A system for concentrated targeting using magnetic and oxygen-tactic bacteria and its method of use

Abstract

A system for acquiring, diagnosing, and treating a subject using magnetotactic bacteria is disclosed. The system comprises a processor and a memory storing program code, which, when executed by the processor, causes the processor to acquire image information of a target zone within the subject, apply a magnetic field having a first magnetic field strength to guide the magnetotactic bacteria toward the target zone within the subject, including a hypoxic region, and apply a magnetic field having a second magnetic field strength lower than the first magnetic field strength to enable the magnetotactic bacteria to follow an oxygen gradient that attracts the bacteria toward the hypoxic region before they lose their motility. A method for using the system is also disclosed.

Description

Technical Field

[0001] This disclosure claims priority based on U.S. Provisional Application No. 63 / 351,938, filed on June 14, 2022, and U.S. Provisional Application No. 63 / 351,950, filed on June 14, 2022. Both provisional applications are incorporated herein by reference.

[0002] This disclosure relates to magnetotaxis, and more specifically, to techniques for inducing magneto-aerotactic-responsive bacteria using magnetic fields for imaging, diagnosis, and / or treatment.

Background Art

[0003] A magnetotactic entity is defined as one in which the propulsion source or system responsible for the entity's displacement is either part of the entity itself, or is coupled to or embedded in the entity and not externally moored. Magnetotactic entities include objects or groups of microorganisms, as well as any biological or hybrid systems, including micro- or nano-level systems or structures composed of biologically derived and / or synthetic (chemical, artificial, etc.) materials and / or components. The directional motion of a magnetotactic entity may be affected by torque induced by a directional magnetic field (e.g., the magnetic field of a permanent magnet) or an electromagnetic field (including electromagnetic fields generated by electric currents flowing through conductors). This method of influencing the direction of motion of a magnetotactic entity with a directional magnetic field is referred to herein as magnetotaxis (magnetotactic entities can be functionalized as needed and coupled to other structures). Magnetotactic entities include, but are not limited to, a single or a group (such as a motility group, aggregate, or aggregate) of flagellated magnetotactic bacteria (MTBs). Furthermore, this includes other bacteria or microorganisms that possess self-propulsion capabilities and are affected by directional magnetic fields for the purpose of directional control. These bacteria or microorganisms may have been pre-modified by various methods, including culture conditions and genetic manipulation, or they may have been modified to enable magnetotaxis-based control, such as being bound to or embedded in other entities, such as other cells (including red blood cells), or bound to synthetic structures affected by directional magnetic fields. It may also include entities that exhibit directional motion and are sensitive to magnetotaxis or directional magnetic fields, formed by adding micro- or nano-level components to such bacteria, cells, or other microorganisms. This also includes hybrid entities (consisting of biological and synthetic elements). A directional magnetic field is, for example, a magnetic field that affects the direction of the needle of a magnetic nanocompass.

[0004] However, environmental factors originating from the human body, such as body temperature, pH, and solute concentration, are hostile to bacteria injected into the subject, reducing the activity and motility of magnetotactic bacteria after injection. Therefore, there remains a need to establish methods to improve the targeting ability of magnetotactic bacteria in order to enable them to reach the target zone before they lose motility or die. [Overview of the project]

[0005] This disclosure relates to a method for modulating a magnetic field used to induce magnetotactic bacteria for the treatment, diagnosis, and / or imaging of a subject.

[0006] This method utilizes the oxygenataxis and magnetotaxis properties of magnetotactic bacteria. Magnetotactic bacteria move along an oxygen gradient from a high-oxygen side to a low-oxygen side, and their movement is also induced by a magnetic field. Magnetotactic bacteria have an affinity for low oxygen concentrations and are attracted to low-oxygen regions (not limited to tumors, but such areas) within the subject where almost no oxygen is present. Magnetotactic bacteria can autonomously move to low-oxygen regions at the target site using their own propulsion system (e.g., flagella). Such movement can occur when magnetotactic bacteria are placed under an oxygen gradient, regardless of the presence of a magnetic field. At this time, the relaxation time of the run-and-tumble motion increases, so the direction of bacterial movement changes from the direction of the magnetic field.

[0007] Therefore, this method relates to adjusting the strength of a magnetic field with the aim of changing the movement pattern of bacteria in accordance with the strength of the magnetic field. The higher the magnetic field strength, the more run-and-reverse motion, a movement that causes displacement of bacteria as they move in accordance with the magnetic field, is promoted. The lower the magnetic field strength, the more magnetotactic bacteria perform run-and-tumble motion during the process of seeking out the oxygen gradient, which leads them to the hypoxic region of the subject.

[0008] This method improves the targeting of magnetotactic bacteria in patients by utilizing the fact that the movement pattern of magnetotactic bacteria changes in response to environmental factors (e.g., magnetic field strength). By changing the magnetic field strength, it is possible to control the movement of bacteria based on run-and-tumble motion. When the magnetic field strength is high, the change in the direction of movement of bacteria associated with run-and-tumble motion decreases, and when the magnetic field strength is low, the change in the direction of movement associated with run-and-tumble motion increases, thereby enabling bacteria to reach hypoxic regions.

[0009] Run-and-reverse movement can typically be described as a mode of movement in which bacteria make long runs in one direction, and during this run, they reverse direction at a specific average non-periodic frequency, resulting in a forward displacement of the bacteria. In contrast, run-and-tumble movement can be characterized as a mode of movement in which the alignment of the direction of movement of magnetotropic bacteria with the direction of the magnetic field is disrupted due to a tumble action (an action in which the direction of movement of bacteria changes from the direction of the magnetic field), and then, in the run state, the alignment with respect to the magnetic field is restored through a relaxation process. If the run state time is shorter than the relaxation time, there is not enough time for the alignment with the magnetotropic bacteria to be restored by the action of the magnetic field before the next tumble action occurs, and the direction of movement of the bacteria fluctuates with respect to the direction of the magnetic field.

[0010] In the method described herein, two to three modes can be employed as modes for inducing magnetotactic and oxytactic bacteria. In the first mode, the magnetic field strength is set so that magnetotaxis is dominant, and the prominent movement of the bacteria is mainly run-and-reverse motion, with the run state during run-and-tumble motion becoming longer relative to the relaxation time as the magnetic field strength increases. In the second mode, by lowering the magnetic field strength, bacterial movement may be caused by both magnetotaxis and oxytaxis, and in this case, the bacteria exhibit more directional changes due to run-and-tumble motion. In the third mode, by lowering the magnetic field strength even further than in the second mode, bacterial movement becomes mainly due to oxytaxis, and therefore the bacteria exhibit directional changes mainly due to run-and-tumble motion.

[0011] It should be understood that, when inducing bacteria, one mode can be used multiple times, and the order of modes can be changed depending on the bacterial migration path, the characteristics of the target site, the location of the target site, etc. The user can select one or more modes in different orders to reach the target site and the desired hypoxic region distribution.

[0012] Run-and-reverse movement allows magnetotactic bacteria to move to different locations within the subject (for example, mainly used to create relatively long-distance displacements). On the other hand, run-and-tumble movement enhances the bacteria's exploration of hypoxic areas. The magnetic field strength can be appropriately adjusted according to the desired movement pattern of the magnetotactic bacteria, the characteristics of the target within the subject (e.g., tissue density, size, degree of concentration of hypoxic areas, etc.), the distribution of bacterial velocity before injection, the decay of bacterial velocity after injection (regression), the volume and dimensions of the bolus, the concentration of magnetotactic bacteria in the bolus, and the distance between the injection site and the target.

[0013] In some embodiments, when magnetotactic bacteria are induced by a magnetic field, the bacteria move faster with increasing magnetic field strength. Therefore, this high magnetic field strength is maintained to induce the bacteria until they reach a target site with one or more oxygen gradients (e.g., oxygen gradients resulting from one or more hypoxic regions). Once the magnetotactic bacteria reach the target site, the magnetic field strength is reduced, and the oxygen gradient alters the movement pattern of the magnetotactic bacteria. In the altered movement pattern, the magnetotactic bacteria perform run-and-tumble movements, resulting in a change in the direction of movement. This change in direction allows the magnetotactic bacteria to reach hypoxic regions, where they may remain until their post-injection velocity drops to zero (e.g., drug bound to the magnetotactic bacteria may be deposited).

[0014] Therefore, when magnetic and oxygen-tactic bacteria are bound to a diagnostic agent, imaging agent, and / or therapeutic agent, if the magnetic and oxygen-tactic bacteria become immobile (e.g., die) in a hypoxic area of ​​the tumor, the diagnostic agent, imaging agent, and / or therapeutic agent are deposited in that hypoxic area. In other words, targeted delivery of the diagnostic agent, imaging agent, and / or therapeutic agent occurs, making it possible to reduce the amount of the agent required for delivery to the subject. This reduces toxicity to the subject.

[0015] In some embodiments, the velocity distribution of magnetotactic bacteria contained in a bolus or sample can be used to predict the behavior of magnetotactic bacteria when injected into a subject and exposed to a magnetic field. For example, if the velocity distribution is narrow, the initial velocities of the magnetotactic bacteria are similar to each other, resulting in a small spread of magnetotactic bacteria when injected into a patient and exposed to a magnetic field (and therefore a small target volume). In contrast, if the velocity distribution is wide, some of the magnetotactic bacteria move at a significantly slower speed than others when exposed to a magnetic field, resulting in a larger spread of magnetotactic bacteria or distance between bacteria (and target volume) over time after injection and exposure to a magnetic field. When bacterial spread is large, targeting to hypoxic regions can become more diffusive when the magnetic field strength is adjusted to increase run-and-tumble motion for searching for hypoxic regions. As bacteria diffuse to more hypoxic regions, the concentration per hypoxic region decreases. In contrast, if the spread of magnetotaxis fibers after injection is small (due to a small velocity distribution), targeting of hypoxic regions can become more concentrated when the magnetic field strength is adjusted to increase run-and-tumble motion for searching for hypoxic regions. This is because when the magnetic field strength is reduced (which promotes changes in the direction of bacteria away from the magnetic field direction due to increased run-and-tumble motion for searching for hypoxic regions), all injected bacteria have already traveled approximately the same distance from the injection site and are located in a narrow volume region within the subject.

[0016] As a result, by utilizing the velocity distribution of magnetotactic bacteria (for example, when preparing injection solutions or selecting injection solutions or samples), it becomes possible to increase or decrease the number of target hypoxic regions and adjust the concentration of magnetotactic bacteria in each target hypoxic region. If it is necessary to dilute the magnetotactic bacteria to more hypoxic regions, a sample with a large velocity distribution can be prepared or selected. If it is necessary to concentrate the magnetotactic bacteria to fewer hypoxic regions (for example, without reducing dilution), a sample with a small velocity distribution can be prepared or selected.

[0017] In one embodiment, with respect to the applied magnetic field, the magnetic field generated by using three pairs of magnetic coils provides a directional torque that can be defined along one or more of the three axes. Each pair of magnetic coils is positioned relative to one axis. Each pair of magnetic heads can generate a three-dimensional convergence point in which a magnetotactic entity moves and converges in that direction. A three-dimensional convergence point (CP) in a magnetic field is a point without spatial boundaries where entities moving along the direction of the magnetic field within a cohesive region (AZ) converge. The magnetic field at the convergence point is substantially zero, and within the cohesive region surrounding the convergence point, the effective magnetic field is directed towards the convergence point from all directions. Since the magnetic field is not a point source, by changing at least one magnetic field source over time, the entity can be moved toward the convergence point and kept near the convergence point.

[0018] Therefore, by maintaining a constant (static) magnetic field along any two axes (x, y, or z) and changing the direction of the magnetic field along the remaining axis, a convergence point can be generated because these two axes remain constant. Similarly, a convergence point can also be generated by maintaining one axis constant and simultaneously (synchronously) changing the directions of the remaining two axes in a time-division manner, or by changing them with a phase delay. However, this change must occur at a frequency to which magnetotactic bacteria can respond appropriately (i.e., a low frequency of about 0.1 Hz to 5 Hz, more preferably about 0.5 Hz). It is also possible to change the direction of all three axes simultaneously in a time-division manner, or with a phase delay. Any combination is possible as long as the magnetic field gradient along at least one axis (x, y, or z) is sequentially changed in a time-division manner at a switching speed suitable for the reaction time of magnetotactic bacteria. U.S. Patent No. 9,905,347, incorporated herein by reference, describes a system for inducing a magnetotactic entity within a subject. Also described in U.S. Patent No. 9,905,347 is a system and method for generating a three-dimensional convergence point using at least three pairs of magnetic field sources arranged along three axes or three planes.

[0019] A broader embodiment relates to a method for acquiring at least one of imaging information, diagnosis, and treatment of a subject using self-propelled, adaptable magnetic-oxygen-tactic bacteria. The method includes acquiring imaging information of a target zone within a subject, wherein a bolus of magnetic-oxygen-tactic bacteria conjugated to at least one of a therapeutic agent, a diagnostic agent, and an imaging agent is injected into the subject; applying a magnetic field having a first magnetic field strength to guide and displace the magnetic-oxygen-tactic bacteria via magnetotaxis toward the target zone having a hypoxic region; and applying a magnetic field having a second magnetic field strength lower than the first magnetic field strength to enable the magnetic-oxygen-tactic bacteria to follow an oxygen gradient that draws the bacteria toward the hypoxic region, wherein the change in the direction of the magnetic field from the direction of movement of the magnetic-oxygen-tactic bacteria increases at the second magnetic field strength compared to the first magnetic field strength, thereby enabling at least one of treatment, diagnosis, and imaging of the subject.

[0020] In some embodiments, the method includes obtaining velocity distribution information of magnetotactic bacteria in the bolus and predicting the spread of the magnetotactic bacteria after injection into the subject when the magnetic field strength is the first magnetic field strength.

[0021] In some embodiments, the bolus is selected from a plurality of magnetic-oxygen-tactic bacterial solutions according to the pre-injection velocity distribution information of magnetic-oxygen-tactic bacteria in the solution, and the selected solution contains magnetic-oxygen-tactic bacteria with a large velocity distribution, depending on the desired target volume for the magnetic-oxygen-tactic bacteria, and the larger the target volume, the more hypoxic regions associated with the target zone can be targeted.

[0022] In some embodiments, when it is required to increase the target volume and thus increase the number of hypoxic regions targeted by the magnetotactic bacteria, the solution with the widest pre-injection velocity distribution of the magnetotactic bacteria is selected from among the multiple solutions.

[0023] In some embodiments, when it is required to perform concentrated targeting to increase the concentration of magneto-oxygen motile bacteria targeting one or more hypoxic regions, among the plurality of solutions, the solution with the narrowest velocity distribution before injection of the magneto-oxygen motile bacteria is selected.

[0024] In some embodiments, the first and second magnetic field strengths are determined taking into account the attenuation of the velocity after injection of the magneto-oxygen motile bacteria.

[0025] In some embodiments, the method includes estimating the position of the magneto-oxygen motile bacteria when the velocity after injection of the magneto-oxygen motile bacteria has decayed to zero.

[0026] In some embodiments, the method includes calculating the distance between the injection site of the bolus and a part of the aggregation region or tumor, and applying a magnetic field having the second magnetic field strength is based on the calculated distance.

[0027] In some embodiments, the magneto-oxygen motile bacteria are bound to an imaging agent, and the imaging agent is a contrast agent.

[0028] In some embodiments, the contrast agent is gadolinium.

[0029] In some embodiments, acquisition of image information of a subject's tumor is performed using an MRI or CT scanner.

[0030] In some embodiments, the image information includes information regarding the arteries of the subject in order to avoid puncturing an artery when injecting a bolus of the magneto-oxygen motile bacteria into the peripheral region of the tumor.

[0031] In some embodiments, the first and second magnetic field strengths are determined and adjusted based on the volume of the bolus, the distance between the administration position of the bolus and the aggregation region, and the elapsed time after injection.

[0032] In some embodiments, the first and second magnetic field strengths are further determined and adjusted based on the concentration of magnetotactic bacteria in the bolus.

[0033] In some embodiments, the bolus of magnetic-oxytomic bacteria is injected into the periphery of the tumor.

[0034] In some embodiments, multiple boluses are injected into different locations in the subject.

[0035] In some embodiments, the method includes reducing the magnetic field strength of the magnetic field to a second magnetic field strength, and then further reducing the magnetic field strength of the magnetic field to a third magnetic field strength lower than the second magnetic field strength, such that the change from the direction of the magnetic field in the direction of movement of the magnetotactic bacteria is greater at the third magnetic field strength than at the second magnetic field strength.

[0036] In some embodiments, the value of the second magnetic field strength is greater than or equal to 0 Gauss and less than 5 Gauss.

[0037] In some embodiments, the first magnetic field strength is at least 15 gauss.

[0038] In some embodiments, the second magnetic field strength is less than 15 gauss and greater than or equal to 5 gauss.

[0039] In some embodiments, the value of the second magnetic field strength is greater than or equal to 0 Gauss and less than 5 Gauss.

[0040] Another broader aspect relates to a system for performing at least one of the acquisition of imaging information, diagnosis, and treatment of a subject using magnetotactic bacteria, wherein the magnetotactic bacteria are adapted to move autonomously by run-and-reverse and run-and-tumble movements after a bolus of magnetotactic bacteria is injected into the subject, and are bound to at least one of a therapeutic agent, a diagnostic agent, and an imaging agent. The system comprises a processor and a memory storing program code, the program code, when executed by the processor, causes the processor to perform the following actions: acquire image information of the target zone of the subject; apply a magnetic field having a first magnetic field strength to guide and displace the magnetotactic bacteria toward the target zone having a hypoxic region via magnetotaxis; and apply a magnetic field having a second magnetic field strength lower than the first magnetic field strength to enable the magnetotactic bacteria to follow the oxygen gradient that attracts the bacteria toward the hypoxic region, wherein the change in the direction of the magnetic field relative to the direction of movement of the magnetotactic bacteria increases at the second magnetic field strength compared to the first magnetic field strength.

[0041] In some embodiments, the system includes a user input interface, and the program code, when executed by the processor, further includes instructions that cause the processor to receive instructions input from a user to the user input interface and to generate the acquired image information.

[0042] In some embodiments, the system comprises one or more magnetic sources.

[0043] In some embodiments, the one or more magnetic sources include three pairs of magnetic coils, each pair of the three pairs of magnetic coils is positioned relative to one of three different axes: x, y, and z.

[0044] In some embodiments, the system includes an imaging device.

[0045] In some embodiments, the imaging device is an MRI device.

[0046] In some embodiments, the first magnetic field strength is at least 15 gauss.

[0047] In some embodiments, the second magnetic field strength is less than 15 gauss and greater than or equal to 5 gauss.

[0048] In some embodiments, the program code for determining the first and second magnetic field strengths takes into account the volume of the bolus, the distance between the bolus injection site and the aggregation region, and the elapsed time after injection.

[0049] In some embodiments, the program code for determining the first and second magnetic field strengths further takes into account the concentration of magnetotactic bacteria in the bolus.

[0050] In some embodiments, the memory includes program code that, when executed by the processor, causes the processor to reduce the magnetic field strength of the magnetic field to a second magnetic field strength, and then further reduce the magnetic field strength of the magnetic field to a third magnetic field strength lower than the second magnetic field strength, such that the change from the direction of the magnetic field in the direction of movement of the magnetotactic bacteria increases at the third magnetic field strength compared to the second magnetic field strength.

[0051] In some embodiments, the memory includes program code that, when executed by the processor, causes the processor to calculate the distance between the bolus injection site and the aggregated region or portion of the tumor, and the application of the magnetic field having the second magnetic field strength is performed taking the calculated distance into account.

[0052] In some embodiments, the memory includes program code that, when executed by the processor, causes the processor to estimate the position of the magnetotactic bacteria when their post-injection velocity drops to zero.

[0053] Other broad embodiments relate to a non-temporary storage medium storing instructions executable by a computer device, the storage medium including at least one instruction for acquiring image information of a target zone within a subject; at least one instruction for applying a magnetic field having a first magnetic field strength to guide and displace magnetotactic bacteria toward the target zone within the subject having a hypoxic region; and at least one instruction for applying a magnetic field having a second magnetic field strength lower than the first magnetic field strength to enable the magnetotactic bacteria to follow an oxygen gradient that attracts the bacteria toward the hypoxic region, wherein the change in the direction of the magnetic field from the direction of movement of the magnetotactic bacteria increases at the second magnetic field strength compared to the first magnetic field strength.

[0054] Other broad embodiments relate to a method for selecting a sample of magnetotactic bacteria conjugated to at least one of imaging agents, targeting agents, and diagnostic agents, in accordance with concentrated or dispersed targeting by the magnetotactic bacteria to target zones within a subject having hypoxic regions, wherein the magnetotactic bacteria are guided to target sites within the subject using a magnetic field by magnetotaxis and further explore hypoxic regions within the subject by oxygenotaxis. The method includes selecting a sample of magnetotactic bacteria from a plurality of samples based on the pre-injection velocity distribution of the magnetotactic bacteria in the sample, wherein a wider pre-injection velocity distribution is selected as the target volume and number of target hypoxic regions increase, and a narrower pre-injection velocity distribution is selected as the target volume and number of target hypoxic regions decrease, for more concentrated targeting by the magnetotactic bacteria.

[0055] Other broad embodiments relate to a method for preparing a solution of magnetotactic bacteria to be administered parenterally to a subject in order to achieve a desired level of target volume at a target site within the subject, wherein the administered magnetotactic bacteria are guided to the target site within the subject using a magnetic field via magnetotaxis and further explore hypoxic regions within the subject via oxygenotaxis. The method comprises selecting magnetotactic bacteria based on their self-propelled velocity via magnetotaxis to obtain a population of magnetotactic bacteria selected for the solution, wherein the velocity distribution of the selected magnetotactic bacteria is large in proportion to the desired target volume, and the method further comprises preparing a solution containing the selected magnetotactic bacteria, wherein the selected magnetotactic bacteria are bound to at least one of imaging agents, contrast agents, and diagnostic agents.

[0056] Other broad embodiments relate to a method for controlling the distribution of magnetotropic bacteria that target tumor regions within a subject using a magnetic field for at least one of imaging information, diagnosis, and treatment. The method involves selecting a magnetic field strength according to the target distribution zone of the magnetotropic bacteria in the tumor, wherein a higher strength is selected for wider distribution zones and a lower strength for narrower distribution zones; generating a magnetic field of the selected strength to induce the magnetotropic bacteria, wherein the velocity distribution of the magnetotropic bacteria under the magnetic field causes them to cover the target distribution zone.

[0057] In some embodiments, the target distribution zone includes a hypoxic region.

[0058] In some embodiments, if the target distribution zone is concentrated at the injection site of the magnetotactic bacteria in the subject, the selected intensity is 0 Gauss.

[0059] In some embodiments, if the target distribution zone is large, the selected intensity is 15 Gauss so that the distribution of the magnetotactic bacteria increases and spreads throughout the entire target distribution zone.

[0060] Other broad embodiments relate to methods for performing at least one of the following on a subject: imaging, diagnosis, and treatment, using self-propelled magnetotactic bacteria. The method includes acquiring imaging information of a target zone within a subject, wherein a bolus of magnetotactic bacteria conjugated to at least one of a therapeutic agent, a diagnostic agent, and an imaging agent is injected into the subject; applying a magnetic field having a first magnetic field strength to guide and displace the magnetotactic bacteria via magnetotaxis toward the target zone having a hypoxic region; and applying a magnetic field having a second magnetic field strength lower than the first magnetic field strength to enable the magnetotactic bacteria to follow an oxygen gradient that draws the bacteria toward the hypoxic region, wherein the change in the direction of the magnetic field from the direction of movement of the magnetotactic bacteria increases at the second magnetic field strength compared to the first magnetic field strength, thereby enabling at least one of the following on a subject: treatment, diagnosis, and imaging.

[0061] In some embodiments, the method includes obtaining velocity distribution information of magnetotactic bacteria in the bolus and predicting the spread of the magnetotactic bacteria after injection into the subject when the magnetic field strength is the first magnetic field strength.

[0062] In some embodiments, the bolus is selected from a plurality of magnetic-oxygen-tactic bacterial solutions according to the pre-injection velocity distribution information of magnetic-oxygen-tactic bacteria in the solution, and the selected solution contains magnetic-oxygen-tactic bacteria with a large velocity distribution, depending on the desired target volume for the magnetic-oxygen-tactic bacteria, and the larger the target volume, the more hypoxic regions associated with the target zone can be targeted.

[0063] In some embodiments, when it is required to increase the target volume and thus increase the number of hypoxic regions targeted by the magnetotactic bacteria, the solution with the widest pre-injection velocity distribution of the magnetotactic bacteria is selected from among the multiple solutions.

[0064] In some embodiments, when intensive targeting is required to increase the concentration of magnetotactic bacteria targeting one or more hypoxic regions, the solution with the narrowest pre-injection velocity distribution of magnetotactic bacteria is selected from among the multiple solutions.

[0065] In some embodiments, the first and second magnetic field strengths are determined taking into account the decay of the injection velocity of the magnetotactic bacteria.

[0066] In some embodiments, the method includes estimating the position of the magnetotactic bacteria when their injection velocity decays to zero.

[0067] In some embodiments, the method includes calculating the distance between the injection site of the bolus and a portion of the aggregated area or tumor, and applying a magnetic field having the second magnetic field strength based on the calculated distance.

[0068] In some embodiments, the magnetic-oxytomic bacteria are bound to an imaging agent, which is a contrast agent.

[0069] In some embodiments, the contrast agent is gadolinium.

[0070] In some embodiments, imaging information of the subject's tumor is obtained using an MRI or CT scanner.

[0071] In some embodiments, the image information includes information about the subject's arteries in order to avoid puncturing arteries when injecting the bolus of the magnetic-oxytomic bacteria into the area surrounding the tumor.

[0072] In some embodiments, the first and second magnetic field intensities are determined and adjusted based on the volume of the bolus, the distance between the bolus injection site and the aggregation region, and the elapsed time after injection.

[0073] In some embodiments, the first and second magnetic field strengths are further determined and adjusted based on the concentration of magnetotactic bacteria in the bolus.

[0074] In some embodiments, the bolus of magnetic-oxytomic bacteria is injected into the periphery of the tumor.

[0075] In some embodiments, multiple boluses are injected into different locations in the subject.

[0076] In some embodiments, the method includes reducing the magnetic field strength of the magnetic field to a second magnetic field strength, and then further reducing the magnetic field strength of the magnetic field to a third magnetic field strength lower than the second magnetic field strength, such that the change from the direction of the magnetic field in the direction of movement of the magnetotactic bacteria is greater at the third magnetic field strength than at the second magnetic field strength.

[0077] In some embodiments, the value of the second magnetic field strength is greater than or equal to 0 Gauss and less than 5 Gauss.

[0078] In some embodiments, the first magnetic field strength is at least 15 gauss.

[0079] In some embodiments, the second magnetic field strength is less than 15 gauss and greater than or equal to 5 gauss.

[0080] In some embodiments, the value of the second magnetic field strength is greater than or equal to 0 Gauss and less than 5 Gauss.

[0081] Another broader aspect relates to a system for performing at least one of the acquisition of imaging information, diagnosis, and treatment of a subject using magnetotactic bacteria, wherein the magnetotactic bacteria are adapted to move autonomously by run-and-reverse and run-and-tumble movements after a bolus of magnetotactic bacteria is injected into the subject, and are bound to at least one of a therapeutic agent, a diagnostic agent, and an imaging agent. The system comprises a processor and a memory storing program code, the program code, when executed by the processor, causes the processor to perform the following actions: acquire image information of the target zone of the subject; apply a magnetic field having a first magnetic field strength to guide and displace the magnetotactic bacteria toward the target zone having a hypoxic region via magnetotaxis; and apply a magnetic field having a second magnetic field strength lower than the first magnetic field strength to enable the magnetotactic bacteria to follow the oxygen gradient that attracts the bacteria toward the hypoxic region, wherein the change in the direction of the magnetic field relative to the direction of movement of the magnetotactic bacteria increases at the second magnetic field strength compared to the first magnetic field strength.

[0082] In some embodiments, the system includes a user input interface, and the program code, when executed by the processor, further includes instructions that cause the processor to receive instructions input from a user to the user input interface and to generate the acquired image information.

[0083] In some embodiments, the system comprises one or more magnetic sources.

[0084] In some embodiments, the one or more magnetic sources include three pairs of magnetic coils, each pair of the three pairs of magnetic coils is positioned relative to one of three different axes: x, y, and z.

[0085] In some embodiments, the system includes an imaging device.

[0086] In some embodiments, the imaging device is an MRI device.

[0087] In some embodiments, the first magnetic field strength is at least 15 gauss.

[0088] In some embodiments, the second magnetic field strength is less than 15 gauss and greater than or equal to 5 gauss.

[0089] In some embodiments, the program code for determining the first and second magnetic field strengths takes into account the volume of the bolus, the distance between the bolus injection site and the aggregation region, and the elapsed time after injection.

[0090] In some embodiments, the program code for determining the first and second magnetic field strengths further takes into account the concentration of magnetotactic bacteria in the bolus.

[0091] In some embodiments, the memory further includes program code, when executed by the processor, that causes the processor to reduce the magnetic field strength of the magnetic field to a second magnetic field strength, and then further reduce the magnetic field strength of the magnetic field to a third magnetic field strength lower than the second magnetic field strength, such that the change from the direction of the magnetic field in the direction of movement of the magnetotactic bacteria increases at the third magnetic field strength compared to the second magnetic field strength.

[0092] In some embodiments, the memory includes program code that, when executed by the processor, causes the processor to calculate the distance between the bolus injection site and the aggregated region or portion of the tumor, and the application of the magnetic field having the second magnetic field strength is performed taking the calculated distance into account.

[0093] In some embodiments, the memory includes program code that, when executed by the processor, causes the processor to estimate the position of the magnetotactic bacteria when their post-injection velocity drops to zero.

[0094] Other broad embodiments relate to a non-temporary storage medium storing instructions executable by a computer device, the storage medium including at least one instruction for acquiring image information of a target zone within a subject; at least one instruction for applying a magnetic field having a first magnetic field strength to guide and displace magnetotactic bacteria toward the target zone within the subject having a hypoxic region; and at least one instruction for applying a magnetic field having a second magnetic field strength lower than the first magnetic field strength to enable the magnetotactic bacteria to follow an oxygen gradient that attracts the bacteria toward the hypoxic region, wherein the change in the direction of the magnetic field from the direction of movement of the magnetotactic bacteria increases at the second magnetic field strength compared to the first magnetic field strength.

[0095] Other broad embodiments include a method for selecting a sample of magnetotactic bacteria conjugated to at least one of imaging agents, targeting agents, and diagnostic agents, in accordance with concentrated or dispersed targeting by the magnetotactic bacteria to target zones within a subject having hypoxic regions, wherein the magnetotactic bacteria are guided to target sites within the subject by magnetotaxis using a magnetic field and explore hypoxic regions within the subject by oxygenotaxis, and the method includes selecting a sample of magnetotactic bacteria from a plurality of samples based on the pre-injection velocity distribution of the magnetotactic bacteria in the sample, wherein a wider pre-injection velocity distribution is selected as the target volume and number of target hypoxic regions increase, and a narrower pre-injection velocity distribution is selected as the target volume and number of target hypoxic regions decrease, for more concentrated targeting by the magnetotactic bacteria.

[0096] Other broad embodiments relate to a method for preparing a solution of magnetotactic bacteria for parenteral administration to a subject in order to achieve a desired level of target volume at a target site in the subject, wherein the administered magnetotactic bacteria are guided to the target site in the subject using a magnetic field via magnetotaxis and explore hypoxic regions in the subject via oxygenotaxis. The method involves selecting magnetotactic bacteria based on their magnetotactic self-propulsion speed to obtain a population of magnetotactic bacteria selected for the solution, wherein the velocity distribution of the selected magnetotactic bacteria is large in proportion to the desired target volume, and a solution containing the selected magnetotactic bacteria is prepared, wherein the selected magnetotactic bacteria are bound to at least one of imaging agents, contrast agents, and diagnostic agents.

[0097] Other broad embodiments relate to methods for controlling the distribution of magnetotropic bacteria to target tumor regions within a subject using a magnetic field for at least one of imaging, diagnosis, and treatment. The method involves selecting the intensity of a magnetic field according to the target distribution zone of the magnetotropic bacteria in the tumor, wherein a higher intensity is selected for wider distribution zones and a lower intensity for narrower distribution zones, and generating a magnetic field of the selected intensity to induce the magnetotropic bacteria, wherein, according to the velocity distribution of the magnetotropic bacteria, the magnetotropic bacteria cover the target distribution zone under the magnetic field.

[0098] In some embodiments, the target distribution zone includes a hypoxic region.

[0099] In some embodiments, if the target distribution zone is concentrated at the injection site of the magnetotactic bacteria in the subject, the selected intensity is 0 Gauss.

[0100] In some embodiments, when the target distribution zone is large, the selected intensity is 15 Gauss so that the dispersion of the magnetotactic bacteria is large and they spread throughout the entire target distribution zone.

[0101] Other broad embodiments relate to a container for injecting a given amount of magnetotactic bacteria into a subject for magnetic field-based treatment, diagnosis, and / or imaging, the container comprising an identifier configured to be scanned to provide one or more characteristics of the magnetotactic bacteria in the container, and the magnetotactic bacteria disposed within the container.

[0102] In some embodiments, the one or more characteristics include one or more of the following: the attenuation of the activity of the magnetotactic bacteria, the density of the magnetotactic bacteria, the polarity or polarity ratio of the magnetotactic bacteria, the response time of the magnetotactic bacteria to a change in the orientation of the magnetic field, and the maximum velocity or velocity distribution of the magnetotactic bacteria.

[0103] In some embodiments, the identifier is a barcode.

[0104] In some embodiments, the identifier is a QR (quick response) code.

[0105] Other broad embodiments relate to methods for characterizing one or more characteristics of magnetotactic bacteria in a sample of magnetotactic bacteria, the methods comprising generating a video of the magnetotactic bacteria observed under a microscope in a magnetic field having known magnetic field strength and orientation, and analyzing the behavior of the magnetotactic bacteria in the video to identify the one or more characteristics.

[0106] In some embodiments, the one or more characteristics include the velocity distribution of the magnetotactic bacteria, and the analysis includes calculating the distance traveled by the magnetotactic bacteria among the magnetotactic bacteria in a predetermined time and dividing the distance by the time.

[0107] In some embodiments, the one or more characteristics include a decrease in the activity of the magnetotactic bacteria, the analysis including repeatedly calculating the distance traveled by the magnetotactic bacteria over a predetermined time at different time intervals, and measuring the velocity of the magnetotactic bacteria over that time by dividing the distance by that time, the decrease in activity is identified from the decrease in velocity measured at different time intervals.

[0108] In some embodiments, the one or more characteristics include the polarity of the magnetotactic bacteria, and the analysis includes measuring the percentage of bacteria moving in a direction corresponding to the north or south of the magnetic field.

[0109] Other broader aspects include a method for inducing self-propelled magnetotactic bacteria, wherein imaging information of the location of a target zone within a specimen is acquired, wherein a bolus of magnetotactic bacteria is injected into the specimen, the magnetotactic bacteria being bound to at least one of a therapeutic agent, a diagnostic agent, and an imaging agent; a magnetic field having a first magnetic field strength is applied to guide and displace the magnetotactic bacteria via magnetotaxis toward the target zone having a hypoxic region, based on the location; a magnetic field having a second magnetic field strength lower than the first magnetic field strength is applied to enable the magnetotactic bacteria to follow an oxygen gradient that draws the bacteria toward the hypoxic region; and in at least one of the treatment, diagnosis, and imaging of the specimen, the change from the direction of the magnetic field in the direction of movement of the magnetotactic bacteria increases at the second magnetic field strength compared to the first magnetic field strength. [Brief explanation of the drawing]

[0110] This invention will be better understood by referring to the attached drawings and reading the detailed description of the invention below.

[0111] [Figure 1]This shows exemplary direction-controllable magnetotactic bacteria bound to an imaging agent, and exemplary direction-controllable magnetotactic bacteria bound to a therapeutic agent.

[0112] [Figure 2] This shows exemplary direction-controllable magnetic-oxygen-tactic bacteria conjugated to both imaging and therapeutic agents.

[0113] [Figure 3] This diagram maps exemplary hypoxic regions through which magnetotactic bacteria migrate from the administration site.

[0114] [Figure 4] This is another diagram mapping the hypoxic regions that magnetotactic bacteria migrate through from the administration site.

[0115] [Figure 5] This is an illustrative graph showing the standard deviation of the velocity of therapeutic complexes formed when magnetotactic bacteria bind to any agent (e.g., therapeutic agents, diagnostic agents, imaging agents). It indicates that when the velocity of the bacteria exceeds a velocity threshold, applying a magnetic field of sufficient strength to promote magnetotaxis will cause at least a large portion of the magnetotactic bacteria to pass through one or more hypoxic zones, resulting in a more concentrated induction of the bacteria into the target hypoxic region further away from the administration site.

[0116] [Figure 6] Another exemplary graph showing the standard deviation of the rates of therapeutic complexes formed when magnetotactic bacteria bind to any agent (e.g., therapeutic agents, diagnostic agents, imaging agents), illustrating that if only a portion of the magnetotactic bacteria have rates above the rate threshold, then when a magnetic field strength that promotes magnetotaxis is applied, the magnetotactic bacteria will diffuse or be diluted more widely over a larger hypoxic region.

[0117] [Figure 7]This flowchart illustrates an exemplary method for modulating a magnetic field to induce magnetotactic bacteria for the treatment, diagnosis, and / or imaging of a subject.

[0118] [Figure 8] This is a block diagram of an exemplary system for targeted imaging, diagnosis, and / or treatment of a subject using magnetic and oxygen-tactic bacteria.

[0119] [Figure 9A] This is a schematic diagram representing a target site with a large target volume.

[0120] [Figure 9B] This is a schematic diagram representing a target site with a small target volume. [Modes for carrying out the invention]

[0121] This disclosure describes a method for modulating a magnetic field used to induce magnetotactic bacteria for the treatment, diagnosis, and / or imaging of a subject. Magnetotactic bacteria respond to both a magnetic field (magnetotaxis) and an oxygen gradient (oxytaxis).

[0122] Therefore, by increasing or decreasing the magnetic field strength according to the position of magnetotactic bacteria, the movement pattern of magnetotactic bacteria can be altered. At magnetic field strengths that induce magnetotaxis, displacement due to run-and-reverse motion is promoted (because the run state lasts longer than the relaxation time, the deviation of the bacterial movement direction from the direction of the magnetic field is reduced), while at magnetic field strengths that primarily induce oxygenataxis, the deviation of the bacterial movement direction from the direction of the magnetic field is promoted via run-and-tumble motion.

[0123] It should be understood that magnetotactic bacteria move autonomously while being guided by the torque of the magnetic field. If it is determined or estimated that the magnetotactic bacteria have reached a target site with an oxygen gradient (e.g., a tumor area), the magnetic field strength may be reduced. At lower magnetic field strengths, the magnetotactic bacteria can follow the oxygen gradient and explore hypoxic areas. When following the oxygen gradient, the magnetotactic bacteria exhibit more run-and-tumble type movement patterns, thus enabling them to move into hypoxic areas. Once the magnetotactic bacteria detect a hypoxic area, they remain in or near that hypoxic area. When the velocity of the magnetotactic bacteria becomes zero (e.g., they die or weaken within the subject), the magnetotactic bacteria deposit one or more agents (therapeutic agents, imaging agents, diagnostic agents) in the hypoxic area. This enables targeted therapy, imaging, and / or diagnosis, reducing the required dosage of drugs, and consequently, minimizing the impact on the patient's health.

[0124] Magnetic and oxygen-tactic bacteria:

[0125] In this disclosure, magnetotactic bacteria refer to a group of bacteria that have flagella for self-propulsion and chains of magnetosomes for orientation, and that respond to a magnetic field (i.e., magnetotactic) by changing orientation and moving along the magnetic field lines of the Earth's magnetic field. The magnetotaxis of magnetotactic bacteria can be conferred by specific intracellular organelles called magnetosomes. These magnetosomes contain membrane-bound nanometer-sized magnetite mineral particles (crystals) arranged in chains by a dedicated cytoskeleton. The magnetite mineral particles may include iron oxides such as magnetite (Fe3O4) or iron sulfides such as grayite (Fe3S4) or pyrite (FeS2). Magnetotactic bacteria also respond to oxygen gradients (i.e., are oxygenotactic). Examples of magnetococci and oxygen-tactic bacteria include bacteria of the Magnetococcus marinus species (e.g., strains MO-1, MSR-1, MC-1, BM-1, or genetically modified versions of these species or strains, including those optimized in the laboratory). Magnetococci and oxygen-tactic bacteria may be naturally occurring, adapted through natural selection, or adapted through artificial intervention such as laboratory manipulation, in which the characteristics of the bacteria (e.g., sensitivity to magnetic fields) have been selected or optimized. Artificial interventions include, but are not limited to, genetic engineering, selection and growth of bacteria exhibiting specific characteristics, and adjustment of the environment and / or culture medium to optimize bacterial characteristics or acquire specific traits. Magnetic and oxygen-tactic bacteria share the following characteristics: they are self-propelled (in some cases, magnetic and oxygen-tactic bacteria are driven by an external source that generates a magnetic torque that displaces the bacteria), they are magnetically induced, and they are adapted to be induced, move, and / or react to external stimuli other than magnetic fields.

[0126] In one embodiment, the magnetic-oxytomic bacteria of the present disclosure are non-pathogenic and, for example, do not grow in a subject and do not cause bacterial infection. Magnetic and oxygen-tactic bacteria are obtained from non-pathogenic Magnetospirillum magneticum strains (e.g., AMB-1), non-pathogenic Magnetotactic coccus strains, non-pathogenic Magnetospirillum magnetotacticum strains, non-pathogenic Magnetospirillum gryphiswaldense strains (e.g., MSR-1), non-pathogenic Magnetospirillum bellicus strains, non-pathogenic facultative anaerobic magnetotactic spirillum strains, or non-pathogenic obligate anaerobic strains, such as Desulfovibrio magneticus strains (e.g., RS-1). In one embodiment, the magnetotactic bacterium is a magnetotactic coccus. It should be understood that non-pathogenic bacteria may possess certain inflammatory bacterial components (e.g., LPS), which may cause mild symptoms in the subject. However, if the bacteria do not proliferate and do not cause serious symptoms in the subject, they are considered non-pathogenic.

[0127] Magnetochotactic bacteria can move along gradients of decreasing oxygen concentration, following down to approximately 2% or less. Normal oxygen concentration in healthy tissue (sometimes referred to as "physoxia") varies by tissue, averaging approximately 5-6% (ranging from approximately 7.5% to 4% depending on the tissue) (McKeown MA, Br J Radiol, 2014, 87(1035)). Therefore, the magnetochotactic bacteria of this disclosure are capable of moving from areas of tissue or organ exhibiting normal oxygen concentration (physoxia), such as areas with oxygen concentrations of approximately 4%-7.5% or 5-6%, to areas with decreased oxygen concentration (hypoxic areas). Hypoxic areas include areas of tissue or organ with oxygen concentrations of approximately 2% or less, approximately 1.75% or less, or approximately 1.5% or less. In one embodiment, the magnetic-oxytomic bacteria of this disclosure have the ability to move along a gradient of decreasing oxygen concentration and follow it until they reach levels where the oxygen concentration is approximately 0.3% to approximately 0.7%, approximately 0.4% to approximately 0.6%, or approximately 0.5%. This property is useful, for example, for reaching hypoxic regions in organs, tissues, or tumors. For example, the median oxygen concentration in most tumors is usually less than 2%, and in some tumors, the median oxygen concentration is less than 1% (McKeown MA, Br J Radiol, 2014, 87(1035)). However, heterogeneity in individual tumors is significant, and therefore, in some areas of a tumor, the oxygen concentration may be less than 1%, for example, about 0.5% to 1%, or even lower. Such hypoxic regions in tumors are usually highly resistant to cancer treatments such as chemotherapy and / or radiotherapy.

[0128] In some cases, the survival time of magnetotactic bacteria within the subject's body may be limited to a short period. Magnetotactic bacteria may die within a short period after administration to the subject. Therefore, if the magnetotactic bacteria contain a therapeutic agent, the agent (e.g., therapeutic agent, diagnostic agent, imaging agent) may remain at the target site (site of interest) once the magnetotactic bacteria die.

[0129] Characteristics of magnetotactic and oxygenotactic bacteria:

[0130] It will be understood that magnetotactic bacteria can be north-directed (pointing towards the north in a magnetic field), south-directed (pointing towards the south in a magnetic field), or a mixture of both. The magnetic field sequences relating to this disclosure may be selected depending on whether the magnetotactic bacteria are north-directed, south-directed, or a mixture of both. Before injection into a subject, a sample or bolus of magnetotactic bacteria may be tested (for example, by exposing the magnetotactic bacteria to a magnetic field of known properties and monitoring the bacteria's response to the magnetic field; in some cases, a microscope may be used, and a video of the behavior of the magnetotactic bacteria may be generated using a camera and microscope). Such tests may determine whether the magnetotactic bacteria are south-directed, north-directed, or a mixture of both (the ratio of south-directed to north-directed bacteria may be determined).

[0131] In some cases, other characteristics of magnetotropic bacteria in a sample of magnetotropic bacteria (e.g., administered to a subject) may be identified (e.g., by using a camera in addition to a microscope to generate a video of the magnetotropic bacteria observed under a microscope). The velocity of magnetotropic bacteria can be calculated by dividing the distance traveled by the magnetotropic bacteria (e.g., average distance traveled) by time. The velocity or velocity distribution can be calculated for a given group or sample of magnetotropic bacteria. The response time of magnetotropic bacteria to changes in magnetic field direction can also be determined by observing the reaction time of the magnetotropic bacteria (i.e., the time it takes for the magnetotropic bacteria to change direction, change their direction of movement, and align themselves in the new magnetic field direction after the direction of the magnetotropic bacteria changes). By using the velocity distribution (or velocity) of magnetotactic bacteria, and / or the reaction time of magnetotactic bacteria to changes in the direction of the magnetic field, it is possible to identify the minimum magnetic field strength that causes displacement of the bacteria (or a magnetic field strength low enough for the bacteria to prioritize searching for hypoxic regions within the subject).

[0132] Furthermore, changes in the magnetic field direction can provide information about the polarity of magnetotactic bacteria in the sample (i.e., north-facing, south-facing, or a mixture of north-facing and south-facing bacteria).

[0133] The density of magnetotactic bacteria in a given sample solution can also be determined (for example, from a video obtained from a microscope, or by directly observing the magnetotactic bacteria under a microscope) by dividing the number of magnetotactic bacteria by a known surface area or volume.

[0134] The decay of the activity of magnetotactic bacteria in a given sample can also be measured. Furthermore, the change in the velocity of magnetotactic bacteria over time can also be measured. Observation of magnetotactic bacteria can be performed using a camera (attached to a microscope), a magnetic field source to apply a magnetic field to the sample containing the magnetotactic bacteria, and one or more sensors characterizing the environment of the magnetotactic bacteria (e.g., a pH monitor to measure pH, a thermometer to measure temperature, etc.). The velocity or velocity distribution of magnetotactic bacteria in the sample can be calculated at predetermined time intervals (e.g., every 5 minutes) (i.e., the distance traveled by the magnetotactic bacteria divided by time). The decay of magnetotactic bacteria activity can be calculated by plotting the decrease in the measured bacterial velocity as a function of time (velocity is calculated from the distance traveled and elapsed time in each measurement time interval). The decay of magnetotactic bacteria activity can also be calculated under different environmental conditions (e.g., the average body temperature of the subject, 37°C).

[0135] The characteristics measured for magnetotactic bacteria may be stored in association with an identifier or code (e.g., a barcode or QR code®) attached to the container of the magnetotactic bacteria sample. This allows scanning the identifier or code to obtain information indicating the characteristics of the magnetotactic bacteria contained in the sample. These characteristics can be used as guidance for the injection of magnetotactic bacteria into the subject and / or for the appropriate application of a magnetic field or magnetic field sequence.

[0136] Definition:

[0137] In this disclosure, “imaging” means a medical technique that enables the acquisition of information about a subject. In some embodiments, imaging makes it possible to observe the anatomical structure of a subject. Such techniques include, but are not limited to, magnetic resonance imaging (MRI), computed tomography (CT scan, CAT scan, etc.), positron emission tomography, single-photon emission computed tomography, electron paramagnetic resonance imaging, ultrasound, and X-ray. In other embodiments, imaging can be used to collect information about a subject, such as the presence of drugs inside or in specific locations within the subject. Imaging may be performed, for example, with a radiation detector having a predetermined aperture (for example, a small aperture) capable of detecting radiation in a specific area of ​​the subject (e.g., near an organ).

[0138] In this disclosure, “bolus” means a single dose of a solution administered to or given to a subject. A bolus has a given volume. In this application, a bolus contains a given concentration of magnetotactic bacteria in a solution, which may be conjugated to a therapeutic agent, a diagnostic agent, and / or an imaging agent.

[0139] In this disclosure, “diagnose” means (i) determining whether a particular condition, disease, or disorder is present; (ii) determining the risk of developing a particular condition, disease, or disorder; or (iii) determining the progression of a particular condition, disease, or disorder (worsening, no change, or improvement).

[0140] In this disclosure, “imaging agent” means an agent for providing more information about organs, cellular processes, tumors, and normal tissues. The imaging agent may be a contrast agent that enhances the contrast of tissues or fluids detectable by medical imaging. Examples of such agents include, but are not limited to, iodine, barium, iron oxide nanoparticle clusters, gadolinium and gadolinium derivatives, magnetic nanoparticles such as iron-platinum particles, manganese-based nanoparticles (e.g., manganese chelates), perflubron, fluorodeoxyglucose, protein-based imaging agents, and microbubble imaging agents. In some examples, the imaging agent may be present within a carrier (vehicle) (e.g., a vesicle) for encapsulating the imaging agent. The imaging agent may be bound to the surface of magnetotactic bacteria or taken up within such bacteria. In some embodiments, the imaging agent may be an agent that emits radiation detectable by a radiation detector (e.g., a radioisotope). In some embodiments, the imaging agent may be taken up by magnetotactic bacteria. For example, if the magnetotactic bacteria are magnetotactic bacteria, they can be incorporated by culturing them in a culture medium containing a radioactive isotope. In some embodiments, a protein donor may be incorporated into the magnetotactic bacteria. In some embodiments, a radiation-emitting imaging agent may be bound to the surface of the magnetotactic bacteria.

[0141] In this disclosure, “subject” means mammals and non-mammals. Mammals include, but are not limited to, humans, any member of the class Mammalia. Non-mammals include birds, reptiles, etc. The term “subject” does not impose any limitations on sex or age.

[0142] In this disclosure, “target site” is also referred to herein as “target zone,” and means a site within a subject to which magnetotactic bacteria are to be induced. The target site includes a hypoxic region to which magnetotactic bacteria follow the oxygen gradient by oxygenatosis. Magnetotactic bacteria may aggregate in the hypoxic region at the target site. The target site may be a tumor or a part thereof, or an organ of the subject.

[0143] In this disclosure, “to treat” or “treatment” means one or more of the following: (i) to prevent (temporarily or permanently) some or all of a particular condition, disease, or disorder; (ii) to suppress or reduce (temporarily or permanently) one or all of a particular condition, disease, or disorder; or (iii) to alleviate (temporarily or permanently) some or all of a particular condition, disease, or disorder.

[0144] In this disclosure, “transpotherapy” refers to a therapeutic mode that enables non-systemic delivery of a therapeutic agent based on directional control or guidance of the delivery of the therapeutic agent (for example, by utilizing a more direct delivery route from the injection (or introduction) site to the physiological area to be treated), and such delivery is achieved by using a transporter such as magnetic or oxygen-tactic bacteria at a specific physiological site in a specific volume. Unlike systemic delivery used in chemotherapy, for example, the main advantage of transpotherapy is that it enhances the therapeutic effect while minimizing systemic toxicity or systemic exposure, and generally, it is possible to increase the amount delivered to the treatment site with a significantly smaller injection volume compared to chemotherapy. With transpotherapy, it is also possible to deliver the therapeutic agent to the tumor mass beyond the diffusion limit of standard chemotherapy agents. In this specification, transpotherapy is used as a comprehensive term and is not limited to, but includes, for example, the non-systemic delivery of diagnostic agents (transpo-diagnostic) and imaging agents (transpo-imaging). For example, transpotherapy used for the delivery of radiosensitizers in radiotherapy can be more specifically called transpo-radiotherapy. Similarly, transporter therapy used in immunotherapy can be called transporter immunotherapy, and transporter chemotherapy refers to the implementation of non-systemic chemotherapy using such transporters. It is also possible to combine multiple treatment modalities using transporter therapy. One such example is the combination of chemotherapy and immunotherapy, which can be called transporter chemoimmunotherapy. Transporters for transporter therapy that carry both therapeutic and diagnostic agents can be called transporter therapy-diagnostic complexes. Many combinations are possible. For example, transporter chemotherapy-imaging can be achieved by simultaneously or separately injecting a transporter chemotherapy complex and a transporter imaging (or transporter contrast) complex. It can also be achieved by using a transporter chemotherapy complex and an imaging complex in a single injection.A similar nomenclature can be applied to other types of transporation therapy complexes and treatment methods, with "transporation therapy" being used as a general term for treatment methods.

[0145] In this specification, “velocity distribution” is also referred to as “velocity standard deviation,” and refers to the degree of variation or deviation in velocity exhibited under magnetotaxis by a group of magnetotactic bacteria contained in a particular sample or solution (e.g., for administration to a subject). The velocity distribution relates to the velocity before injection. The velocity distribution can be estimated or obtained using known imaging techniques (e.g., microscopy). It can also be estimated or obtained by generating cultures of magnetotactic bacteria and, in some cases, by a selection process of combining such cultures according to velocity (furthermore, it is also possible to prepare a sample or solution by selecting cultures that exhibit a desired velocity or velocity range).

[0146] Magnetic and oxygen-tactic bacteria:

[0147] Now, refer to Figure 1. This figure is a simplified diagram illustrating magnetic-oxygen-tactic bacteria 100 and magnetic-oxygen-tactic bacteria 200.

[0148] Magnetic and oxygen-tactic bacteria 100 and 200 have a magnetosome chain 104 that is sensitive to magnetic fields (e.g., applied magnetic fields or geomagnetic fields).

[0149] In some embodiments, the therapeutic agent 202 may be encapsulated within the carrier 201. The carrier 201 may have a hydrophilic portion and a hydrophobic portion (e.g., a phospholipid bilayer). Depending on whether the therapeutic agent is hydrophobic or hydrophilic, the therapeutic agent may reside in either the hydrophilic or hydrophobic portion of the carrier 201. In some embodiments, the therapeutic agent 202 is encapsulated within a carrier made of a biodegradable polymer. It should be understood that the therapeutic agent 202 can be encapsulated within any carrier that effectively encapsulates the therapeutic agent 202 being transported.

[0150] In some embodiments, the therapeutic agent 202 can be conjugated to magnetotactic bacteria 200 using, for example, a ligand 205 (such as an antibody). The ligand 205 can be attached to the therapeutic agent 202, in which case the ligand may be adapted to bind to the outer surface of the magnetotactic bacteria 200. The ligand 205-therapeutic agent 202 conjugate can be placed in the same solution as the magnetotactic bacteria 200, thereby allowing the ligand-therapeutic agent conjugate to bind to the magnetotactic bacteria 200.

[0151] In some embodiments, the therapeutic agent 202 is present within a vesicle 201, which can be conjugated to a ligand (e.g., an antibody) adapted to bind to a magnetotactic bacterium 200. The vesicle 201 can then be conjugated to the membrane of the magnetotactic bacterium via the ligand 205.

[0152] In some embodiments, the magnetic and oxygen-tactic bacteria 200 may be forcibly activated with lipopolysaccharides so that they adhere to a carrier 201 containing the therapeutic agent 202. For example, the magnetic and oxygen-tactic bacteria 200 may be mixed in a culture medium with a low nutrient concentration to adjust to a new concentration, and then the carrier carrying the therapeutic agent may be injected into a new culture medium containing the given concentration.

[0153] It should be understood that it is also possible to immobilize the therapeutic agent 202 on the magnetotactic bacteria 200 using other mechanisms without departing from the teachings of this disclosure. For example, see Taherkhani et al., “Covalent Binding of Nanoliposomes to the Surface of Magnetotactic Bacteria for the Synthesis of Self-Propelled Therapeutic Agents” (ACS Nano, May 27, 2014, 8(5):5049~60). This document is incorporated herein by reference.

[0154] Depending on the application, other binding means may be provided between the magnetic / oxytactic bacteria and the therapeutic agent 202.

[0155] In some embodiments, the magnetotactic bacteria do not include a carrier 201, and the therapeutic agent 202 may be directly bound to the magnetotactic bacteria 200 or incorporated into the magnetotactic bacteria 200 (in which case the magnetotactic bacteria function as a carrier for transporting the therapeutic agent).

[0156] In some examples, the therapeutic agent 202 and the imaging agent 101 are the same agent (for example, in the case of superparamagnetic iron oxide nanoparticles).

[0157] In some embodiments, both the imaging agent 101 and the therapeutic agent 202 are immobilized on magnetotactic bacteria (e.g., magnetotactic bacteria 300 in Figure 2).

[0158] In some embodiments, the imaging agent 101 can be conjugated to magnetotactic bacteria 100 using, for example, a ligand 105 (such as an antibody). The ligand 105 can be attached to the imaging agent 101, in which case the ligand 105 may be adapted to bind to the outer surface of the magnetotactic bacteria 100. The ligand-imaging agent conjugate can be placed in the same solution as the magnetotactic bacteria 100, thereby allowing the ligand-imaging agent conjugate to bind to the magnetotactic bacteria 100.

[0159] It should be understood that it is also possible to immobilize the imaging agent 101 on the magnetic-oxytotactic bacteria 100 using other mechanisms, without departing from the teachings of this disclosure.

[0160] In some embodiments, the proportion, quantity, and / or concentration (or relative concentration) of magnetotactic bacteria, as well as the magnetotactic bacteria themselves, are known. By observing the amount of magnetotactic bacteria that reach and / or remain at the target site (e.g., at least one hypoxic region in a tumor) and the amount (or proportion) that diffuses from the target site, it is possible to estimate, based on the known proportion, quantity, or concentration of magnetotactic bacteria containing the therapeutic agent, that a similar proportion of the magnetotactic bacteria containing the therapeutic agent remains at the target site, and that the amount that diffuses from the site can also be estimated. This information can be a useful indicator, for example, for evaluating the toxicity of the therapeutic agent (based on the amount of therapeutic agent that diffuses into the body without being transported to the target site). For example, this information can be used as an indicator of the required dosage of the therapeutic agent, based on the estimation that a given proportion of magnetotactic bacteria carrying the therapeutic agent does not reach or remain at the target site.

[0161] It should be understood that solutions containing magnetic and oxygen-tactic bacteria can be prepared using known techniques for the purpose of preserving the bacteria and administering them to subjects.

[0162] In some embodiments, magnetotactic bacteria are frozen before being bound to an imaging agent, a therapeutic agent, or both. The thawed magnetotactic bacteria can be grown. Alternatively, they can be added to the solutions described herein to be bound to a contrast agent, a therapeutic agent, or both.

[0163] An exemplary system for targeting aggregation regions using magnetic and oxygen-tactic bacteria:

[0164] Next, refer to Figure 8. This figure shows a system 800 that implements a method for targeting aggregated regions within a subject using magnetotactic bacteria. The system adjusts the strength of the magnetic field according to the progress of the magnetotactic bacteria within the subject.

[0165] System 800 includes a processor 801 and memory 802.

[0166] System 800 is capable of communicating with, and may include, one or more pairs (e.g., three pairs) of magnetic sources 804 (e.g., magnetic coils) aligned with any one of the three axes (x, y, and z). System 800 is capable of communicating with, or may include, an imaging device 805.

[0167] System 800 includes a display 803. System 800 also includes a user input interface 806.

[0168] The processor 801 may be a general-purpose programmable processor. In this embodiment, the processor 801 is shown as a single unit, but the processor 801 may be multi-core or distributed (e.g., a multiprocessor).

[0169] Computer-readable memory 802 stores program instructions and data used by processor 801. Memory 802 may be non-temporary memory. Although computer-readable memory 802 is shown as a single unit for simplification in this embodiment, it may comprise multiple memory modules and / or caches. Specifically, the memory may comprise multiple memory layers such as a hard drive and an external drive (e.g., SD card storage), and faster, smaller random access memory (RAM) modules. The RAM modules can store data and / or program code that processor 801 is currently processing, has recently processed, or will soon process, as well as cache data and / or program code from the hard drive. The hard drive stores program code and is accessed by processor 801 for retrieval and execution of said code. It is also accessed by processor 801 to store image information about the subject, values ​​related to the velocity of magnetotactic bacteria, the volume of the injection bolus, program code for operating the magnetic source 804, etc. Memory 802 may have a recycling architecture for storing, for example, image information related to the subject. In this architecture, when memory 802 is full or nearing full, old data files are deleted, or old data files that have been stored in memory 802 for a predetermined period of time are deleted.

[0170] The processor 801 and memory 802 may be connected via a BUS connection.

[0171] The user input interface 806 allows the user to interact with the system 800 by providing input to it, and can be used, for example, to set the magnetic field strength generated by the magnetic source 804 or to identify a cluster region or 3D convergence point on the display 803. The user input interface 806 is, for example, a mouse, keyboard, and / or controller, and can be used to receive user input from the user.

[0172] It will also be understood that other user input interfaces, such as touchscreens, joysticks, microphones, and one or more proximity sensors for detecting user movement, can be used in accordance with the teachings of this application.

[0173] In some embodiments, system 800 does not have a user input interface 806. In this case, system 800 receives user input from a remote computer device (e.g., a remote computer, smartphone, tablet, laptop, etc.) via an input / output interface such as a transceiver (not shown).

[0174] The display 803 provides a graphical user interface for controlling the system 800 and can present the user with image information generated by the imaging device 805, parameters of the magnetic source 804, and the like. The display 803 may also have a touchscreen function and can therefore also serve as the user input interface 806.

[0175] The magnetic source 804 may include one or more pairs of magnetic coils. Each pair of magnetic coils may be arranged along one axis. For example, the magnetic source 804 may include three pairs of magnetic coils, each pair of which may be arranged along one of the three axes (x, y, z). The magnetic source 804 may generate 3D convergence points (and aggregate regions) as described herein or in U.S. Patent No. 9,905,347.

[0176] The imaging device 805 generates image information of the subject, which can be used, for example, to define aggregated regions and 3D convergence points, to provide visual information about the subject's body (e.g., a tumor), to provide visual information about hypoxic regions of the tumor, and to monitor the movement of magnetotactic bacteria injected into the subject. The imaging device 805 enables the performance of the imaging described herein. For example, the imaging device 805 may be a PET (positron emission tomography) device, an MRI (magnetic resonance imaging) device, a CT (computed tomography) device, etc.

[0177] Exemplary methods of targeting using magnetotactic bacteria:

[0178] Next, refer to Figure 7. This figure shows an exemplary method 700 for targeting aggregated regions within a subject using magnetotactic bacteria by changing the magnetic field strength.

[0179] For the sake of explanation, referring to an exemplary system 800, method 700 is carried out by having a processor 801 execute program code stored in memory 802, and the processor 801 executes instructions of the program code stored in memory 802. The processor 801 can control the magnetic field strength of the magnetic field generated by the magnetic source 804 by controlling the power / current supplied to the magnetic source 804.

[0180] The processor 801 can also control the imaging device 805 to acquire image information. In some embodiments (for example, when the imaging device 805 is not part of system 800), the image information is sent to system 800 and then to the processor 801 via an input / output (I / O) interface (e.g., a transceiver).

[0181] In step 710, image information of the subject is acquired. Image information can be acquired by imaging as described herein. Image information can provide information about target sites, tissues, or organs in the subject (e.g., tumors, anatomical structures or parts thereof of the subject, hypoxic areas in tumors, etc.).

[0182] In some embodiments, image information can be enhanced by using magnetotactic bacteria in combination with an imaging agent. Since the imaging agent is detectable by imaging, it can improve the information collected by imaging.

[0183] In step 720, one or more boluses of magnetotactic bacteria are injected into one or more sites. In some embodiments, the magnetotactic bacteria are injected into the peripheral region of the tumor. In some embodiments, the injection may be systemic. The bolus contains magnetotactic bacteria at a given concentration and has a given volume. The magnetotactic bacteria may be conjugated to one or more of the following: therapeutic agents (e.g., anticancer agents), diagnostic agents, and / or imaging agents.

[0184] When applying the first mode, in step 730, a magnetic field is applied with a magnetic field strength sufficient to act on the magnetotactic bacteria with directional torque. This makes magnetotaxis and run-and-reverse motion dominant in the movement of the bacteria (the increase in run states and the reduction in relaxation time allow the direction of bacterial movement to realign with the direction of the magnetic field, resulting in less change in the direction of bacterial movement compared to run-and-tumble motion). The first mode is a mode that displaces the bacteria from their current location (e.g., injection site) to a new location (e.g., target site). In some embodiments, the first magnetic field strength is at least 15 gauss. However, as will be readily apparent to those skilled in the art, the value of the first magnetic field strength may vary depending on the characteristics of the magnetotactic bacteria, such as their sensitivity to magnetic fields. The magnetic field strength can be calculated based on, and may depend on, the volume of the injected bolus, the distance traveled by the magnetotactic bacteria (for example, this can be calculated from the known velocity of the bacteria and the time elapsed after injection), the time elapsed after injection (because the velocity and / or responsiveness of the bacteria decreases with residence time in the body), the surface area of ​​the bolus, and / or the concentration of magnetotactic bacteria in the bolus.

[0185] Magnetic fields can induce magnetotactic bacteria in one to three dimensions, for example, by generating 3D convergence points as described herein.

[0186] Optionally, in step 740, information regarding the progress of magnetotactic bacteria moving within the subject may be generated (for example, by monitoring the movement of magnetotactic bacteria or estimating the location of magnetotactic bacteria). For example, monitoring of magnetotactic bacteria is performed by combining the bacteria with an imaging agent that can be perceived by imaging. In some embodiments, monitoring of magnetotactic bacteria is performed by calculating the elapsed time since the bacteria were injected into the subject. The distance can be estimated based on the distribution of bacterial velocity before injection, the decay of bacterial velocity after injection, and the elapsed time after injection. In some embodiments, the distance value can also be calculated from the volume and surface area of ​​the injected bolus.

[0187] In the second mode, when it is observed or estimated that magnetotactic bacteria are approaching a target site, in step 750, a magnetic field with a second magnetic field strength lower than the first magnetic field strength is applied. The target site has an oxygen gradient resulting from a hypoxic region located at the target site, or is located near such a gradient. By reducing the magnetic field strength, the movement pattern of the magnetotactic bacteria changes. The magnetotactic bacteria exhibit more run-and-tumble motion compared to when a higher first magnetic field strength is applied. This change in movement pattern to run-and-tumble motion allows the magnetotactic bacteria to explore hypoxic regions of the tumor by oxygenataxis. In some embodiments, the second magnetic field strength is less than 15 gauss and greater than 5 gauss. However, as will be readily apparent to those skilled in the art, the value of the second magnetic field strength may vary depending on the characteristics of the magnetotactic bacteria, such as their sensitivity to magnetic fields. The magnetic field strength can be calculated based on, and may depend on, the volume, viscosity, shape, and anisotropic orientation of the injected bolus, the distance traveled by magnetotactic bacteria (for example, this can be calculated from the known velocity of the bacteria and the time elapsed after injection), the time elapsed after injection (because the velocity and responsiveness of the bacteria decrease with residence time in the body), and the concentration of magnetotactic bacteria in the bolus.

[0188] In some embodiments, in the third mode, when the magnetotactic bacteria reach the oxygen gradient of a hypoxic region at the target site, in step 760, the magnetic field strength is reduced to less than 5 gauss, and further reduced to 0 gauss. This reduction in magnetic field strength is performed to keep the magnetotactic bacteria within the hypoxic region of the target site, and the bacteria change direction further than in step 750 as a result of run-and-tumble motion, which is inconsistent with the direction of the magnetic field. The magnetotactic bacteria can be bound to one or more therapeutic agents, diagnostic agents, and / or imaging agents, so that when the post-injection velocity of the bacteria in the hypoxic region decreases to 0 (for example, by the bacteria dying or weakening) (for example, a few minutes after introduction into the subject), the therapeutic agent, diagnostic agent, and / or imaging agent are deposited in the hypoxic region of the subject. This allows the compound to be administered to the subject in a targeted manner. Furthermore, this method makes it possible to specifically target hypoxic areas of tumors, which were difficult to target with conventional compound administration due to the tumor's vascular and cellular structure.

[0189] This method utilizes the changes in the responsiveness and integrity of magnetotactic bacteria that occur after administration to a subject. Since the specific low-temperature environmental conditions necessary for the survival of magnetotactic bacteria are not provided within the human body, their lifespan is shortened upon introduction into the human body. Furthermore, the longer the residence time of magnetotactic bacteria within the human body, the slower their movement becomes, and they eventually die. Therefore, it is desirable to utilize the time (after injection) when magnetotactic bacteria are moving rapidly to improve the accuracy of targeting. Accordingly, by applying a magnetic field strength increased to its maximum value after injection (to avoid the problem of repolarization), the magnetotactic response of the magnetotactic bacteria is promoted, allowing the bacteria to exhibit optimal activity and travel long distances to reach the oxygen gradient adjacent to the hypoxic region of the target area. As the magnetotactic bacteria approach the oxygen gradient formed by the tumor, their movement speed decreases due to the hostile environment within the subject's body. However, the distance that magnetotactic bacteria travel when exploring hypoxic areas of tumors may be short, and such slowing of movement is acceptable.

[0190] Exemplary methods for controlling the diffusion of magnetotactic bacteria during targeting:

[0191] For example, when performing the targeting described above using method 800, even if the bacterial concentration in each hypoxic region at the target site decreases, it may be desirable to target more hypoxic regions with a single injection, or it may be desirable to concentrate targeting on a few hypoxic regions with a higher bacterial concentration. Therefore, it may be beneficial to adjust the target volume at the target site according to the desired result. Targeting that disperses bacteria more will result in a larger target volume, while targeting that suppresses bacterial dispersion (at a higher concentration) will result in a smaller target volume at the target site.

[0192] The increase or decrease in the target volume of the target site reached by the injected bacteria can be achieved by preparing or selecting a sample based on the dispersion rate of the bacterial sample to be administered. Referring to Figure 9A, the figure shows an example of a large target volume 900A that includes multiple hypoxic regions 910 at the target site. Figure 9B shows a small target volume 900B that includes fewer hypoxic regions at the target site. It should be understood that the exemplary target volumes 900A and 900B, and the hypoxic regions, are simplified for illustrative purposes and can take various shapes depending on the anatomical structure of the subject and the target organ, tissue, or tumor. Target volumes 900A and 900B are used to illustrate the difference in size of the target volume. For a target volume 900A, a bacterial sample with a wide velocity distribution can be selected. For a target volume 900B, a bacterial sample with a narrow velocity distribution can be selected.

[0193] When the velocity distribution is wide, the target volume or diffusion is large. When the velocity distribution is narrow, the target volume or diffusion is small.

[0194] By evaluating this velocity distribution (e.g., the standard deviation of bacterial velocities in the sample), users can select a bacterial sample based on a desired target volume at the target site.

[0195] Furthermore, the pre-injection velocity of bacteria in the sample may be compared to a velocity threshold for passing through one or more hypoxic regions near the injection site. Bacteria exhibiting velocities above the threshold are likely to move through these hypoxic regions. On the other hand, bacteria exhibiting velocities below the threshold are likely to remain aggregated within these hypoxic regions. Therefore, information regarding the post-injection bacterial migration pattern can be obtained not only from the bacterial velocity distribution, but also from the pre-injection velocity of the bacteria itself.

[0196] The velocity distribution of the sample can be evaluated (and determined, for example, if it is not provided by the bacterial sample manufacturer) before injection into the patient. If the velocity distribution of the sample does not match what is needed to achieve the desired target volume at the target site, the sample may be discarded or replaced with another sample showing a more desirable velocity distribution.

[0197] Furthermore, samples may be prepared according to the desired diffusion or target volume at the target site. For example, multiple bacterial cultures associated with a given velocity or velocity range (e.g., "low velocity," "medium velocity," "high velocity") may be cultured. If a small velocity distribution is desired, a sample may be prepared from a single culture exhibiting a relatively uniform velocity. On the other hand, if a large velocity distribution is desired, a sample may be prepared from multiple cultures exhibiting different velocity levels, thereby obtaining a sample containing both slow and fast bacteria (increasing the overall velocity distribution of the prepared sample). It should also be understood that it is possible to select cultures with multiple velocity ranges from the same culture.

[0198] In other embodiments, the user may select samples focusing on the decay of post-injection velocity. For example, some bacteria may have higher tolerance to the environment within the test body than others, and therefore take longer to decay. On the other hand, some bacteria may have lower tolerance and decay rapidly after injection. If the predicted decay profile of post-injection velocity is known, the user may select a sample with a large range of change in post-injection velocity (even if the velocity distribution is relatively narrow) when the target volume is large, and a sample with a small range of change in post-injection velocity when the target volume is small. When a sample with a large range of change in post-injection velocity is used, the range of distance the bacteria travel when the magnetic field strength is adjusted to promote bacterial magnetotaxis and run-and-reverse motion increases because the velocity distribution of the bacteria administered to the test body widens over time due to the large distribution of change in post-injection velocity.

[0199] Exemplary methods for controlling the distribution of magnetotactic bacteria

[0200] This disclosure also describes a method for controlling the size of the target distribution zone for magnetotactic bacteria in tumors within a subject.

[0201] A bolus of magnetotactic bacteria contains magnetotactic bacteria with a variety of velocities, from high to low. By utilizing the strength of the magnetic field, it becomes possible to control the size of the target distribution zone of bacteria based on the velocity distribution of the magnetotactic bacteria.

[0202] Under high-intensity magnetic fields, faster-moving magnetotropic bacteria are induced by the magnetic field and move along its direction. This displacement is related to the bacteria's velocity. Faster magnetotropic bacteria tend to travel further than slower ones. Increasing the magnetic field strength promotes the run-and-reverse movement of magnetotropic bacteria, causing faster bacteria to travel further and slower bacteria to remain near the injection site. As a result, magnetotropic bacteria are dispersed over a wide area within the tumor. This is because faster magnetotropic bacteria are more likely to travel the furthest from the injection site. The dispersion of magnetotropic bacteria with varying velocities allows them to spread across the target distribution area. Therefore, such magnetotropic bacteria target larger tumor areas (e.g., more hypoxic zones within the tumor). Furthermore, magnetotactic bacteria gradually weaken upon exposure to the surrounding environment inside the subject's body, eventually ceasing to move over time. As a result, even the fastest magnetotactic bacteria, upon exposure to the surrounding environment inside the subject's body, will have their velocity approach zero over time, their movement inhibited, and eventually come to a complete halt.

[0203] On the other hand, reducing the magnetic field strength inhibits even the fastest magnetotactic bacteria from moving far (increasing run-and-tumble movement), causing them to remain near the injection site. As a result, the spread of magnetotactic bacteria is reduced, and the target distribution zone shrinks. However, as the target distribution zone shrinks, the concentration of magnetotactic bacteria in that zone increases. Therefore, when a high concentration of magnetotactic bacteria is required in a specific area of ​​the tumor (e.g., a hypoxic area), it is appropriate to set the magnetic field strength low.

[0204] In some embodiments, the magnetic field strength can be kept at 0 Gauss, in which case the injected magnetotactic bacteria, both fast and slow, concentrate around the injection site.

[0205] In another embodiment, the magnetic field strength can be set to 15 gauss, in which case fast magnetotactic bacteria are displaced more significantly from the injection site, while slower magnetotactic bacteria remain near the injection site. Higher magnetic field strengths result in a wider distribution of magnetotactic bacteria within the tumor, and a larger target distribution zone (because the fastest magnetotactic bacteria are more likely to travel the furthest from the injection site, and gradually approach immobility when exposed to the surrounding environment within the subject's body).

[0206] As those skilled in the art will understand, other magnetic field strength values ​​(e.g., any value between 0 and 15 Gauss, or other magnetic field strength ranges) may also be used, and the selected magnetic field strength may depend on factors such as the characteristics of magnetotactic bacteria (e.g., characteristics resulting from the selection of a particular strain of magnetotactic bacteria).

[0207] Exemplary study:

[0208] The following exemplary studies are intended to facilitate the understanding of this disclosure by those skilled in the art. These are illustrative and representative examples and are added solely for illustrative purposes and not to limit the scope of this disclosure. It is also possible to further explain and illustrate this disclosure using other exemplary studies without departing from the teachings of this disclosure, as will be understood by those skilled in the art.

[0209] Tumor regions with hypoxic conditions, known as hypoxic zones (HZs), exhibit resistance to various treatments such as chemotherapy and radiotherapy, and acquire a more malignant and invasive phenotype, which negatively impacts the patient's prognosis. For example, low oxygen levels in hypoxic zones reduce the effectiveness of radiotherapy, and chemotherapy agents cannot adequately reach deep-seated hypoxic zones, primarily due to diffusion limitations. Therefore, targeting hypoxic zones using a special propulsion vector capable of transporting therapeutic agents or other payloads to them can significantly improve therapeutic efficacy. In this specification, this propulsion vector is referred to as a therapeutic agent transporter (TT), and the propulsion vector with a therapeutic agent or other payload attached is referred to as a therapeutic agent transport complex (TC).

[0210] The specific magnetotropic bacteria described herein can be suitably used as therapeutic agent transporters for targeting hypoxic zones and delivering payloads. These magnetotropic bacteria are typically microaerophilic and have the property of seeking out oxygen levels that are similar to or close to the oxygen concentration in the hypoxic zone of a tumor. Tumor hypoxia generally occurs with an oxygen concentration of less than 1%, and for limited periods, it is often less than 0.1% (less than 7.5-0.75 mmHg) (Br J Radiol, March 2014, 87(1035):20130676). The Target Oxygen Level (TOL) as defined herein means the oxygen concentration targeted by the therapeutic agent transporter or therapeutic agent transport complex in the Aerotactic Targeting Mode (ATM). ATM refers to a state in which the directional displacement of the therapeutic agent transport complex is influenced solely by the oxygen gradient and is not affected by other factors such as directional magnetic fields that could influence its directional swimming behavior. TOL is appropriate as the oxygen concentration of the hypoxic zone, or sufficiently close to it, and as a result, it is possible to obtain appropriate targeted therapeutic effects that cannot be achieved with other conventional treatments, such as radiotherapy and chemotherapy, which are the two main examples.

[0211] Magnetic-oxygen-tactic therapeutic agent transporters or therapeutic agent transport complexes are most commonly guided to regions containing hypoxia zones using various magnetic field configurations (MFCs). A single magnetic field configuration, or a series of magnetic field configurations referred to as a magnetic field configuration sequence (MFCS), constitutes therapeutic agent transport targeting sequence events (TTSEs) (or simply referred to as events) that form a specific directional displacement pathway (DP) for the therapeutic agent transport complex, using one of three magnetotactic targeting modes (MTMs) (direct magnetotactic targeting (DMT), spatially restricted magnetotactic targeting (SMT), and aggregated region targeting (AZT)) or their derivatives. By combining all the necessary therapeutic agent transport targeting sequence events into a single sequence, a targeting sequence (TTS) is formed. Within the magnetic field configuration, the therapeutic agent transport complex is acted upon by a directional vector (DV), which guides the therapeutic agent transport complex to follow a specific displacement pathway. Each directional vector consists of two components: the magnitude and direction of the magnetic field configuration applied to the therapeutic agent transport complex at a given time. The magnitude of the magnetic field configuration refers to the magnetic field strength. Magnetic tactic oxygen targeting (MATM), which utilizes the magnetic field strength in the directional vector, is used in relation to the hypoxic zone and its impact on the operational behavior associated with the desired directional displacement of the therapeutic agent transport complex. In this method, the magnitude of the magnetic field configuration for targeting the hypoxic zone is appropriately adjusted, taking into account specific key parameters listed in the following sections.

[0212] Main parameters:

[0213] Magnetic field intensity range (FIR)

[0214] These magnetotactic and oxygen-tactic bacterial therapeutic agent transporters are suitable for targeting hypoxic zones and typically contain motile magnetotactic and oxygen-tactic bacteria. Here, magnetotactic and oxygen-tactic means that the bacteria can move and operate using only the magnetotactic mode, only the oxygen-tactic mode, or both modes simultaneously. The selection of the appropriate displacement or targeting mode for the magnetotactic and oxygen-tactic bacterial therapeutic agent transporter is performed by exposing the therapeutic agent transporter or transport complex to a predetermined range of directional magnetic flux densities. Here, the magnetic field intensity ranges are defined in order of increasing intensity as the aerotactic magnetic field intensity range, the magneto-aerotactic magnetic field intensity range, and the magnetotactic magnetic field intensity range, respectively, corresponding to the oxygen-tactic, magneto-aerotactic, and magnetotactic displacement or targeting modes. These ranges can also vary depending on the therapeutic agent transport complex, specific physiological environment, and other factors, but the typical magnetotaxis magnetic field strength range is between approximately 5 gauss and 15 gauss, with the lower limit defined by the oxygenotaxis magnetic field strength range (lower limit is 0 gauss) and the upper limit defined by the magnetotaxis magnetic field strength range (the upper limit of the magnetotaxis magnetic field strength range is limited because higher magnetic field strengths can cause repolarization (RPL) of the therapeutic agent transport complex, potentially reversing the direction of displacement of the transporter in the opposite direction to the intended direction).

[0215] Magnetochotic bacteria, which function as therapeutic agent transporters, exhibit oxygenataxis through a combination of run-and-tumble and run-and-reverse movements. Within the cells of magnetochotic bacteria, magnetosome chains containing magnetic particles are 10 -15 A m 2The magnetic moment is on the order of , which induces magnetic torque even in relatively weak directional magnetic fields. This magnetic torque acts to align the direction of motion of magnetotactic bacterial cells along the direction of the magnetic field. However, tumble motion disrupts this alignment. Therefore, run-and-tumble motion is characterized by the disruption of alignment caused by tumble and the subsequent relaxation process in the run state to align with the directional magnetic field. If the run state is shorter than the relaxation time, there is not enough time for the magnetic field to restore alignment with the magnetotactic bacteria before the next tumble occurs, and as a result, the direction of bacterial motion deviates from the direction of the magnetic field. The viscosity of the medium and the geometric properties of the therapeutic agent transport complex are only two of the factors that can affect the relaxation time. For example, since the rotational friction coefficient is proportional to the viscosity (η) of the medium, the induced torque required to rotate the cells is large, and as a result, the distribution of tumble angles is expected to be biased towards smaller angles. By increasing the strength of the directional magnetic field, the magnetic torque acting on the magnetosome chain increases, resulting in a shorter relaxation time and a smaller range of variation from the direction of the magnetic field.

[0216] Within the magnetic-oxygen-taxis magnetic field strength range, the directional displacement of the therapeutic agent transport complex is still affected by the directional magnetic field, but especially at low magnetic field strengths, the relaxation time increases, resulting in a greater range of variation in the direction of movement relative to the magnetic field. This increased range of variation contributes to keeping the therapeutic agent transport complex within the hypoxic zone when it passes through it at least temporarily within the magnetic-oxygen-taxis magnetic field strength range. Furthermore, when the magnetic field strength decreases to the oxygen-taxis magnetic field strength range (i.e., from 0 to the minimum magnetic field strength within the magnetic-oxygen-taxis magnetic field strength range), it contributes to keeping the complex within the hypoxic zone for a period longer than the total motion time (MT). However, when the directional magnetic field increases to the magnetic-taxis magnetic field strength range, the effect of the tumble state decreases significantly, and in the run state, the therapeutic agent transport complex moves along the magnetic axis in the direction of the magnetic field without showing substantial stagnation due to tumble, even within the hypoxic zone. If, following a running state, a reverse state occurs in which the therapeutic agent transport complex moves backward along the magnetic field axis (or, in the case of polar magnetotropic bacterial cells, a state in which they retreat while facing the direction of the magnetic field), the propulsive force or flagella of polar magnetotropic bacteria are located on only one side of the bacterium (unlike axial magnetotropic bacteria, they are not located on both sides). As a result, the forward movement speed is faster (more efficient) than the backward movement speed, and in north-directed (NS) magnetotropic bacteria, the net movement results in a displacement of the magnetic field towards the North Pole. (Note that magnetotropic bacteria can also repolarize to south-directed (SS) as needed, and vice versa.)

[0217] When therapeutic agent transport complexes are exposed to a directional magnetic field with a magnetic field configuration within the magnetotaxis field strength range, the magnetic directional torque acting on the magnetosome chains within magnetotaxis-oxygenotaxis bacterial cells increases, thereby restricting exploratory motion by oxygenotaxis run-and-tumble movement. Therefore, when exposed to a directional magnetic field within the magnetotaxis field strength range, run-and-reverse movement is likely to become dominant. Magnetotaxis-oxygenotaxis bacteria typically perform long unidirectional movements interrupted by reversal movements occurring at a specific average aperiodic frequency, resulting in the displacement of the bacteria forward.

[0218] When a therapeutic agent transport complex operates within the oxygen-tactic magnetic field strength range, the complex located in the target oxygen concentration region performs run-and-tumble and run-and-reverse movements to maintain its position at that oxygen concentration. As a result, the therapeutic agent transport complex forms a band-like distribution within or near the target hypoxic zone. The volume occupied by the therapeutic agent transport complex located within or near the hypoxic zone corresponds to the OATZ (Oxic-Anoxic Transition Zone). The period during which the therapeutic agent transport complex remains within the target oxygen concentration region (aerobic-anaerobic transition zone), such as the hypoxic zone, is referred to herein as the TOLRD (Target Oxygen Concentration Retention Period), which is defined as the mean or median period during which the therapeutic agent transport complex maintains its position within the target oxygen concentration region, such as the tumor hypoxic zone. Within the oxytaxis field strength range, a target oxygen concentration retention period of 1.0 means that the therapeutic agent transport complex remains in the target oxygen concentration region after reaching it until the post-injection velocity becomes 0 (the post-injection velocity is defined in the next section). Within the magnetotaxis field strength range, a target oxygen concentration retention period of 0 means that the therapeutic agent transport complex passes through the target oxygen concentration region without exhibiting any clear retention (due to the absence of clear tumbling and the occurrence of only short reverse motion with a long run phase). Within the magnetotaxis field strength range, the target oxygen concentration retention period changes from a value slightly above 0 near the lower limit of the magnetotaxis field strength range (MIN-MAFIR) to a value slightly below 1.0 near the upper limit (MAX-MAFIR).

[0219] Velocity after injection (VPI)

[0220] Post-injection velocity (VPI) is typically characterized by the rate reduction behavior exhibited by therapeutic agent transport complexes, including magnetic and oxygen-tactic bacteria, when exposed to physiological environmental factors that can reduce their motility. One such factor is the physiological temperature of 37°C. The post-injection velocity at a given time after injection can be corrected by applying correction factors to compensate for the effects of specific physiological or neoplastic microenvironments. The post-injection velocity is given by the formula (e.g., post-injection velocity = 0.4(0.1t)). 2 (-8.1t + 188), where t represents the time after injection in minutes, and the injection velocity is μm. s-1 It can be expressed by (represented by ), and can also be expressed in graph format, tabular format, etc. Post-injection velocity is the translational velocity, i.e., the velocity calculated based on the distance between two points by a straight line. Cells of magnetotactic bacteria are not normally passively pushed or pulled by flagella located on one side of the cell. These cells contribute to displacement efficiency and further increase the displacement velocity in the fluid environment by performing helical motion due to the angle between the long axis of the magnetosome chain and the axis of the flagellar propulsion structure. For this reason, the actual velocity or instantaneous velocity of the therapeutic transporter or therapeutic transport complex is related to the total helical distance in the helical motion. Thus, instantaneous velocity (velocity in the lateral or rotational direction) is different from translational velocity (velocity in the longitudinal direction), and it is the latter (translational velocity) that is actually considered when estimating the total distance traveled.

[0221] Standard deviation of therapeutic agent transport complex velocity (VTCSD)

[0222] Here, the velocity of the therapeutic agent transport complex (VTC) is defined as the post-injection velocity at t=0, when the post-injection velocity is equal to the velocity of the therapeutic agent transport complex. The velocity of the therapeutic agent transport complex here refers to the velocity recorded before shipment from the manufacturing site, and VDI represents the velocity during injection. The standard deviation (SD) of the velocity of the therapeutic agent transport complex is an indicator of the magnitude of the variation in the velocity of the therapeutic agent transport complex, and is referred to as VTCSD in this specification. A small standard deviation of the velocity of the therapeutic agent transport complex indicates that the velocity values ​​of multiple injected therapeutic agent transport complexes tend to be close to the average velocity of those complexes. On the other hand, a large standard deviation of the velocity of the therapeutic agent transport complex indicates that the velocity of the therapeutic agent transport complex is distributed over a wider range.

[0223] Total Bolas Exit Time (TBET)

[0224] Total Bolus Escape Time (BES) refers to the total time it takes for an injected therapeutic agent transport complex bolus to dissolve, i.e., the total time required for all mobile therapeutic agent transport complexes to detach from the bolus. For a single bolus, the total bolus escape time increases with increasing therapeutic agent transport complex concentration (TCC). Increasing the TCC may reduce the volume (VOL) required to inject a given therapeutic dose, which is particularly desirable in cases of high-volume administration. In addition, increasing the TCC increases the viscosity of the bolus, improving its retention within the tissue. For a single therapeutic agent transport complex, the total bolus escape time also depends on the bolus escape surface (BES). In this specification, the bolus escape surface refers to the surface of the injected therapeutic agent transport complex bolus that faces north in the magnetic field configuration applied to a north-facing therapeutic agent transport complex. Therapeutic agent transport complexes located on the bolus escape surface generally detach first from the injected bolus. Depending on the shape of the bolus, the bolus release surface may change as the bolus volume decreases, or it may remain constant. Therefore, by appropriately using various injection techniques to form the injected bolus into an appropriate shape, it is possible to adjust the bolus release surface.

[0225] Major magnetic-oxygen-tactic targeting methods (MATM)

[0226] The magnetic-oxygen-tactic targeting method can be either a single magnetic-oxygen-tactic targeting method (SMATM) or a combined magnetic-oxygen-tactic targeting method (CMATM). Single magnetic-oxygen-tactic targeting methods include targeting of the hypoxia zone based on the air magnetic field strength range, targeting of the hypoxia zone based on the post-injection velocity, targeting of the hypoxia zone based on the velocity standard deviation of the therapeutic agent transport complex, and targeting of the hypoxia zone based on the total bolus withdrawal time. A combined magnetic-oxygen-tactic targeting method is a combination of two or more single magnetic-oxygen-tactic targeting methods.

[0227] Targeting low-oxygen zones based on magnetic field strength range

[0228] In targeting hypoxia zones using magnetic field strength ranges, the hypoxia zone is targeted by changing the magnetic field strength range along the displacement path generated by the magnetotaxis targeting mode or its derived modes. A simple example is shown in Figure 3. In Figure 3, hypoxia zone 2 is the target. A directional magnetic field configuration, represented by the arrow in Figure 3, is applied from the injection site (INJ). To target hypoxia zone 2, a directional magnetic field configuration in the magnetotaxis magnetic field strength range is applied to pass through hypoxia zone 1, and then, once the therapeutic agent transport complex is sufficiently close to or inside hypoxia zone 2, a directional magnetic field configuration in the oxygenotaxis magnetic field strength range is applied.

[0229] Targeting hypoxia zones using magnetic field strength ranges allows for highly precise control over the selection of target hypoxia zones, but requires appropriate information regarding the distribution and location of these zones. Such information is typically collected and estimated in advance using appropriate medical imaging techniques. Furthermore, it is necessary to estimate the distance traveled by the therapeutic agent transport complex from the injection site with relative accuracy in relation to the location of the target hypoxia zone. To maximize the effectiveness of targeting hypoxia zones using magnetic field strength ranges, the standard deviation of the therapeutic agent transport complex's velocity must be relatively low, and the average value must be sufficiently high, thereby ensuring a sufficient HBPIT (Hypoxic Bypass Post Injection Time). In this specification, HBPIT refers to the maximum time after injection during which the injected therapeutic agent transport complex is in a state where it can escape the hypoxia zone when operating within the magnetotaxis magnetic field strength range. This time is primarily determined by the decay (decrease) of the post-injection velocity (and consequently, the decrease in the distance traveled from the hypoxic zone), and this decay is particularly pronounced in magnetotactic bacterial species suitable as therapeutic agent transporters for hypoxic zone targeting. This limits the time during which hypoxic zone targeting by magnetic field strength range is applicable, and the total distance from the injection site that can pass through the hypoxic zone. However, this total distance may be relatively long depending on the post-injection velocity. It is noteworthy that by using hypoxic zone targeting by magnetic field strength range, it is also possible to target different hypoxic zones with therapeutic agent transport complexes of different concentrations. For example, hypoxic zone 2 and hypoxic zone 3 can be targeted with therapeutic agent transport complexes of different ratios. In the example shown in Figure 3, this can be achieved by setting a magnetotactic magnetic field strength range for hypoxic zone 1, a magnetotactic and oxygenotactic magnetic field strength range for hypoxic zone 2, and an oxygenotactic magnetic field strength range for hypoxic zone 3. Note that this is only a simple example, and other embodiments are possible.

[0230] Targeting the hypoxic zone based on injection velocity

[0231] The post-injection velocity-hypoxia zone targeting (VPI-HZT) method typically operates within the magnetotaxis field strength range (although the magnetotaxis-oxygenotaxis field strength range can also be used), targeting the hypoxia zone based on the decay of the post-injection velocity (decrease in post-injection velocity). This method is simplified and shown in Figure 4, where t represents the elapsed time after injection.

[0232] In the simple example shown in Figure 4, the post-injection velocity of the therapeutic transport complex is sufficiently high that it passes through hypoxic zone 1 upon application of a directional magnetic field configuration within the magnetotaxis field strength range. As the therapeutic transport complex continues to move along the displacement path created by the magnetic field configuration, its decayed post-injection velocity decreases further until it reaches a velocity at which it can no longer pass through the hypoxic zone. At this point, the distance traveled in the run state due to the decay of the post-injection velocity is insufficient to reach a position unaffected by the oxygen gradient associated with the hypoxic zone. Therefore, the therapeutic transport complex remains in a specific hypoxic zone, shown as hypoxic zone 2 in the example shown in Figure 4.

[0233] When targeting a specific hypoxic zone using post-injection velocity targeting, it is necessary to accurately estimate the rate of decay of the post-injection velocity and the corresponding distance traveled along the migration path. Because post-injection velocity targeting strongly depends on the decay of the post-injection velocity, the range controllable by the operator is limited. Furthermore, the targeting range from the injection site is limited and defined by the decay of the post-injection velocity.

[0234] Targeting of hypoxic zones by standard deviation of the rate of therapeutic agent transport complexes

[0235] The hypoxic zone targeting method using the standard deviation of the therapeutic agent transport complex velocity (VTCSD-HZT) targets the hypoxic zone by utilizing the standard deviation of the therapeutic agent transport complex velocity, and consequently, the variability in the post-injection velocity of the injected complex. The basic concept of the hypoxic zone targeting method using the standard deviation of the therapeutic agent transport complex velocity is shown in Figures 5 and 6, and mainly depends on the standard deviation of the therapeutic agent transport complex velocity, the average injection velocity of the therapeutic agent transport complex, and the decay rate of the injection velocity, indicated by the leftward-pointing arrows in Figures 5 and 6.

[0236] Figure 5 shows an example of the standard deviation of therapeutic agent transport complex velocities in a batch of therapeutic agent transport complexes, measured at the factory. In this example, only the fastest therapeutic agent transport complexes are selected by excluding the slowest ones. Assuming that the injection velocity, i.e., the post-injection velocity at t=0, is the same as the therapeutic agent transport complex velocity, when operating within the magnetotactic magnetic field intensity range, the therapeutic agent transport complex is likely to pass through the first hypoxic zone near the injection site. This is because the injection velocity and the post-injection velocity immediately after injection for the entire population of therapeutic agent transport complexes exceed the magnetotactic magnetic field intensity range velocity during injection threshold, which is the minimum velocity required to pass through the hypoxic zone. Although the post-injection velocity decreases over time after injection, the hypoxic region is targeted by maintaining the magnetotactic magnetic field intensity range. The rate and distribution of hypoxic zone targeting within the hypoxic zone along the displacement path depend on the standard deviation of the therapeutic agent transport complex velocity. When the standard deviation of the drug transport complex velocity is high, magnetotactic bacteria are widely dispersed within the hypoxic zone along the displacement pathway, resulting in a lower density of drug transport complexes per hypoxic zone. On the other hand, when the standard deviation of the drug transport complex velocity is low, the distribution of drug transport complexes in the hypoxic zone becomes more localized (less spread), resulting in a higher density of drug transport complexes per hypoxic zone.

[0237] Figure 6 shows the injection velocity threshold for the magnetotaxis field strength range, which is set to the average injection velocity of the drug transport complex. This threshold allows for targeting not only hypoxic zones near the injection site but also other hypoxic zones located further away, within the magnetotaxis field strength range. Therefore, even with batches using the same type of drug transport complex, the targeting of hypoxic zones according to the distribution and location of hypoxic zones within the tumor volume can be optimized by designing the system to vary the ratio of drug transport complex populations located to the left or right of the injection velocity threshold based on the velocity standard deviation. This approach is the fundamental idea behind the hypoxic zone targeting method using the velocity standard deviation of drug transport complexes. Generally, when targeting a large portion of hypoxic zones that may be relatively far apart from each other, relying solely on the hypoxic zone targeting method using the velocity standard deviation of drug transport complexes, drug transport complexes with a large velocity standard deviation, i.e., drug transport complexes with a wide distribution of velocity variability, are more suitable.

[0238] Targeting the hypoxic zone based on total bolus withdrawal time.

[0239] The Total Bolus Release Time-Based Hypoxic Zone Targeting (TBET-HZT) method targets hypoxic zones within tumor volume by combining the dissolution rate of the injected therapeutic agent transport complex bolus with the decay of its post-injection velocity. This is typically performed using a directional vector operating within the magnetotaxis field strength range (although the magnetotaxis-oxygenotaxis field strength range can also be used). If the radius of the spherical injection bolus is r, then the bolus surface area = 12.57r. 2 Volume = 4.189 r 3This is expressed as follows. Considering such a surface area-to-volume (S / V) ratio of the injection bolus, dividing a single injection bolus into multiple smaller injection boluses increases the effective bolus release surface area, resulting in a reduction in the total bolus release time. A similar idea applies when the bolus's volumetric shape is changed. For example, if the bolus is cylindrical and positioned so that its longitudinal axis is perpendicular to the direction vector, it will have a higher surface area-to-volume ratio, resulting in a shorter total bolus release time. When a magnetic field configuration within the magnetotaxis field strength range is applied, the total bolus release time is generally equal to or less than the injection velocity threshold within the magnetotaxis field strength range in order to target hypoxic zones within the tumor volume located further away from the injection site. This is because if the total bolus release time exceeds the allowable time corresponding to this injection velocity threshold, a portion of the therapeutic agent transport complex population will remain at the injection site. However, such retention may be desirable in some cases. In addition to the bolus release surface area, there are other factors that influence the total bolus release time and can be used for adjustment; the concentration of the therapeutic agent transport complex is one example.

[0240] Regarding the therapeutic agent transport complexes located on the bolus detachment surface that detach from the bolus and move toward the hypoxic zone, therapeutic agent transport complexes present within the bolus are also exposed to physiological environmental factors that can reduce their motility. Therefore, as time passes after injection, the injection velocity of therapeutic agent transport complexes detaching from the bolus decreases, and the injection velocity of the last therapeutic agent transport complex to detach from the bolus is lower than that of the first therapeutic agent transport complex to detach from the bolus.

[0241] If the total bolus departure time is equal to the motion time of the direction vector in the magnetotaxis field strength range, and the average injection velocity is sufficiently higher than the injection velocity threshold in that field strength range, and furthermore, the standard deviation of the drug transport complex velocity is extremely small, then the drug transport complex that first departs from the injected bolus can be expected to pass through the first hypoxia zone without targeting it, and then target a hypoxia zone further from the injection site once its post-injection velocity has decayed to below the injection velocity threshold in that field strength range. Subsequently, the drug transport complex that departs from the bolus reaches a hypoxia zone located at a shorter distance from the injection site, and therefore targets a hypoxia zone closer to the injection site. On the other hand, the drug transport complex that last departs from the injected bolus cannot pass through the hypoxia zone closest to the injection site due to insufficient post-injection velocity, and as a result, this closest hypoxia zone is also targeted. If the total bolus departure time is adjusted to be significantly shorter, targeting of hypoxia zones close to the injection site may not occur, and instead, targeting of hypoxia zones further from the injection site may increase.

[0242] Combined magnetic-oxygen tactic targeting method

[0243] A combined magnetic-oxygen-tactic targeting method (CMATM) is a combination of two or more individual magnetic-oxygen-tactic targeting methods described in the previous section.

[0244] Although the present invention has been described with reference to preferred embodiments, it will be obvious to those skilled in the art that various modifications and improvements are possible. Such modifications and variations should be understood to be within the spirit and scope of the present invention.

[0245] In the above description, representative non-limiting embodiments of the present invention have been described in detail with reference to the accompanying drawings. This detailed description is intended solely to teach those skilled in the art further details for carrying out preferred embodiments of the present invention and is not intended to limit the scope of the present invention. Furthermore, the various additional features and teachings described above and below may be used alone or in combination with other features and teachings.

[0246] Furthermore, the various combinations of features and steps described in the detailed description and experimental examples above are not essential for carrying out the broadest aspects of the present invention, but are merely provided to specifically illustrate exemplary embodiments of the present invention. Moreover, the features shown in the exemplary embodiments described above, and the features described in the following independent and dependent claims, can be used in combination, even if combinations are not specifically indicated, to provide additional useful embodiments of the teachings.

Claims

1. A system for performing at least one of the acquisition of image information of a subject, diagnosis, and treatment using magnetic and oxygen-tactic bacteria, wherein the magnetic and oxygen-tactic bacteria, after a bolus of magnetic and oxygen-tactic bacteria is injected into the subject, self-propels by run-and-reverse motion and run-and-tumble motion, and is bound to at least one of a therapeutic agent, a diagnostic agent, and an imaging agent, and the system is Processor and The system comprises a memory that stores program code, and when the program code is executed by the processor, the processor... To acquire image information of the target zone within the subject, Applying a magnetic field having a first magnetic field strength to guide and displace the magnetic and oxygen-tactic bacteria toward the target zone having a hypoxic region by magnetotaxis, A magnetic field having a second magnetic field strength lower than the first magnetic field strength is applied so that the magnetic-oxygen-tactic bacteria can follow the oxygen gradient that attracts the bacteria to the hypoxic region. A system that performs the following actions, wherein the change in the direction of movement of the magnetic-oxygen-tactic bacteria from the direction of the magnetic field increases at the second magnetic field strength compared to the first magnetic field strength.

2. The system further includes a user input interface, and when the program code is executed by the processor, the processor will... The system according to claim 1, further comprising an instruction to receive an instruction from a user into the user input interface and to execute the process of generating the acquired image information.

3. The system according to claim 1 or 2, further comprising one or more magnetic sources.

4. The system according to claim 3, wherein the one or more magnetic sources include three pairs of magnetic coils, each pair of the three pairs of magnetic coils is positioned with respect to one different axis from the three axes x, y, and z.

5. The system according to claim 1 or 2, further comprising an imaging device.

6. The system according to claim 5, wherein the imaging device is an MRI device.

7. The system according to claim 1 or 2, wherein the first magnetic field strength is at least 15 gauss.

8. The system according to claim 1 or 2, wherein the value of the second magnetic field strength is less than 15 gauss and 5 gauss or more.

9. The method according to claim 1 or 2, wherein the value of the second magnetic field strength is less than 5 gauss.

10. The program code for determining the first and second magnetic field strengths is: The volume of the bolus, The distance between the injection site of the bolus and the aggregation zone, and The system according to claim 1 or 2, which takes into account the time elapsed after injection.

11. The system according to claim 10, wherein the program code for determining the first and second magnetic field strengths further takes into account the concentration of magnetotactic bacteria in the bolus.

12. The system according to claim 1 or 2, wherein the memory further includes program code that, when executed by the processor, causes the processor to reduce the magnetic field strength of the magnetic field to a second magnetic field strength, and then further reduce the magnetic field strength of the magnetic field to a third magnetic field strength lower than the second magnetic field strength, and the change in direction from the direction of the magnetic field in the movement of the magnetotactic bacteria is greater at the third magnetic field strength than at the second magnetic field strength.

13. The system according to claim 1 or 2, wherein the memory further includes program code, when executed by the processor, causing the processor to calculate the distance between the injection site of the bolus and the aggregated zone or portion of the tumor, and the application of a magnetic field having the second magnetic field strength is performed based on the calculated distance.

14. The system according to claim 1 or 2, wherein the memory further includes program code that, when executed by the processor, causes the processor to estimate the position of the magnetotactic bacteria when the injection velocity of the magnetotactic bacteria has decayed to zero.

15. The system according to claim 1 or 2, wherein the memory further includes program code that, when executed by the processor, causes the processor to determine the first and second magnetic field strengths taking into account the decay of the injection velocity of the magnetotactic bacteria.

16. A non-temporary storage medium that stores instructions executable by a computer device, At least one command to acquire image information of a target zone within the subject, At least one command for applying a magnetic field having a first magnetic field strength to guide and displace magnetotactic bacteria toward a target zone within the subject having a hypoxic region, A medium comprising at least one instruction to apply a magnetic field having a second magnetic field strength lower than the first magnetic field strength, such that the magnetotactic bacteria can follow an oxygen gradient that draws the bacteria to the hypoxic region, wherein the change in direction from the direction of the magnetic field in the movement of the magnetotactic bacteria increases at the second magnetic field strength compared to at the first magnetic field strength.

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