the sensor data comprises a concentration of trigger species 121 ,122, and / or a concentration of releasable species 134,136. In some example, the method is a non-therapeutic method.

[0105] In some examples, the method 300 is used to control the exposure of samples or microorganisms to the first species or the corresponding first releasable species.

[0106] In some examples, the method 300 is for performing treatment of a human or non- human animal with controlled exposure of the first trigger species and / or the corresponding first releasable species. In some of these examples, the method 300 further comprises introducing the hydrogel into a human or a non-human animal. In some of these examples, providing 330 the first trigger species into said interaction region comprises implanting the outlet and / or a device comprising said outlet into a human or a non-human animal.

Claims

CLAIMS1 . A system for orthogonal bioorthogonal release of at least two types of releasable species, the system (100) comprises a device (110) comprising one or more reservoirs (111 ), transport means (112) and an outlet (113), wherein said one or more reservoirs (111 ) are arranged to contain at least two types of trigger species (121 ,122), and the system (100) further comprises at least two types of target species (133,135) comprised in an interaction region (130) outside the device (110), wherein each of said target species (133,135) comprise a releasable species (134,136) or a precursor thereof,- wherein said outlet (113) is arranged in and / or at the interaction region (130), and wherein the device (110) is arranged to transport each trigger species (121 ,122) from said one or more reservoirs (111 ) into said interaction region (130) via said outlet (113) utilizing said transport means (112),- wherein said target species (133,135) are arranged to be fluidly connected to said outlet (113), thereby being accessible for said trigger species (121 ,122) transported into the interaction region (130) via said outlet (113),- wherein the device (110) comprises a computer (115) arranged to control said transport means (112) and being arranged to control transporting said trigger species (121 ,122) into said interaction region (130) via said outlet (113),- wherein each type of trigger species (121 ,122) is configured to react with a corresponding target species (133,135), and wherein said reaction comprises releasing the corresponding releasable species (134,136), and- wherein said reactions between trigger species (121 ,122) and corresponding target species (133,135) are bioorthogonal reactions, and wherein two or more of said reactions are mutually orthogonal to each other.

2. The system according to claim 1 , wherein the computer (115) is arranged to control the timing of transport and the amount transported of each trigger species (121 ,122) into the interaction region (130) based on predetermined instructions comprising a schedule for transport rates of two or more trigger species (121 ,122) over a period of time.

3. The system according to claim 1 or 2, comprising a sensor arranged (116) in and / or at the interaction region (130), wherein said sensor (116) is connected to saidcomputer (115), and wherein said computer (115) is arranged to control the timing of transport and the amount transported of each trigger species (121 ,122) into the interaction region (130) based on a sensor signal from said sensor and a set of criteria.

4. The system according to any of claims 1 -3, wherein said computer (115) is arranged to store data indicative of amounts of transported trigger species (121 ,122) and / or sensor signals from the sensor (116), and is arranged to control transport of each trigger species (121 ,122) based on said stored data and a set of criteria.

5. The system according to any preceding claim, wherein the transport means (112) comprise electrokinetic transport means arranged to provide electrokinetic transport of trigger species (121 ,122) from said one or more reservoirs (111 ) into said interaction region (130) via said outlet (113).

6. The system according to any preceding claim, wherein the system (100) comprises a scaffold and / or hydrogel arranged in said interaction region (130), and wherein said at least two types of target species (133,135) are covalently bound to said scaffold and / or hydrogel.

7. The system according to claim 6, wherein each of said at least two types of target species (133,135) is covalently bound to a corresponding region of said scaffold and / or hydrogel, thereby allowing targeted release of releasable species (134,136) in a specific region in said interaction region (130) functionalized with the corresponding target species (133,135).

8. The system according to claim 6 or 7, wherein a region of said scaffold and / or hydrogel comprises at least one type of capture species (137,138) each configured to, upon reacting with a corresponding trigger species (121 ,122), covalently bind and / or modify said trigger species (121 ,122), wherein a modified trigger species is unable to chemically activate the corresponding target species (133,135) so as to cause release of said releasable species (134,136).

9. The system according to claim 8, wherein said at least one capture species (137,138) is covalently bound to at least one region of said scaffold and / or hydrogel.

10. The system according to any preceding claim, wherein said one or more reservoirs (111 ) are arranged to contain at least three types of trigger species (121 ,122), and wherein the device (110) is arranged to transport each trigger species (121 ,122) from said one or more reservoirs (111 ) into said interaction region (130) via said outlet (113) utilizing said transport means (112).

11. The system according to any preceding claim, wherein one reservoir (111 ) is arranged to contain two or more types of trigger species (121 ,122), and wherein the transport means (112) are configured to simultaneously transport said two or more types of trigger species (121 ,122) into said interaction region (130) from said one reservoir (111 ).

12. The system according to any preceding claim, wherein the at least two types of trigger species (121 ,122) each comprise at least one of the following moieties: tetrazine, Tz, fluoride, trans-cyclooctene, TCO, bicyclononyne, BCN, cyclooctyne, and cycloalkynes.

13. The system according to any preceding claim, wherein the at least two types of target species (133,135) each comprise at least one of the following moieties: trans-cyclooctene, TCO, iminosydnone, mesoionic compounds, ortho- carbamoylmethylene silyl-phenolic ethers, tetrazine, Tz, or derivatives thereof.

14. A method for orthogonal bioorthogonal release of at least two types of releasable species, the method (300) comprising- providing (310) at least two types of target species (133,135) comprised in an interaction region (130), wherein said target species (133,135) each comprise a corresponding releasable species (134,136) or a precursor thereof, wherein each target species (133,135) is configured to, upon reacting with a corresponding trigger species (121 ,122), release the corresponding release species (134,136);- determining (320) a target release of said release species (134,136) utilizing computer (115); and- providing (330) trigger species (121 ,122) into said interaction region (130) based on the determined target release of said releasable species (134,136); wherein said reactions between trigger species (121 ,122) and corresponding target species (133,135) are bioorthogonal reactions, and wherein two or more of said reactions are mutually orthogonal to each other.

15. The method according to claim 14, wherein the method (300) comprises measuring (340) the state of said interaction region utilizing a sensor (116) arranged to determine sensor data, and wherein determining (320) the target release of said releasable species (134,136) is based on obtained sensor data indicative of a measured state in said interaction region (130).

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