Systems for focused targeting of magnetic-oxygen responsive bacteria and methods of use thereof

Abstract

A method of improving targeted delivery of at least one of a therapeutic agent, an imaging agent and a diagnostic agent attached to a magnetic-oxygen responsive bacterium; the method includes adjusting the total bolus escape time of a solution of magnetic-oxygen-responsive bacteria to affect targeting of a target region having an anoxic region in a patient.

Description

[0001] This disclosure claims priority to U.S. Provisional Patent Application No. 63 / 351,938, filed June 14, 2022, which is incorporated herein by reference; and claims priority to U.S. Provisional Patent Application No. 63 / 351,950, filed June 14, 2022, which is incorporated herein by reference. Technical Field

[0002] This disclosure relates to magnetotaxis, and more specifically to the use of magnetic fields to guide magnetotactic oxygen-responsive bacteria for imaging, diagnosis and / or treatment. Background Technology

[0003] The use of magnetic fields in medical interventions is becoming increasingly common. For example, the following patent documents describe existing medical applications utilizing magnetic fields: U.S. Patent No. 9,655,539, U.S. Patent No. 9,381,063, U.S. Patent No. 9,220,425, U.S. Patent No. 8,986,214, U.S. Patent No. 8,684,010, U.S. Patent No. 8,457,714, US2013,000,6100, US2012,031,0111, US2012,028,982, US2012,028,883, US2011,009,2808, U.S. Patent No. 7,873,401, US2010,003,054,02, US2009,027,582, US2009,024,8014, US2005,009,589, etc.

[0004] Recently, the use of magnetotactic entities under an applied magnetic field has also been studied for medical purposes.

[0005] A magnetotactic entity is defined as an unbound entity whose propulsion source or system responsible for its displacement is part of, attached to, or embedded in the entity itself. Magnetotactic entities include a group of objects or microorganisms, as well as any biological or hybrid system comprising micron- or nanoscale systems or structures made of biological and / or synthetic materials (including chemical, artificial, etc.), and / or components whose directional motion can be influenced by torque induced from a directional magnetic field (e.g., from a permanent magnet) or an electromagnetic field (magnetic fields herein include those generated by an electric current flowing in a conductor), a method referred to herein as magnetotactic, wherein the direction of motion of such magnetotactic entities is influenced by a directional magnetic field (these magnetotactic entities may also be functionalized and attached to other structures). Examples of such magnetotactic entities include, but are not limited to, single or grouped (e.g., clusters, aggregates, clumps, etc.) flagellated magnetotactic bacteria (MTBs), or other bacteria or microorganisms capable of self-propulsion and directional control by a directional magnetic field; these bacteria or microorganisms may be pre-modified by a variety of methods, including but not limited to culture parameters, genetic modification, or attachment to / embedding other entities modified to allow magnetotactic control, such as other cells (including red blood cells), or attachment to synthetic structures that can be affected by a directional magnetic field, or the addition of micro / nanoscale components to bacteria, cells, or other microorganisms to make the realization of such directional movement (including hybrid realizations composed of biological and synthetic components) sensitive to magnetotaxis or directional magnetic fields (such as magnetic fields capable of influencing the orientation of magnetic nanocompasses).

[0006] However, the internal environment of the human body (such as body temperature, pH, and solute concentration) is hostile to magnetotactic bacteria after injection into the subject, reducing the activity and motility of the bacteria. Therefore, protocols are still needed to improve the targeting of magnetotactic bacteria, enabling them to reach the target area before losing their motility or dying. Summary of the Invention

[0007] This disclosure relates to methods for modulating magnetic fields used to guide magneto-oxygen-responsive bacteria for the treatment, diagnosis, and / or imaging of subjects.

[0008] These methods utilize the oxytaxis and magnetotaxis of magnetotactic bacteria. Magnetotactic bacteria are adapted to follow an oxygen gradient from hyperoxygen to hypooxygen and are guided by a magnetic field. This affinity for decreasing oxygen concentration causes the bacteria to be attracted to hypoxic areas within the subject's body (e.g., tumors, but not limited to), where the hypoxic zone is virtually anaerobic. The bacteria are able to move autonomously to the hypoxic area of ​​the target region using their self-propelled systems (e.g., their flagella). Once the bacteria are within the oxygen gradient, this can occur with or without a magnetic field, where the recovery time of the run-and-roll motion increases, causing the bacteria to change direction from the magnetic field.

[0009] Therefore, this method involves adjusting the magnetic field strength to alter bacterial motility patterns based on changes in magnetic field strength. Higher magnetic field strengths favor run-and-roll movements (causing bacterial displacement) because the bacteria will follow the direction of the magnetic field. Under lower magnetic field strengths, magnetotactic bacteria exhibit more run-and-roll movements as they seek oxygen gradients, thus guiding them to hypoxic areas within the subject's body.

[0010] This method utilizes changes in environmental factors (such as magnetic field strength) to alter the motility of magnetotactic bacteria, thereby improving their targeting within the patient's body. By adjusting the magnetic field strength, bacterial movement induced by running and rolling can be controlled. Higher magnetic field strengths reduce changes in the direction of bacterial movement caused by running and rolling, while lower magnetic field strengths increase changes in the direction of movement caused by running and rolling, thus enabling bacteria to locate hypoxic areas.

[0011] Run-reverse motion refers to the long-distance unidirectional swimming of bacteria, typically interrupted by brief reversal events at a specific average non-periodic frequency, resulting in forward displacement. In contrast, run-tumble motion involves the alignment between the direction of movement and the magnetic field direction of magneto-oxygen-responsive bacteria being disturbed by tumble (where the bacterial direction of movement changes from the magnetic field direction), followed by restoration of alignment with the oriented magnetic field during the run phase. If the run phase is shorter than the recovery time, the magnetic field will not have sufficient time to realign with the magneto-oxygen-responsive bacteria before the next tumble, causing the bacterial cells to deviate from the magnetic field direction.

[0012] The method described in this paper can guide magnetotactic-oxygen-responsive bacteria in two or three modes. In the first mode, the magnetic field strength is set to predominate magnetotaxis, where the main bacterial movement is run-tumble; higher magnetic field strength prolongs the run state, exceeding the recovery time during the run-tumble phase. In the second mode, by reducing the magnetic field strength, bacterial movement can be induced by both magnetotaxis and oxygen chemotaxis, where bacteria exhibit more directional changes due to run-tumble movement. In the third mode, the magnetic field strength is even lower than in the second mode, and bacterial movement is primarily induced by oxygen chemotaxis, where bacteria exhibit the main directional changes due to run-tumble movement.

[0013] Understandably, a particular pattern can be used multiple times when guiding bacteria, and the order of patterns can vary depending on the given trajectory of the bacteria, the characteristics of the target area, the location of the target area, etc. Users can select one or more patterns and use them in different orders to reach the target area and achieve the desired accumulation of the anoxic zone.

[0014] The run-and-roll motion enables magnetotactic bacteria to move to different locations within the subject's body (e.g., primarily for longer distances), with the run-and-roll motion improving their ability to locate hypoxic areas. The magnetic field strength can be adjusted based on the desired movement pattern of the magnetotactic bacteria, the characteristics of the target within the subject's body (e.g., tissue density, size, concentration in hypoxic areas), the pre-injection velocity distribution, the post-injection velocity decay, the volume and size of the drug bolus, the concentration of magnetotactic bacteria within the bolus, and the distance between the injection site and the target.

[0015] In some embodiments, because the Magnetic-Oxidative Responsive Bacteria (MORB) move faster when guided by a magnetic field of higher strength, a higher magnetic field strength can be maintained to guide them until one or more oxygen gradients bring them to the target area (e.g., caused by one or more anoxic zones). Once the MORB reaches the target area, reducing the magnetic field strength by lowering the oxygen gradient causes a change in their movement pattern. In this different movement pattern, the MORB follows a run-tumble motion, resulting in a change in their direction of movement. These changes in direction allow the MORB to localize to anoxic zones, where they may remain until their velocity drops to zero after injection (e.g., leading to deposition of the agent adhering to the MORB).

[0016] Therefore, when magnetotactic responsive bacteria attach to diagnostic, imaging, and / or therapeutic agents, they remain (e.g., die) in these hypoxic areas of the tumor, thus leaving the diagnostic, imaging, and / or therapeutic agents at the hypoxic site. In effect, because the diagnostic, imaging, and / or therapeutic agents provide targeted delivery of the drug, this results in a smaller dose required to deliver it to the subject. This reduces toxicity to the subject.

[0017] In some embodiments, the velocity distribution of magnetotactic bacteria in a bolus or sample can be used to predict the behavior of magnetotactic bacteria after injection into a subject or exposure to a magnetic field. For example, a narrower velocity distribution results in less diffusion (and a smaller target volume) of magnetotactic bacteria after injection into a patient and exposure to a magnetic field, because the bacteria have similar initial velocities. However, a wider velocity distribution will result in greater diffusion or distance (and a larger target volume) of magnetotactic bacteria after injection and exposure to a magnetic field over time, because some bacteria move significantly slower than others when exposed to a magnetic field. Greater diffusion among bacteria may cause more dispersed targeting of hypoxic areas when adjusting the magnetic field strength to increase run-tumble motion to locate hypoxic areas. However, because the bacteria are distributed across more hypoxic areas, the bacterial concentration in each hypoxic area will be lower. Conversely, less distance diffusion between magnetotactic bacteria after injection (due to a narrower velocity distribution) will result in more concentrated targeting of hypoxic areas when adjusting the magnetic field strength to increase run-tumble motion to locate hypoxic areas. This is because when the magnetic field strength decreases (which is beneficial for bacteria to change direction from the magnetic field direction to find hypoxic areas due to increased run-tumble motion), the bacteria are already located in a smaller volume in the subject's body, where all injected bacteria have traveled approximately the same distance from the injection site in the subject's body.

[0018] Therefore, the velocity distribution of magnetotactic bacteria can be used (e.g., in preparing injection solutions; in selecting injection solutions or samples) to increase or decrease the number of target anoxic regions and to target the concentration of magnetotactic bacteria in each target anoxic region. If it is desired that the magnetotactic bacteria be diluted in more anoxic regions, a sample with a larger velocity distribution can be selected or prepared. If it is desired that the magnetotactic bacteria be concentrated in fewer anoxic regions (e.g., at a lower dilution), a sample with a smaller velocity distribution can be selected or prepared.

[0019] In one embodiment, with respect to the applied magnetic field, the generated magnetic field provides a directional torque, which can be defined on one or more of three axes using three pairs of magnetic coils, each pair of coils being arranged relative to one axis. These magnetic head pairs can generate a three-dimensional convergence point, towards which magnetotropic entities will be guided and converge. The three-dimensional convergence point (CP) in the magnetic field is an unbounded point in space, towards which entities following the direction of the magnetic field in the aggregation zone (AZ) will move and converge. The effective magnetic field at the convergence point is zero, and around the convergence point in the aggregation zone, the effective magnetic field points towards the convergence point from all directions. Since the magnetic field is not a point source, at least one magnetic field source needs to be time-varying to cause entities to move towards and remain near the convergence point.

[0020] Therefore, maintaining a constant (static) magnetic field on any two axes (x, y, or z), and changing the direction of the third axis while keeping the other two axes constant, will generate a convergence point. Similarly, maintaining one axis constant and changing the directions of the other two axes simultaneously (synchronously) or with phase (delay) in a time-multiplexed manner will be used to generate a convergence point, provided that the frequency (e.g., low frequency, approximately 0.1 to 5 Hz, or preferably approximately 0.5 Hz) at which the magnetotactic responsive bacteria can respond appropriately is changed. The directions of the three axes can also be changed simultaneously or with delay between each of the three axes in a time-multiplexed manner. All combinations are feasible provided that the magnetic field gradient of at least one axis (x, y, or z) is changed in a time-multiplexed manner and the switching speed is suitable for the reaction time of the magnetotactic responsive bacteria. U.S. Patent No. 9,905,347 (the entire contents of which are incorporated herein by reference) describes a system for guiding magnetotactic entities within a subject. U.S. Patent No. 9,905,347 describes 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 in three planes.

[0021] A broad aspect is a method for acquiring imaging information, diagnosis, and treatment of a subject using magnetotactic responsive bacteria adapted for self-movement. The method includes: acquiring imaging information of a target region within the subject, wherein a phallus of magnetotactic responsive bacteria is injected into the subject, the bacteria attaching to at least one of a therapeutic agent, a diagnostic agent, and an imaging agent; applying a magnetic field of a first magnetic field strength to guide and induce displacement of the magnetotactic responsive bacteria toward the target region having a hypoxic area by magnetotaxis; and applying a magnetic field of a second magnetic field strength (lower than the first magnetic field strength) to cause the magnetotactic responsive bacteria to follow an oxygen gradient, thereby attracting the bacteria to the hypoxic area, wherein the change in the direction of movement of the magnetotactic responsive bacteria from the magnetic field direction is increased to a greater extent under the second magnetic field strength than under the first magnetic field strength, for the purpose of treating, diagnosing, and imaging the subject.

[0022] In some embodiments, the method may include acquiring velocity distribution information of magnetotactic bacteria in the drug bolus to predict post-injection diffusion of magnetotactic bacteria in a subject when the magnetic field strength is at a first magnetic field strength.

[0023] In some embodiments, a drug pellet can be selected from a variety of magnetically oxidative response bacterial solutions based on the pre-injection velocity distribution information of the magnetically oxidative response bacteria in the solutions, wherein the selected solutions may contain magnetically oxidative response bacteria whose velocity distribution widens with increasing desired target volume, wherein a larger target volume allows for targeting of more hypoxic regions associated with the target region.

[0024] In some embodiments, when it is desirable to target a larger volume to increase the number of hypoxic regions targeted by the magnetotactic bacteria, the magnetotactic bacteria solution with the widest pre-injection velocity distribution can be selected from these solutions.

[0025] In some embodiments, when it is desirable to concentrate the target to increase the concentration of magnetotactic bacteria that target one or more hypoxic regions, the magnetotactic bacterial solution with the narrowest velocity distribution before injection can be selected from these solutions.

[0026] In some embodiments, the first magnetic field strength and the second magnetic field strength may be determined based on the decay of the velocity after injection of magneto-oxygen-responsive bacteria.

[0027] In some embodiments, the method may include estimating the location of the magnetically oxidative bacteria when the velocity decays to zero after injection.

[0028] In some embodiments, the method may include: calculating the distance between the injection site of the drug bolus and the aggregation area or tumor portion, wherein a magnetic field with a second magnetic field strength is applied based on the calculated distance.

[0029] In some embodiments, magnetotactic bacteria may attach to an imaging agent, wherein the imaging agent is a contrast agent.

[0030] In some embodiments, the contrast agent may be gadolinium.

[0031] In some embodiments, imaging information of the subject's tumor can be obtained using an MRI or CT scanner.

[0032] In some embodiments, the imaging information may include information about the subject's arteries to avoid puncturing the arteries when performing the injection of magnetotactic bacteria into the peripheral region of the tumor.

[0033] In some embodiments, the first magnetic field strength and the second magnetic field strength may be determined and adjusted based on the volume of the drug pellet, the distance between the injection point of the drug pellet and the aggregation area, and the time elapsed after injection.

[0034] In some embodiments, the first magnetic field strength and the second magnetic field strength may be further determined and adjusted based on the concentration of magnetotactic bacteria in the drug bolus.

[0035] In some embodiments, magnetotactic bacteria can be injected into the peripheral region of the tumor.

[0036] In some embodiments, multiple volumes may be injected into the subject at different sites.

[0037] In some embodiments, the method may include: after reducing the magnetic field strength of the magnetic field to a second magnetic field strength, further reducing the magnetic field strength of the magnetic field to a third magnetic field strength lower than the second magnetic field strength, wherein the magnetotactic bacteria may also exhibit a greater degree of change in the direction of movement of the magnetotactic bacteria from the direction of the magnetic field under the third magnetic field strength than under the second magnetic field strength.

[0038] In some embodiments, the value of the second magnetic field strength may be 0 Gauss or higher, but less than 5 Gauss.

[0039] In some embodiments, the first magnetic field strength may be at least 15 Gauss.

[0040] In some embodiments, the second magnetic field strength may be less than 15 Gauss, but greater than or equal to 5 Gauss.

[0041] In some embodiments, the value of the second magnetic field strength may be 0 Gauss or higher, but less than 5 Gauss.

[0042] Another broad aspect is a system for acquiring imaging information, diagnosis, and treatment of a subject after injecting a certain amount of magnetically oxidative bacteria into the subject, wherein the magnetically oxidative bacteria are adapted to move spontaneously by run-invert and run-roll movements, and the magnetically oxidative bacteria attach to at least one of a therapeutic agent, a diagnostic agent, and an imaging agent. The system includes: a processor; and a memory storing program code that, when executed by the processor, causes the processor to acquire imaging information of a target area within the subject's body; applying a magnetic field of a first magnetic field strength to guide and induce displacement of the magnetically oxidative bacteria toward a target area having a hypoxic region by magnetotaxis; and applying a magnetic field of a second magnetic field strength lower than the first magnetic field strength to cause the magnetically oxidative bacteria to follow an oxygen gradient, thereby attracting the bacteria to the hypoxic region, wherein the change in the direction of movement of the magnetically oxidative bacteria from the magnetic field direction is greater under the second magnetic field strength than under the first magnetic field strength.

[0043] In some embodiments, the system may include a user input interface, wherein the program code further includes instructions that cause the processor to receive user input instructions from the user input interface when executing the program code, in order to generate the acquired imaging information.

[0044] In some embodiments, the system may include one or more magnetic field sources.

[0045] In some embodiments, the one or more magnetic field sources may include three pairs of magnetic coils, wherein each of the three pairs of magnetic coils may be arranged relative to an independent axis of three axes: x, y, z.

[0046] In some embodiments, the system may include an imaging device.

[0047] In some embodiments, the imaging device may be an MRI machine.

[0048] In some embodiments, the first magnetic field strength may be at least 15 Gauss.

[0049] In some embodiments, the second magnetic field strength may be less than 15 Gauss, but greater than or equal to 5 Gauss.

[0050] In some embodiments, the program code used to determine the first magnetic field strength and the second magnetic field strength may take into account the following items: the volume of the drug bolus, the distance between the injection point of the drug bolus and the aggregation area; and the time elapsed after injection.

[0051] In some embodiments, the program code used to determine the first magnetic field strength and the second magnetic field strength may also take into account the concentration of magnetotactic bacteria in the bolus.

[0052] In some embodiments, the memory may contain program code that, when executed by a processor, causes the processor to further reduce the magnetic field strength to a third magnetic field strength below the second magnetic field strength after reducing the magnetic field strength to a second magnetic field strength, wherein the magnetotactic bacteria also exhibit a greater degree of change in the direction of movement of the magnetotactic bacteria from the direction of the magnetic field at the third magnetic field strength than at the second magnetic field strength.

[0053] In some embodiments, the memory may contain program code that, when executed by a processor, causes the processor to calculate the distance between the injection site of the drug bolus and the aggregation area or tumor portion, wherein a magnetic field with a second magnetic field strength is applied based on the calculated distance.

[0054] In some embodiments, the memory may contain program code that, when executed by a processor, causes the processor to estimate the location of the magnetically oxidative bacteria when its speed decays to zero after injection.

[0055] Another broad aspect is a non-transitory storage medium storing instructions executable by a computing device, including: at least one instruction for acquiring imaging information of a target area within a subject's body; at least one instruction for applying a magnetic field of a first magnetic field strength to guide and induce displacement of magnetotactic bacteria toward a target area with a hypoxic region within the subject's body; and applying a magnetic field of a second magnetic field strength lower than the first magnetic field strength to cause the magnetotactic bacteria to follow an oxygen gradient, thereby attracting the bacteria to the hypoxic region, wherein the change in the direction of movement of the magnetotactic bacteria from the direction of the magnetic field is increased to a greater extent under the second magnetic field strength than under the first magnetic field strength.

[0056] Another broad aspect is a method for selecting magnetically oxidative responsive bacterial samples, the bacteria being attached to at least one of an imaging agent, a targeting agent, and a diagnostic agent. This selection is based on concentrated or diluted targeting of a target region with hypoxic areas within a subject's body. The magnetically oxidative responsive bacteria are guided to the target region within the subject's body via magnetotaxis using a magnetic field and seek out hypoxic areas within the subject's body through oxidative attraction. The method includes selecting magnetically oxidative responsive bacterial samples from a variety of samples based on their pre-injection velocity distribution. Specifically, when the target volume is large and the number of target hypoxic areas is large, samples with a larger pre-injection velocity distribution are selected; and when the target volume is small and the number of target hypoxic areas is small, samples with a smaller pre-injection velocity distribution are selected for more concentrated targeting of the magnetically oxidative responsive bacteria.

[0057] Another broad aspect is a method for preparing a magnetically oxidative-responsive bacterial solution for parenteral administration to a subject to achieve a desired level of target volume at a target area in the subject, wherein a magnetic field is used to guide the administered magnetically oxidative-responsive bacteria to the target area in the subject's body via magnetotaxis, and oxygen tropism is used to locate hypoxic areas in the subject's body. The method includes: selecting magnetically oxidative-responsive bacteria based on their magnetotactic self-movement velocity to obtain a selected group of magnetically oxidative-responsive bacteria for the solution, wherein the velocity distribution of the selected magnetically oxidative-responsive bacteria increases with increasing desired target volume; and preparing a solution containing the selected magnetically oxidative-responsive bacteria, wherein the selected magnetically oxidative-responsive bacteria are attached to at least one of an imaging agent, a contrast agent, and a diagnostic agent.

[0058] Another broad aspect is a method for improving the targeted delivery of at least one of a therapeutic agent, imaging agent, and diagnostic agent attached to magnetically oxytactic (MOO)-responsive bacteria, the MOO-responsive bacteria being adapted to move spontaneously in the presence of a magnetic field, depositing at least one of the therapeutic agent, imaging agent, and diagnostic agent in a hypoxic region within a patient's body. The method includes: adjusting the total puff escape time of the MOO-responsive bacterial solution upon injection into the patient to influence targeting of a target region with hypoxia within the patient's body, wherein shortening the total puff escape time facilitates targeting of the MOO-responsive bacteria to one or more hypoxic regions distant from the injection site of the MOO-responsive bacteria, while taking into account the reduced post-injection velocity of the MOO-responsive bacteria when exposed to the patient's environment.

[0059] In some embodiments, the total flotation escape time can be adjusted by adjusting one or more of the following: the concentration of magnetotactic bacteria in the solution; the volume of the flotation; the viscosity of the solution; and the flotation's exposure to a surface facing north of the magnetic field when the magnetotactic bacteria are north-seeking, or the flotation's exposure to a surface facing south of the magnetic field when the magnetotactic bacteria are south-seeking.

[0060] In some embodiments, the total drug escape time can be adjusted by dividing the drug bolus into smaller boluses and injecting them into the patient during injection, thereby reducing the bolus volume because the individual volume of each smaller bolus is smaller than the volume of the bolus.

[0061] In some embodiments, the total drug escape time can be extended by heating the solution in the injection device before injecting it into the patient.

[0062] In some embodiments, the total drug escape time can be prolonged by reducing the volume rate at which the drug is injected into the patient.

[0063] In some embodiments, the total pill escape time can be adjusted by changing the shape of the pill.

[0064] In some embodiments, shortening the total drug escape time can be achieved to access tumor portions inaccessible to the injection device, requiring magneto-oxygen-responsive bacteria to travel from the injection point to the tumor portion.

[0065] In some embodiments, the total bolus escape time can be adjusted by selecting an appropriate syringe to influence the shape of the bolus.

[0066] In some embodiments, the total drug escape time can be shortened by increasing the surface area to volume ratio of the injected drug mass.

[0067] In some embodiments, when it is desirable to target hypoxic areas closer to the injection site, it may be desirable to prolong the total drug escape time.

[0068] Another broad aspect is a method for influencing the total drug escape time of a solution of magnetically oxytactic bacteria attached to at least one of a therapeutic agent, imaging agent, and diagnostic agent injected into a patient, wherein the magnetically oxytactic bacteria are adapted to move spontaneously when subjected to a magnetic field, and the magnetically oxytactic bacteria deposit at least one of the therapeutic agent, imaging agent, and diagnostic agent in a target area with an anaerobic zone within the patient's body. The method includes: shortening the total drug escape time of magnetically oxygen-responsive bacteria by at least one of the following: increasing the surface area to volume ratio of the drug float, decreasing the concentration of magnetically oxygen-responsive bacteria in the solution, decreasing the viscosity of the solution, and increasing the exposure of the drug float to the surface north of the magnetic field for north-seeking magnetically oxygen-responsive bacteria, and increasing the exposure of the drug float to the surface south of the magnetic field for south-seeking magnetically oxygen-responsive bacteria; or prolonging the total drug escape time of magnetically oxygen-responsive bacteria by at least one of the following: decreasing the surface area to volume ratio of the drug float, increasing the concentration of magnetically oxygen-responsive bacteria in the solution, increasing the viscosity of the solution, and decreasing the exposure of the drug float to the surface north of the magnetic field for north-seeking magnetically oxygen-responsive bacteria, and decreasing the exposure of the drug float to the surface south of the magnetic field for south-seeking magnetically oxygen-responsive bacteria, wherein the shorter total drug escape time of the magnetically oxygen-responsive bacteria is used to target one or more hypoxic areas in a target region of the patient body that is far from the injection site.

[0069] In some embodiments, when the injection device cannot access areas near one or more hypoxic zones within the tumor that are further from the injection site, and magnetotactic responsive bacteria need to travel further from the injection site to the hypoxic zones, targeting these hypoxic zones may be considered. This method includes shortening the total drug escape time of the magnetotactic responsive bacteria by at least one of the following: increasing the surface area to volume ratio of the drug pouch, reducing the concentration of magnetotactic responsive bacteria in the drug pouch, reducing the viscosity of the drug pouch, and increasing the exposure of the drug pouch to the surface north of the magnetic field for north-seeking magnetotactic responsive bacteria, and increasing the exposure of the drug pouch to the surface south of the magnetic field for south-seeking magnetotactic responsive bacteria.

[0070] In some embodiments, the method may include increasing at least one of the surface area to volume ratio of the drug pellet by dividing the pellet into smaller pellets.

[0071] In some embodiments, magnetotactic bacteria may attach to the therapeutic agent.

[0072] In some embodiments, the drug pellets are prepared by further considering the velocity distribution of magnetotactic bacteria within the pellet.

[0073] Another broad aspect is a container that uses a magnetic field to supply a quantity of magnetically oxidative bacteria into a subject for treatment, diagnosis, and / or imaging, the container comprising: an identifier adapted to be scanned to provide one or more characteristics of the magnetically oxidative bacteria within the container; and the magnetically oxidative bacteria located within the container.

[0074] In some embodiments, the one or more characteristics include at least one of the following: the attenuation of the activity of magnetotactic bacteria; the density of magnetotactic bacteria; the polarity or polarity ratio of magnetotactic bacteria; the reaction time of magnetotactic bacteria to a change in the direction of a magnetic field; and the maximum velocity or velocity distribution of magnetotactic bacteria.

[0075] In some embodiments, the identifier may be a barcode.

[0076] In some embodiments, the identifier may be a QR (quick response) code.

[0077] Another broad aspect is a method for characterizing one or more properties of magnetically oxidative bacteria in a sample of magnetically oxidative bacteria, comprising: generating a video of the magnetically oxidative bacteria observed under a microscope while the bacteria are subjected to a magnetic field having a known magnetic field strength and direction; and analyzing the behavior of the magnetically oxidative bacteria in the video to determine the one or more properties.

[0078] In some embodiments, the one or more characteristics include the velocity distribution of magnetotactic bacteria, wherein the analysis includes: calculating the distance traveled by the magnetotactic bacteria over a period of time and dividing the distance by the time.

[0079] In some embodiments, the one or more characteristics include the decay of activity of magnetotactic bacteria, wherein the analysis includes: periodically calculating the distance traveled by magnetotactic bacteria over a period of time at different time intervals, and dividing the distance by time to determine the velocity of magnetotactic bacteria at a given time, wherein the decay of activity is determined by measuring the decrease in velocity over different time intervals.

[0080] In some embodiments, the one or more properties include the polarity of magnetotactic bacteria, wherein the analysis includes measuring the proportion of bacteria guided along a direction corresponding to the magnetic north or south pole of a magnetic field.

[0081] In some embodiments, the viscosity of the bacterial solution can be determined by selecting the solvent constituting the solution, adjusting the concentration of solutes or salts present in the solution, adjusting the concentration of magnetotactic bacteria in the solution, and influencing the viscosity. Attached Figure Description

[0082] The invention will be better understood through the following detailed description of embodiments of the invention with reference to the accompanying drawings: Figure 1 This is a schematic diagram of exemplary guided magneto-oxygen-responsive bacteria attached to an imaging agent and exemplary guided magneto-oxygen-responsive bacteria attached to a therapeutic agent; Figure 2 This is a schematic diagram of an exemplary guideable magneto-oxygen-responsive bacterium attached to both an imaging agent and a therapeutic agent; Figure 3 It is a schematic map illustrating an exemplary hypoxic zone that magnetotactic bacteria will traverse from the injection site; Figure 4 This is another schematic map illustrating an exemplary hypoxic zone that magnetotactic bacteria will traverse from the injection site; Figure 5 This is an exemplary graph showing the standard deviation of the velocity-therapeutic transport complex of magnetotactic responsive bacteria coupled with a drug (e.g., a therapeutic agent; a diagnostic agent; an imaging agent), where the velocity of the magnetotactic responsive bacteria is greater than a threshold velocity. This indicates that when a magnetic field strength favorable to magnetotaxis is applied, at least most of the magnetotactic responsive bacteria will cross one or more hypoxic zones, resulting in more concentrated targeting of the magnetotactic responsive bacteria to the target hypoxic zone farther from the injection site. Figure 6This is another exemplary graph showing the standard deviation of the velocity-therapeutic transport complex of magnetotactic responsive bacteria coupled with agents (e.g., therapeutic agents; diagnostic agents; imaging agents), where only a portion of the magnetotactic responsive bacteria have a velocity greater than the threshold velocity, indicating that magnetotactic responsive bacteria have greater diffusion or dilution in more hypoxic areas when a magnetic field strength favorable to magnetotaxis is applied. Figure 7 This is a flowchart of an exemplary method for modulating a magnetic field for guiding magnetotactic bacteria to treat, diagnose, and / or image a subject; Figure 8 This is a block diagram of an exemplary system for targeted imaging, diagnosis, and / or treatment of subjects using magnetotactic bacteria; Figure 9A is a schematic diagram showing the volume of a large target in the target area; Figure 9B is a schematic diagram showing the volume of a small target in the target area. Detailed Implementation

[0083] This disclosure describes a method for modulating a magnetic field to guide magneto-aerotactic-responsive bacteria for the treatment, diagnosis, and / or imaging of a subject. The magneto-aerotactic-responsive bacteria respond to a magnetic field (magnetism) and an oxygen gradient (oxytaxis).

[0084] Therefore, depending on the location of the magnetotactic bacteria, the magnetic field strength can be increased and / or decreased to alter the movement pattern of the magnetotactic bacteria. Specifically, the magnetic field strength that induces magnetotaxis promotes displacement from run-and-reverse motion (where the change in the direction of movement of the bacteria along the direction of the magnetic field is reduced because the running state is longer than the recovery time), while the magnetic field strength that mainly induces oxygenation promotes a change in the direction of movement of the bacteria relative to the direction of the magnetic field through run-and-tumble motion.

[0085] It should be understood that magnetotactic responsive bacteria move by self-propulsion, guided by the torque applied by a magnetic field. Once the arrival of the magnetotactic responsive bacteria in a target area with an oxygen gradient (e.g., a tumor) is identified or estimated, the magnetic field strength can be reduced. The lower magnetic field strength allows the magnetotactic responsive bacteria to seek out hypoxic areas by following the oxygen gradient. The magnetotactic responsive bacteria follow the oxygen gradient, employing a more run-and-roll movement pattern, thus allowing them to migrate to hypoxic areas. Once located in a hypoxic area, the magnetotactic responsive bacteria remain in or near these areas. Once the velocity of the magnetotactic responsive bacteria reaches zero (e.g., death or weakening in the subject), the bacteria deposit one or more agents (therapeutic; imaging; diagnostic) in the hypoxic area. This enables targeted therapy, imaging, and / or diagnostics, reduces the required agent dosage, and consequently reduces the impact of the agent on the subject's health.

[0086] Magneto-oxygen-responsive bacteria: In this disclosure, magnetotactic-oxygen-responsive bacteria refer to a group of bacteria possessing flagella for self-propulsion and chains of magnetosomes for orientation, which respond to magnetic fields by orienting and migrating along geomagnetic field lines (i.e., magnetotaxis). The magnetotactic properties of magnetotactic-oxygen-responsive bacteria can be conferred by specific intracellular organelles (called magnetosomes), which comprise nanoscale, membrane-encapsulated magnetic iron mineral particles (crystals) organized into chains by a specialized cytoskeleton. The magnetic iron mineral particles may include iron oxides, such as magnetite (Fe3O4), or iron sulfides, such as pyrite (Fe3S4) or pyrite (FeS2). Magnetotactic-oxygen-responsive bacteria also respond to oxygen gradients (i.e., oxygen tropism). Examples of magnetotactic-oxygen-responsive bacteria include various species of Magnetococcus marinus (e.g., MO-1 strain, MSR-1 strain, MC-1 strain, BM-1 strain, or genetically modified versions of the species or strains, including those optimized in the laboratory). Magneto-oxygen-responsive bacteria can be naturally occurring, adapted through natural selection, or adapted through human intervention (such as in a laboratory setting), where certain bacterial characteristics may be selected or optimized (e.g., bacterial sensitivity to magnetic fields). Such human interventions may include, but are not limited to, gene manipulation, selecting and propagating bacteria exhibiting certain characteristics, and modifying the environment and / or culture medium to optimize certain characteristics or acquire certain bacterial traits. A common characteristic of magneto-oxygen-responsive bacteria is their ability to self-propel (in some cases, they can be actuated by an external source that induces magnetic torque, enabling them to move). Magneto-oxygen-responsive bacteria can be magnetically guided and modified to respond to external stimuli other than magnetic fields, enabling them to move and / or react.

[0087] In one embodiment, the magnetotactic responsive bacteria according to this disclosure are non-pathogenic, for example, they do not proliferate in the subject or cause bacterial infection. The magnetotactic responsive bacteria may be derived from non-pathogenic magnetotactic spirilla strains (e.g., AMB-1), non-pathogenic magnetotactic coccus strains, non-pathogenic magnetotactic spirilla strains, non-pathogenic magnetotactic spirilla strains (e.g., MSR-1), non-pathogenic magnetotactic bellicus strains, non-pathogenic facultative anaerobic magnetotactic spirilla strains, or non-pathogenic obligate anaerobic strains, such as non-pathogenic desulfovibrio magneticus strains (e.g., RS-1). In one embodiment, the magnetotactic bacteria are magnetotactic coccus strains. It should be understood that non-pathogenic bacteria may still cause mild symptoms in the subject due to the presence of bacterial components that trigger mild inflammation (such as LPS), but are still considered non-pathogenic if they do not proliferate extensively in the subject or do not cause more severe symptoms.

[0088] Magneto-oxygen-responsive bacteria are able to follow a decreasing oxygen gradient until they reach an oxygen level below about 2%. Normal oxygen levels (sometimes referred to as “physiological oxygen”) in normal tissues vary by tissue, averaging about 5-6% (ranging from about 7.5% to 4% depending on the tissue) (McKeown MA, (Br J Radiol. ) 2014; 87(1035)). Therefore, the magneto-oxygen-responsive bacteria according to this disclosure can migrate from areas of tissue or organ with normal oxygen levels (physiological oxygen) (e.g., about 4% to about 7.5% or about 5-6%) to areas with reduced oxygen levels (hypoxia), such as areas of tissue or organ with oxygen levels of about 2% or less, about 1.75% or less, or about 1.5% or less. In one embodiment, the magneto-oxygen-responsive bacteria according to this disclosure are able to follow a decreasing oxygen gradient until they reach an oxygen level of about 0.3% to about 0.7%, about 0.4% to about 0.6%, or about 0.5%. For example, this property can be used to reach hypoxic areas within organs, tissues, or tumors. Most tumors typically have a median oxygen level of 2% or less, and some have 1% or less (McKeown MA, (Br J Radiol.) 2014; 87(1035)). However, significant heterogeneity exists within individual tumors, and therefore some areas of a tumor have oxygen levels below 1%, for example, about 0.5% to 1% or lower. Such hypoxic areas of a tumor are often more resistant to cancer treatments such as chemotherapy and / or radiation therapy.

[0089] In some examples, the survival time of magnetotactic responsive bacteria in the subject may be only a shortened period. The magnetotactic responsive bacteria may die within a short time after being administered to the subject. Therefore, when magnetotactic responsive bacteria contain a therapeutic agent, once the magnetotactic responsive bacteria die, the agent (e.g., for treatment; diagnosis; imaging) can remain at the site of interest.

[0090] Characteristics of magnetotactic bacteria: It should be understood that magnetotactic responsive bacteria can be north-seeking (tending towards the magnetic north pole), south-seeking (tending towards the magnetic south pole), or a mixture of both. The magnetic field sequence selected according to this disclosure may depend on whether the magnetotactic responsive bacteria are north-seeking, south-seeking, or a combination of both. A sample or bolus of magnetotactic responsive bacteria can be tested before injection into a subject (e.g., by subjecting the bacteria to a magnetic field with known properties and monitoring their response to the magnetic field; in some examples, a microscope may be used, and a video of the bacteria's behavior may be generated using a camera and microscope) to determine whether the characteristics of the magnetotactic responsive bacteria are south-seeking, north-seeking, or a combination of both (the ratio of south-seeking to north-seeking magnetotactic responsive bacteria can also be determined).

[0091] In some cases, other characteristics of the magnetically oxidative bacteria in a sample (e.g., to be administered to a subject) can be determined (e.g., by generating a video of the magnetically oxidative bacteria observed under a microscope using a microscope combined with a camera). The velocity of the magnetically oxidative bacteria can be established by dividing the distance traveled (e.g., average distance traveled) by time. A velocity or velocity distribution can be established for a given bacterial population or sample. The response time of the magnetically oxidative bacteria to a change in magnetic field orientation can also be determined by monitoring the reaction time of the bacteria (i.e., the time required for the bacteria to change direction to realign with the new magnetic field direction after a change in magnetic field orientation). The velocity distribution (or velocity) of the magnetically oxidative bacteria and / or their response time to a change in magnetic field orientation can be used to determine the minimum magnetic field strength required to cause displacement of the bacteria (or a magnetic field strength low enough to allow the bacteria to preferentially seek out hypoxic regions within the subject).

[0092] Changes in the direction of the magnetic field can also provide information about the polarity of the magnetotactic bacteria in the sample (i.e., north-seeking, south-seeking, or a mixture of both).

[0093] The density of magnetically oxidative bacteria can also be determined by dividing the number of magnetically oxidative bacteria in a solution of a given sample by a known surface area or volume (e.g., by video taken with a microscope, or by direct observation of the magnetically oxidative bacteria using a microscope).

[0094] The activity decay of magnetically oxidative bacteria in a given sample can also be measured. The velocity change of magnetically oxidative bacteria over time can be measured. Magnetically oxidative bacteria can be observed using a camera (mounted to a microscope), a magnetic field source for applying a magnetic field to the sample containing the bacteria, and one or more sensors (e.g., a pH monitor for measuring pH, a thermometer for measuring temperature, etc.) to characterize the environment of the bacteria. At a given time interval (e.g., every five minutes), the velocity or velocity distribution of the magnetically oxidative bacteria in the sample can be calculated (i.e., by dividing the distance traveled by the bacteria by time). The activity decay of the bacteria can then be calculated by plotting the measured velocity decay of the bacteria over time (the velocity is calculated based on the distance measured at different time intervals). The activity decay of the bacteria can be calculated under different environmental conditions (e.g., at 37 °C, i.e., the average temperature in the subject's body).

[0095] The measurable characteristics of magnetotactic bacteria can be stored in association with an identifier or code (e.g., barcode or QR code) affixed to the sample container of the magnetotactic bacteria. Scanning the identifier or code extracts information defining the characteristics of the magnetotactic bacteria in a given sample. These characteristics can be used to guide the injection of the magnetotactic bacteria into a subject and / or to apply an appropriate magnetic field or magnetic field sequence.

[0096] definition: In this disclosure, "imaging" refers to medical techniques that allow the acquisition of information about a subject. In some examples, imaging allows observation of the subject's anatomical structures. Such techniques include, but are not limited to, magnetic resonance imaging (MRI), computed tomography (CT scan, CAT scan), positron emission tomography (PET), single-photon emission computed tomography (SPECT), electron paramagnetic resonance imaging (EPMRI), ultrasound, X-rays, etc. In other examples, imaging can be used to collect information about a subject, such as the presence of a drug at a specific location within the subject's body. For example, imaging can be a radiation detector with a defined aperture (e.g., a small aperture in some cases) capable of detecting radiation at a given site on the subject (e.g., near an organ).

[0097] In this disclosure, a "drug bolus" refers to a dose of solution administered to a subject, thereby occupying a space within the subject's body at the injection site. After or during injection, it has a volume occupying the subject's body at the injection site. Therefore, a drug bolus has a given volume. In this application, a drug bolus includes a concentration of magnetotactic responsive bacteria in a solution to which imaging agents, diagnostic agents, and / or imaging agents may be attached. The shape of the drug bolus can be influenced by injection techniques, such as moving the needle while injecting the solution of magnetotactic responsive bacteria into the subject, or can be altered depending on the injection device used (e.g., selection of a syringe with a specific injection head—a multi-head syringe, a single-head syringe, etc.).

[0098] In this disclosure, “to make a diagnosis” or “diagnosis” means to determine (i) the presence of a condition, disease or disorder, (ii) the risk of developing a condition, disease or disorder, or (iii) the state of a condition, disease or disorder (worsening, no change or improvement).

[0099] In this disclosure, "imaging agent" refers to an agent used to provide more information about internal organs, cellular processes, and tumors and normal tissues. It may be a contrast agent capable of enhancing the contrast of structures or liquids that can be captured using medical imaging. Such contrast agents may 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), perfluorobromoethane, fluorodeoxyglucose, protein imaging agents, microbubble imaging agents, etc. In some examples, the imaging agent may be present in a carrier (e.g., vesicles) for containing the imaging agent. The imaging agent may be attached to or bound to the surface of magnetotactic bacteria. In some examples, the imaging agent may be an agent capable of emitting radiation that can be captured by a radiation detector (e.g., a radioactive isotope). In some examples, the imaging agent may be bound to magnetotactic bacteria, for example, when the magnetotactic bacteria are magnetotactic bacteria, the magnetotactic bacteria may be cultured in a medium containing a radioactive isotope. In some examples, protein supply may also be integrated into magnetotactic-oxygen-responsive bacteria. In some examples, radiation-emitting imaging agents may be attached to the surface of magnetotactic-oxygen-responsive bacteria.

[0100] In this disclosure, "subject" refers to both mammals and non-mammals. Mammals refer to any member of the class Mammalia, including but not limited to humans. Non-mammals include birds, reptiles, etc. The term "subject" is not limited by sex or age.

[0101] In this disclosure, the term "target area," also referred to herein as "target region," refers to the site within a subject's body that magnetotactic bacteria must navigate toward. The target area includes hypoxic regions toward which magnetotactic bacteria can move by following an oxygen gradient through oxygen attraction. Magnetotactic bacteria can aggregate in the hypoxic areas of the target area. The target area can be a tumor, part of a tumor, an organ of the subject (where at least part of the tumor includes a hypoxic area), etc. The target area may include one or more hypoxic avascular regions within the subject's body caused by or associated with conditions such as ischemic stroke, pulmonary hypertension, ischemic heart disease, diabetic retinopathy, etc.

[0102] In this disclosure, “to treat” or “treatment” means (i) preventing some or all of a symptom, disease or disorder (temporary or permanent), (ii) suppressing or stopping some or all of a symptom, disease or disorder (temporary or permanent), and (iii) alleviating some or all of a symptom, disease or disorder (temporary or permanent).

[0103] In this disclosure, "transport therapy" refers to a therapy based on directional control or guided transport of therapeutic agents, providing a non-systemic delivery approach to therapeutic agents that utilizes transporters (such as magnetotactic bacteria) to deliver therapeutic agents in specific volumes at specific physiological sites (e.g., using a more direct delivery pathway between the injection (or entry) site and the physiological region to be treated). Unlike systemic delivery used in, for example, chemotherapy, the main advantage of transport therapy is minimizing systemic toxicity or systemic exposure while improving therapeutic efficacy, typically achieving a higher delivery dose to the treatment site with a lower injection dose compared to chemotherapy. Transport therapy can also transport therapeutic agents to tumor masses, overcoming the diffusion limitations of standard chemotherapy agents. Here, transport therapy is used as a general term and includes, but is not limited to, non-systemic delivery of diagnostic (transport diagnostic) and imaging (transport imaging) agents. Transport therapy used for, for example, delivering radiosensitizers in radiotherapy can specifically refer to transport radiotherapy. Similarly, the use of transport therapy in immunotherapy can be termed transport immunotherapy, while transport chemotherapy refers to non-systemic chemotherapy using such transporters. Combined therapy with transport therapy can also be used. One example is the combination of chemotherapy and immunotherapy, which can be termed transport chemotherapy immunotherapy. A transporter carrying both a therapeutic agent and a diagnostic agent can be called a transport therapy complex. Many combinations are possible. For example, transport chemoimaging can be achieved by simultaneously or separately injecting a transport chemotherapy complex with a transport imaging (or transport contrast) complex; or by using a transport chemoimaging complex only in a single injection. The same type of nomenclature applies to other types of transport therapy complexes or therapeutic agents; transport therapy remains the common term.

[0104] In this disclosure, "velocity distribution," also referred to as "velocity standard deviation," refers to the velocity distribution or deviation of a group of magnetotactic responsive bacteria found in a given sample or solution (e.g., for administration to a subject) under magnetotactic influence. The velocity distribution is the pre-injection velocity. The velocity distribution can be estimated or obtained using known imaging techniques (e.g., by microscopy) or through a selection process in which magnetotactic responsive bacteria are first cultured, and optional combinations of their velocities are made (and samples or solutions can be further prepared by selecting cultures exhibiting a target velocity or velocity range)...

[0105] Magneto-oxygen-responsive bacteria: For reference Figure 1 An exemplary simplified schematic diagram of magnetotactic bacteria 100 and magnetotactic bacteria 200 is shown.

[0106] Magneto-oxygen responsive bacteria 100 and 200 have magnetosome chains 104 that are sensitive to magnetic fields (such as an applied magnetic field or the geomagnetic field).

[0107] In some embodiments, the therapeutic agent 202 may be contained within a carrier 201. The carrier 201 may have a hydrophilic portion and a hydrophobic portion (e.g., a phospholipid bilayer). Depending on the hydrophobicity or hydrophilicity of the therapeutic agent, the therapeutic agent may be located within the hydrophilic or hydrophobic portion of the carrier 201. In some examples, the therapeutic agent 202 may be contained within a biodegradable polymer carrier. It should be understood that the therapeutic agent 202 may be contained within any carrier that effectively accommodates the therapeutic agent 202 to achieve its delivery.

[0108] In some examples, therapeutic agent 202 may use, for example, a ligand 205 (such as an antibody) to attach to magnetotactic-responsive bacteria 200. Ligand 205 may attach to therapeutic agent 202, wherein the ligand may be adapted to bind to the exterior of magnetotactic-responsive bacteria 200. Ligand 205-therapeutic agent 202 and magnetotactic-responsive bacteria 200 may be co-presented in solution, wherein the ligand-therapeutic agent attaches to magnetotactic-responsive bacteria 200.

[0109] In some examples, therapeutic agent 202 may be present in vesicle 201, wherein vesicle 201 may be linked to a ligand (such as an antibody) adapted to bind to magnetotactic-responsive bacteria 200. Vesicle 201 may then be conjugated to the membrane of magnetotactic-responsive bacteria via ligand 205.

[0110] In some examples, to enable the magnetic-oxygen-responsive bacteria 200 to adhere to the carrier 201 containing the therapeutic agent 202, the magnetic-oxygen-responsive bacteria 200 may be forced to activate lipopolysaccharide. For example, the magnetic-oxygen-responsive bacteria 200 may be first mixed in a medium with a low nutrient concentration and then injected into a new medium containing a concentration of the therapeutic agent carrier.

[0111] It should be understood that other mechanisms for immobilizing therapeutic agent 202 to magnetotactic bacteria 200 may be used without departing from this teaching. 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. 2014 May 27;8(5):5049-60, which is incorporated herein by reference.

[0112] Depending on the application, other coupling mechanisms may be provided between the magnetotactic bacteria and the therapeutic agent 202.

[0113] In some embodiments, the magnetic-oxygen-responsive bacteria do not have any carrier 201, wherein the therapeutic agent 202 may be directly attached to the magnetic-oxygen-responsive bacteria 200 or may be bound to the interior of the magnetic-oxygen-responsive bacteria 200 (wherein the magnetic-oxygen-responsive bacteria may be used as a carrier for transporting the therapeutic agent).

[0114] In some examples, the therapeutic agent 202 and the imaging agent 101 may be the same (e.g., in the case of superparamagnetic iron oxide nanoparticles).

[0115] In some examples, both imaging agent 101 and therapeutic agent 202 can be immobilized on magnetotactic bacteria (e.g., ...). Figure 2 Magnetotaxis-responsive bacteria (300).

[0116] In some examples, imaging agent 101 may use, for example, ligand 105 (such as an antibody) to attach to magneto-oxygen-responsive bacteria 100. Ligand 105 may attach to imaging agent 101, wherein the ligand may be adapted to bind to the exterior of magneto-oxygen-responsive bacteria 100. The ligand combined with the imaging agent may be co-presented in solution with magneto-oxygen-responsive bacteria 100, wherein the ligand combined with the imaging agent attaches to magneto-oxygen-responsive bacteria 100.

[0117] It should be understood that other mechanisms for immobilizing the imaging agent 101 on the magneto-oxygen-responsive bacteria 100 may also be used without departing from this teaching.

[0118] In some examples, the ratio, number, and / or concentration (or relative concentration) of magnetotactic responsive bacteria to other magnetotactic responsive bacteria are known. When the number of magnetotactic responsive bacteria arriving at and / or remaining at the site of interest (e.g., at least one hypoxic area of ​​a tumor) and the number (or proportion) of magnetotactic responsive bacteria diffusing from the site of interest are observed, based on the known proportion, number, or concentration of magnetotactic responsive bacteria carrying the therapeutic agent, it is possible to estimate a similar proportion of magnetotactic responsive bacteria carrying the therapeutic agent remaining at the site of interest and the number of diffusing magnetotactic responsive bacteria carrying the therapeutic agent. For example, this information can provide an indication useful in assessing the toxicity of a therapeutic agent (based on the amount of therapeutic agent diffusing throughout the body, rather than delivery to the site of interest). For example, this information can also be used to provide an indication of the required dose of a therapeutic agent, where a certain proportion of magnetotactic responsive bacteria carrying the therapeutic agent (and therefore, the therapeutic agent carried) has failed to reach or remain at the site of interest.

[0119] It should be understood that solutions containing magnetotactic bacteria can be prepared according to methods known in the art for the purpose of preserving the magnetotactic bacteria and for further administration of the magnetotactic bacteria to subjects.

[0120] In some examples, the magnetotactic bacteria can be frozen before being conjugated to an imaging agent and / or a therapeutic agent. The thawed magnetotactic bacteria can be replicated and can be placed in the solutions described herein to conjugate the magnetotactic bacteria to a contrast agent, a therapeutic agent, or both.

[0121] An exemplary system for targeting aggregation zones using magnetotactic bacteria: For reference Figure 8 An exemplary system 800 is shown for implementing a method of targeting aggregation zones with magnetically oxidative bacteria by adjusting the magnetic field strength of a magnetic field according to the progress of magnetically oxidative bacteria in a subject.

[0122] System 800 includes processor 801 and memory 802.

[0123] System 800 communicates with one or more pairs (e.g., three pairs) of magnetic sources 804 (e.g., magnetic coils) and may include one or more pairs (e.g., three pairs) of magnetic sources, each pair of magnetic sources being aligned with one of three axes (x, y, z). System 800 communicates with imaging device 805 and may include imaging device.

[0124] System 800 may include a display 803. System 800 may include a user input interface 806.

[0125] Processor 801 may be a general-purpose programmable processor. In this example, processor 801 is shown as a single unit, but processor 801 may also be multi-core or distributed (e.g., multiprocessor).

[0126] Computer-readable storage 802 stores program instructions and data used by processor 801. Storage 802 may be non-transitory. Although shown as a single unit in this example for simplicity, computer-readable storage 802 may include multiple storage modules and / or caches. Specifically, it may include several storage layers, such as hard disks, external drives (e.g., SD card memory), and faster, smaller RAM modules. RAM modules may store data / program code currently, recently, or soon to be processed by processor 801, as well as cached data and / or program code from the hard disk. The hard disk may store program code and be accessed to retrieve such code for processor 801 to execute, and may be accessed by processor 801 to store imaging information of the subject, velocity values ​​of magneto-oxygen-responsive bacteria, volume of the injected drug bolus, program code operating the magnetic source 804, etc. Storage 802 may have a recycling architecture for storing, for example, imaging information of the subject, wherein old data is deleted when storage 802 is full or nearly full, or after old data files have been stored for a certain period of time.

[0127] The processor 801 and memory 802 can be connected via a bus connector.

[0128] User input interface 806 is used to allow a user to provide input to system 800 to interact with system 800, such as setting the magnetic field strength generated by magnetic source 804, marking gathering areas or 3D convergence points on display 803, etc. User input interface 806 may be a mouse, keyboard and / or controller, and is used to receive user input from the user.

[0129] It should be understood that other user input interfaces, such as touchscreens, joysticks, microphones, one or more proximity sensors that detect user movement, may also be used in accordance with this teaching.

[0130] In some examples, system 800 may not have user input interface 806, wherein system 800 may receive user input set on a remote computing device (e.g., remote computer, smartphone, tablet, laptop, etc.) through an input / output interface (such as a transceiver, not shown).

[0131] The display 803 provides a graphical user interface for controlling the system 800, and can display imaging information generated by the imaging device 805, parameters of the magnetic source 804, etc. to the user. The display 803 may include touch functionality, and thus also serve as a user input interface 806.

[0132] Magnetic source 804 may include one or more pairs of magnetic coils. Each pair of magnetic coils may be aligned along an axis. For example, magnetic source 804 may have three pairs of magnetic coils, wherein each pair of magnetic source 804 is aligned with one of three axes (x, y, and z). Magnetic source 804 may generate 3D convergence points (and convergence regions) as described herein or in U.S. Patent No. 9,905,347.

[0133] The imaging device 805 generates imaging information of the subject, such as defining aggregation areas and 3D convergence points, providing visual information about the subject's body (e.g., tumors), providing visual information about hypoxic areas of the tumor, and monitoring the movement of magneto-oxygen-responsive bacteria after injection into the subject. The imaging device 805 enables the imaging described herein. For example, the imaging device 805 can be a PET (positron emission tomography) machine, an MRI (magnetic resonance imaging) machine, a CT (computed tomography) machine, etc.

[0134] An exemplary method for targeting using magnetotactic bacteria: For reference Figure 7 An exemplary method 700 is shown to target aggregation areas in a subject’s body using magnetotactic bacteria by modulating the magnetic field strength of a magnetic field.

[0135] For ease of illustration, with reference to exemplary system 800, method 700 may be implemented by processor 801 executing program code stored in memory 802, the processor 801 executing instructions of the program code stored in memory 802. Processor 801 may adjust the power / current supplied to magnetic source 804 to adjust the magnetic field strength of the magnetic field generated by magnetic source 804.

[0136] The processor 801 can also control the imaging device 805 to acquire imaging information. In some embodiments (e.g., when the imaging device 805 is not part of the system 800), the imaging information can also be transmitted to the system 800 and the processor 801 via an input / output interface (I / O) (e.g., a transceiver).

[0137] In step 710, imaging information of the subject is acquired. Imaging information can be acquired through imaging methods as defined herein. The imaging information may provide information about the subject's target area, tissue, or organ (e.g., tumor, anatomical structure or part thereof of the subject, hypoxic area of ​​the tumor, etc.).

[0138] In some embodiments, imaging information can be enhanced by using magneto-oxygen-responsive bacteria that bind to imaging agents. The imaging agents can be captured by the imaging device, thereby enhancing the information collected through imaging.

[0139] In step 720, one or more puffs of magnetotactic responsive bacteria are injected into one or more sites on the subject. In some embodiments, the magnetotactic responsive bacteria may be injected into the peripheral region of a tumor. In some embodiments, the injection may be systemic. When a solution of magnetotactic responsive bacteria is injected into a patient, the puff has a given concentration of magnetotactic responsive bacteria and a given volume. The magnetotactic responsive bacteria may be conjugated to one or more therapeutic agents (e.g., anticancer agents), diagnostic agents, and / or imaging agents.

[0140] In step 730, a first mode is applied, which applies a magnetic field of sufficient strength to apply a directional torque to the magnetotactic bacteria, wherein the movement of the bacteria is primarily magnetic and run-and-flip (as the run phase increases and the recovery time decreases, the bacteria change direction less frequently from run-and-flip, thus realigning the direction of movement of the bacteria with the direction of the magnetic field). This first mode is used to displace the bacteria from their current location (e.g., the injection site) to a new location, such as a target area. In some embodiments, the first magnetic field strength may be at least 15 Gauss. However, those skilled in the art will understand that the first magnetic field strength value may vary depending on the characteristics of the magnetotactic bacteria (such as their sensitivity to the magnetic field). The magnetic field strength may be calculated from the volume of the injected bolus, the distance traveled by the magnetotactic bacteria (e.g., calculated from the known speed of the magnetotactic bacteria and the time after injection), the time after injection (as the bacterial residence time in the human body increases, its speed and responsiveness gradually decrease), the surface area of ​​the bolus, and / or the concentration of the magnetotactic bacteria in the bolus, and may depend on this.

[0141] Magnetic fields can guide magneto-oxygen-responsive bacteria in one to three dimensions by, for example, generating 3D convergence points as described herein.

[0142] Optionally, in step 740, information on the progress of the magnetically oxidative bacteria as they move within the subject's body can be generated (e.g., by monitoring the movement of the magnetically oxidative bacteria; estimating the location of the magnetically oxidative bacteria). For example, this can be monitored by combining the magnetically oxidative bacteria with an imaging agent that can be sensed by imaging. In some examples, monitoring can be performed by calculating the time after injection for the magnetically oxidative bacteria to reach the subject's body. The distance can be estimated from the pre-injection velocity distribution, post-injection velocity decay, and post-injection time. In some examples, the distance value can also be calculated from the volume and surface area of ​​the injected bolus.

[0143] In step 750, in the second mode, when the magnetotactic bacteria are observed or estimated to be approaching the target area (the target area has or is adjacent to an oxygen gradient generated by the hypoxic area at the target area), a magnetic field with a second magnetic field strength lower than the first magnetic field strength is applied. This reduction in magnetic field strength leads to a change in the movement pattern of the magnetotactic bacteria. Compared to when a higher first magnetic field strength is applied, the magnetotactic bacteria employ more run-tumble movements. This change in movement type to run-tumble allows the magnetotactic bacteria to seek out hypoxic areas of the tumor through oxygen attraction. In some examples, the second magnetic field strength may be below 15 Gauss but equal to or higher than 5 Gauss. However, those skilled in the art will understand that 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 from the volume, viscosity, shape, anisotropic orientation, distance traveled by the magnetotactic bacteria (e.g., calculated from the known speed of the magnetotactic bacteria and the time after injection), time after injection (as the bacteria remain in the human body for longer periods, their speed and responsiveness gradually decrease), and concentration of the magnetotactic bacteria in the drug bolus, and can depend on these factors.

[0144] In some embodiments, in step 760, in the third mode, once the magnetotactic bacteria reach the oxygen gradient in the hypoxic zone of the target area, the magnetic field strength can be reduced to below 5 Gauss up to 0 Gauss. Compared to step 750, this reduction in magnetic field strength further causes the magnetotactic bacteria to remain in the hypoxic zone of the target area, and the bacteria further change direction due to run-roll motion, thus deviating from the direction of the magnetic field. Since the magnetotactic bacteria can bind to one or more therapeutic agents, diagnostic agents, and / or imaging agents, once the post-injection velocity of the magnetotactic bacteria drops to 0 in the hypoxic zone (e.g., they die or weaken) (e.g., several minutes after their introduction into the subject), the therapeutic agent, diagnostic agent, and / or imaging agent is deposited in the subject's hypoxic zone. This achieves targeted administration of the compound to the subject. Furthermore, this method achieves specific targeting of the hypoxic zone of the tumor, where conventional administration of the compound to the subject has proven difficult due to the vascular and cellular structure of the tumor.

[0145] This method utilizes changes in the responsiveness and integrity of magnetotactic bacteria after administration to a subject. Because magnetotactic bacteria require colder temperatures and specific environmental conditions unavailable to the human body to survive, their introduction into the human body results in a shorter lifespan. Furthermore, the longer magnetotactic bacteria remain in the body, the slower they become, eventually leading to death. Therefore, it is advantageous to utilize the precision of their targeting during the period when they travel at a relatively high speed (post-injection). Thus, applying a high magnetic field strength to its maximum after injection (to avoid repolarization problems) benefits the magnetotactic response of the magnetotactic bacteria, allowing them to travel a greater distance to reach the oxygen gradient near the hypoxic zone of the target area when they exhibit optimal activity. Once the magnetotactic bacteria approach the oxygen gradient generated by the tumor, they slow down due to the harsh environment within the subject's body. However, this slowness is acceptable because magnetotactic bacteria may travel shorter distances by finding hypoxic areas in the tumor.

[0146] An exemplary method for controlling the spread of magneto-oxygen-responsive bacteria during targeted treatment: When performing targeting, for example, relative to the exemplary method 800, it may be desirable to reach more hypoxic areas with each injection while reducing the bacterial concentration in each hypoxic area at the target region; or to achieve concentrated targeting of a few hypoxic areas using a higher bacterial concentration. Adjusting the target volume at the target region depending on the expected outcome may be advantageous, where targeting with more bacterial diffusion will result in a larger target volume at the target region, while targeting with less bacterial diffusion (more concentrated) will result in a smaller target volume at the target region.

[0147] To increase or decrease the target volume at the point where injected bacteria reach the target area, this can be achieved by preparing or selecting bacterial samples based on the sample distribution rate. (Reference) Figure 9A An exemplary large target volume 900A with multiple hypoxic zones 910 at the target region is shown. Figure 9B A smaller target volume 900B with less hypoxia at the target region is shown. It should be understood that example target volumes 900A and 900B are simplified for illustrative purposes, as are the hypoxia regions, and they can take on various shapes depending on the subject's anatomy and the targeted organ, tissue, or tumor. Target volumes 900A and 900B are used to illustrate the order of magnitude difference between target volumes. For target volume 900A, a bacterial sample with a larger velocity distribution can be selected. For target volume 900B, a bacterial sample with a narrower velocity distribution can be selected.

[0148] A larger velocity distribution will result in a larger target volume or diffusion. A smaller velocity distribution will result in a smaller target volume or diffusion.

[0149] Assessing this velocity distribution (e.g., the standard deviation of the velocity of bacteria in a sample) allows users to select samples based on the desired target volume at the target region.

[0150] Furthermore, the pre-injection velocity of bacteria in the sample can be compared to a velocity threshold for overcoming one or more hypoxic zones near the injection site. Bacteria with velocities above the threshold will travel through these hypoxic zones, while bacteria with velocities below the threshold will remain in these hypoxic zones. Therefore, the pre-injection velocity of bacteria itself (in addition to its velocity distribution) can also provide information about the post-injection movement pattern of bacteria.

[0151] The velocity distribution of a sample before injection into the patient can be assessed (and in some cases, if the bacterial sample manufacturer does not provide this data). If the velocity distribution of the sample does not meet the requirements for achieving the desired target volume at the target site, the sample can be discarded or replaced with another sample that has a more desirable velocity distribution.

[0152] Furthermore, samples can be prepared based on the desired diffusion or target volume at the target region. For example, multiple bacterial cultures can be cultured, each associated with a given rate or rate range (e.g., "slow," "medium," "fast"). If a small rate distribution is desired, a sample can be prepared from a culture with similar rates. However, if a large rate distribution is required, samples can be prepared from multiple cultures, each with different rate levels, thus including slower and faster bacteria (increasing the overall rate distribution of the prepared sample). It should be understood that a rate range can also be selected from the same culture.

[0153] In other embodiments, the user can focus on post-injection velocity decay to select samples. For example, some bacteria may be more resistant to the environment within the subject's body, and therefore require a longer time to decrease in velocity. In other cases, bacteria may be less resistant, and therefore decrease in velocity rapidly after injection. Using known curves of expected post-injection velocity decay, the user selects samples with a larger range of velocity variation after injection when the target volume is large (even if the velocity distribution is relatively narrow), and samples with a smaller range of velocity variation after injection when the target volume is small. Once the bacteria are administered to the subject, if the sample exhibits a large range of velocity variation after injection, the change in velocity variation after injection will manifest as a broadening of the velocity distribution over time, thereby increasing the distance the bacteria travel by adjusting the magnetic field strength to magnetotactic and through run-and-reverse movement.

[0154] An exemplary method for improving targeting of magneto-oxygen-responsive bacteria by adjusting the total drug escape time: Total drug escape time refers to the total time it takes for the injected magnetic-oxygen-responsive bacteria drug pulsar to completely dissolve, that is, the total time required for all motile magnetic-oxygen-responsive bacteria to escape the injected drug pulsar.

[0155] Total pouch escape time can be a factor in improving targeting of specific regions within a patient's body using magnetotactic oxytocin (MOO)-responsive bacteria. Improved targeting can be achieved when total pouch escape time is considered in conjunction with the post-injection velocity of MOO in the patient's body (or the velocity of bacteria after being placed in an environment detrimental to their motility, such as the human body). More specifically, the longer it takes for MOO to leave the pouch (at the injection site), the longer the MOO remains in an environment that reduces bacterial motility (and velocity). Therefore, the longer the escape time for MOO, the slower the MOO is, as its velocity is at least partially attenuated by this delay.

[0156] Injection can be performed within the target area (e.g., inside the tumor) or around the target area (e.g., around the tumor).

[0157] Therefore, a shorter total drug escape time allows for a larger target volume at a greater distance from the injection site. This is because the injected magnetotactic bacteria have a high velocity upon injection into the patient and upon escaping the drug bolus, and thus can travel a greater distance post-injection. Maintaining a short total drug escape time (high post-injection velocity of the bacteria) can be used for targeting areas within the patient body that are difficult to access with a needle or injection device. For example, if the injection device cannot reach one side of a tumor or organ, the magnetotactic bacteria can be administered at a location accessible to the injection device (but at a distance from the target location). The magnetic field then guides the magnetotactic bacteria to the target location, where their post-injection velocity is sufficient to propagate them.

[0158] Conversely, when the target area is near the injection point, a longer total drug escape time is preferable, which results in at least a partial reduction in the post-injection velocity of the magnetotactic bacteria. In these embodiments, it is preferable that the magnetotactic bacteria remain near the injection point after injection. Therefore, a reduction in post-injection velocity is preferred, wherein the displacement of the magnetotactic bacteria is limited after injection. The magnetotactic bacteria will remain near the injection point, the drug bolus, and the target area.

[0159] In some examples, reducing the rate of magnetotactic bacteria before injection can increase the total drug escape time (e.g., in the injection device), such as heating the solution to above room temperature (but preferably below body temperature) before injection into the subject, or exposing the solution to ultraviolet light, can increase the total drug escape time.

[0160] It should be understood that when using the location of magnetotactic bacteria in a patient for targeting, adjusting or taking into account the total drug escape time can be used alone or in combination with any other parameters and / or techniques described herein, such as modulating the magnetic field strength, selecting magnetotactic bacteria samples with specific velocity distribution profiles, etc.

[0161] In the preparation of a magnetically oxygen-responsive bacterial solution for administration to a patient (which can be done by the medical personnel administering the injection or by the manufacturer of the magnetically oxygen-responsive bacterial solution), the total drug escape time can be influenced, whereby the properties of the drug can be selected to affect the total drug escape time.

[0162] For example, such properties include, but are not limited to, pharmacoplasma concentration and pharmacoplasma viscosity (which may be related to its concentration).

[0163] During injection (e.g., when a physician administers a magnetotactic responsive bacterial solution to a patient), other properties that may affect the total drug escape time can be adjusted, such as the drug's surface area to volume ratio, the surface area of ​​the drug exposed to a north-facing magnetic field for north-seeking magnetotactic responsive bacteria, and the drug's volume for south-seeking magnetotactic responsive bacteria exposed to a south-facing magnetic field. These properties can be affected by the injection rate (injection volume varying over time), for example, whether the syringe is withdrawn from the initial injection point while the magnetotactic responsive bacteria are being continuously injected into the body.

[0164] Therefore, the total drug escape time can also be modified by delaying the injection of the drug bolus into the patient (e.g., a solution containing magnetotactic bacteria is retained in the injection device for a specific period of time). The choice of syringe type used for injection can also affect the properties of the drug bolus (e.g., the shape of the bolus, the surface area of ​​the bolus, etc.). Exemplary syringes include, but are not limited to, multi-hole syringes, single-hole syringes, multi-side-hole syringes, etc.

[0165] The concentration of magneto-oxygen-responsive bacteria in the solution used for injection can affect the solution viscosity, thereby influencing the total drug escape time. Higher concentrations result in higher viscosity, and vice versa. Higher concentrations lead to a longer total drug escape time due to increased viscosity, while lower concentrations lead to a shorter total drug escape time due to decreased viscosity.

[0166] The viscosity of the bacterial solution (and therefore the total flotation escape time) can also be affected by one or more other components added to the flotation (e.g., the nature of the flotation excipient).

[0167] The surface area to volume ratio of a drug bolus can also affect the total drug escape time, with a larger ratio resulting in a shorter escape time. Conversely, a smaller ratio results in a longer escape time. The surface area to volume ratio can be adjusted by changing the shape of the bolus (e.g., spheres versus cylinders) or by dividing the volume of a single bolus into smaller boluses (which are then injected independently into the patient).

[0168] In some implementations, the ratio of the surface area of ​​the drug floc facing the magnetic north pole for north-seeking magnetotactic-responsive bacteria and facing the magnetic south pole for south-seeking magnetotactic-responsive bacteria also affects the total drug floc escape time. A larger surface area ratio facing the magnetic north pole for north-seeking magnetotactic-responsive bacteria and facing the magnetic south pole for south-seeking magnetotactic-responsive bacteria will promote the escape of magnetotactic-responsive bacteria and thus reduce the total drug floc escape time. Conversely, a smaller surface area ratio facing the magnetic north pole for north-seeking magnetotactic-responsive bacteria and facing the magnetic south pole for south-seeking magnetotactic-responsive bacteria will hinder the escape and prolong the total drug floc escape time.

[0169] It should be understood that magnetotactic bacteria can be polar or axial: polar magnetotactic bacteria can be north-seeking or south-seeking, while axial magnetotactic bacteria can be both south-seeking and north-seeking. Axial magnetotactic bacteria can move along a north-south axis and, for example, change direction along that axis when encountering an obstacle.

[0170] When the injection device can only dispense a fixed volume over time, a larger volume can lead to a longer total drug flotation time. Therefore, reducing the volume can reduce the total drug flotation time. However, reducing the volume leads to an increase in concentration and thus an increase in viscosity, which can also prolong the total drug flotation time.

[0171] Total drug escape time can also be adjusted by reducing the injection rate to the patient (e.g., varying the volume injected into the patient over time). A reduced injection rate will result in a prolonged total drug escape time. Conversely, an increased injection rate will reduce the total drug escape time.

[0172] It should be understood that the properties of the drug mass that can affect the total drug mass escape time described in this article are merely exemplary, and other properties of the drug mass may also affect the total drug mass escape time.

[0173] Certain drug particle properties that can affect the overall drug particle escape time can be adjusted by the manufacturer of the magnetically oxygen-responsive bacterial solution (e.g., in vials or injection devices) intended for injection into patients. These properties include, but are not limited to, drug particle concentration and viscosity.

[0174] Certain properties of the drug bolus can be modified by the physician administering the bolus. These properties include, but are not limited to, the surface area to volume ratio, the volume of the injected bolus, and the shape of the injected bolus. These properties of the bolus can be affected by the injection technique used by the physician or the tools used by the physician performing the injection (e.g., the type of syringe). For example, a larger bolus volume can be achieved by gradually moving the tip of the syringe during injection. Thus, by not injecting the magnetotactic bacteria at the same point in the subject's body, the volume of magnetotactic bacteria injected into the patient can be increased. Depending on the orientation, displacement, rotation, etc., of the syringe during injection, these movements can also affect the shape of the bolus and thus the total bolus escape time. In some embodiments, the characteristics of the tissue at the injection site (e.g., density, shape, etc.) can also affect the shape of the bolus, where the injected solution may form sacs within a denser tissue capsule. Therefore, environmental tissue can also be used to adjust the shape of the bolus and thus also adjust the total bolus escape time.

[0175] It should also be understood that physicians may delay injecting the drug bolus into the patient, thereby prolonging the total drug bolus escape time.

[0176] Exemplary study: The following exemplary studies are provided to enable those skilled in the art to better understand this disclosure. Because they are merely illustrative and representative examples, they should not be used to limit the scope of this disclosure, but are intended for illustrative and representative purposes only. It should be understood that other exemplary studies may be used to further illustrate and represent this disclosure without departing from these teachings.

[0177] Tumor regions with hypoxic conditions, known as hypoxic zones (HZs), cause tumor cells to develop resistance to multiple treatment modalities, such as chemotherapy and radiotherapy, while exhibiting a more aggressive and malignant phenotype, leading to poor prognosis for patients. For example, the low oxygen levels in hypoxic zones reduce the effectiveness of radiotherapy, while chemotherapy agents are primarily unable to penetrate these hypoxic zones due to limited diffusion. Therefore, targeting hypoxic zones using specialized propulsion carriers (referred to in this paper as transpotherapeutic transporters, or transpotherapeutic complexes, TC) capable of delivering therapeutic drugs or other medications to hypoxic zones can significantly improve treatment efficacy.

[0178] The specific magnetotactic responsive bacteria described in this article are well-suited for use as transport therapeutic vectors to target and transport payloads to hypoxic regions. These magnetotactic responsive bacterial cells are typically microaerophilic or similar types that seek oxygen levels similar to those found in hypoxic tumor regions. Since tumor hypoxia typically occurs at oxygen levels <1% and is usually limited to a duration of <0.1% oxygen (7.5 to <0.75 mmHg) (Br J Radiol. March 2014; 87(1035): 20130676), the target oxygen level (TOL) is defined here as the oxygen level targeted by a transport therapy vehicle or transport therapy complex operating in Aerotactic Targeting Mode (ATM). That is, the directional displacement of the transport therapy complex is only affected by the oxygen gradient and not by other sources (such as directional magnetic fields that can affect its directional movement). They may be suited to or close enough to the hypoxic area to produce a suitable targeted therapeutic effect that is superior to that achieved by other conventional treatments such as radiotherapy and chemotherapy (taking only the two major therapies as examples).

[0179] Magneto-oxygen transport therapy vehicles or transport therapy complexes are typically guided to hypoxic regions using various magnetic field configurations (MFCs). A sequence of magnetic field configurations, or magnetic field configuration sequences (MFCSs), forms a Transpotherapeutic Targeting Sequence Event (TTSE, or simply Event) that uses one of three magnetic tactic targeting modes (MTMs) or derivatives thereof: Direct Magnetic Targeting (DMT), Spatially-limited Magnetotactic Targeting (SMT), and Aggregation Zone Targeting (AZT). This results in a specific displacement path (DP) for the transport therapy complex, while all desired transport therapy targeting sequence events in a sequence lead to a TTS (Targeting Sequence). In these magnetic field configurations, the transport therapy complex is acted upon by a directional vector (DV) that causes the transport therapy complex to follow a specific displacement path. Any directional vector at a given moment consists of two components: the amplitude and direction of the magnetic field configuration applied to the transport therapy complex. For the magnetic field configuration, the amplitude is the magnetic field strength. Magneto-aerotactic targeting methods (MATM) using the magnetic field strength in the directional vector are employed to adjust the amplitude of the magnetic field configuration, taking into account certain key parameters listed in subsequent chapters, to achieve precise targeting of the hypoxic region, considering the hypoxic zone and its related effects on the transport therapy complex's motion behavior.

[0180] Main parameters Magnetic field intensity range (FIR) Magneto-oxygen-responsive bacterial transport therapy vehicles suitable for targeting hypoxic regions are typically motile magneto-oxygen-responsive bacteria, meaning they can operate in magnetotactic mode only, oxygen-responsive mode only, or both simultaneously. The appropriate displacement or targeting mode of the magneto-oxygen-responsive bacterial transport therapy vehicle is selected by exposing the transport therapy vehicle or transport therapy complex to a specific range of directional magnetic flux densities. Magnetic field strength ranges in ascending order of density (referred to herein as oxygen-attracting magnetic field strength ranges, magneto-oxygen-responsive magnetic field strength ranges, and magnetotactic magnetic field strength ranges) are thus defined for oxygen-responsive, magneto-oxygen-responsive, and magnetotactic displacement or targeting modes, respectively. While these ranges vary depending on the transport therapy complex, specific physiological environments, and other factors, a typical magneto-oxygen-responsive magnetic field strength range can be approximately 5 to 15 Gauss. The lower and upper limits of the magneto-oxygen-responsive magnetic field strength range are defined as follows: the lower limit extends to 0 Gauss, while the upper limit is limited by the potential repolarization (RPL) of the transport therapy vehicle, which will reverse the expected direction of the transport therapy vehicle's displacement at higher magnetic field strengths.

[0181] Magneto-oxygen-responsive bacteria, used as therapeutic transport carriers, employ a combination of run-tumble and run-inverted movements for oxygen attraction. The magnetic moment of the magnetosome chains containing magnetic particles in the cells of these bacteria is 10⁻⁶. -15 Am 2 A relatively weak directional magnetic field can induce a magnetic torque that aligns the movement of magneto-oxygen-responsive bacterial cells with the direction of the magnetic field. However, tumbling disturbs this alignment, and therefore run-tumble motion manifests as a disturbance of this alignment caused by tumbling, followed by a return to alignment with the directional magnetic field during the run phase. If the run phase is shorter than the recovery time, the magnetic field does not provide sufficient time for the magneto-oxygen-responsive bacterial cells to realign before the next tumble causes the movement of the cells to deviate from the magnetic field direction. The viscosity of the medium and the geometry of the transport therapy complex are only two factors that can affect the recovery time. For example, since the coefficient of rotational friction is proportional to the viscosity (η) of the medium, a larger induced torque is required to rotate the cells, and therefore the distribution of tumble angles is expected to be biased towards smaller angles. Increasing the strength of the directional magnetic field increases the magnetic torque induced on the magnetosome chain and thus reduces the recovery time, thereby reducing the amplitude of deviations from the magnetic field direction.

[0182] Within the range of magnetotactic magnetic field strength, the directional displacement of the transport therapy complex is still influenced by the directional magnetic field, but the recovery time is longer, especially at lower magnetic field strength levels, leading to increased fluctuations in the direction of movement deviating from the magnetic field direction. This increased fluctuation helps the transport therapy complex to remain in the tumor hypoxic zone, at least temporarily, within the range of magnetotactic magnetic field strength when crossing the hypoxic zone, and rises and exceeds the total transport therapy complex motion time (MT) when the magnetic field strength decreases to the range of oxytactic magnetic field strength (i.e., from 0 to the minimum magnetic field strength within the range of magnetotactic magnetic field strength). However, when the directional magnetic field increases to the range of magnetotactic magnetic field strength, the effect of the tumbling phase is significantly reduced, and during the running phase, the transport therapy complex moves along the magnetic axis with the direction of the magnetic field, without a significant stationary phase caused by tumbling in the hypoxic zone. When the running phase is followed by the reversal phase (in which the transport therapeutic complex moves backward along the magnetic field axis while maintaining the polar magnetotactic-oxygen-responsive bacterial cell head facing the magnetic field direction), the forward swimming speed of the polar magnetotactic-oxygen-responsive bacteria is faster (more efficient) than the backward movement because the propulsion force or flagella are located only on one side of the polar magnetotactic-oxygen-responsive bacterial cell (rather than on both sides as in axial magnetotactic-oxygen-responsive bacteria). The net displacement of the North Seeking (NS) magnetotactic-oxygen-responsive bacteria is towards the North Pole of the magnetic field. (Note that, if needed, the magnetotactic-oxygen-responsive bacteria can repolarize to the South Seeking (SS) type, and vice versa.)

[0183] Therefore, when the transport therapy complex is exposed to a directional magnetic field configured within the range of magnetotactic magnetic field strength, it will exhibit a run-tumble oxygen-seeking movement that is limited by the large directional magnetic torque induced on the magnetosome chains within the magnetotactic-oxygen-responsive bacterial cells. Thus, the run-tumble strategy is most likely to dominate when exposed to a directional field within the range of magnetotactic magnetic field strength. Magnetotactic-oxygen-responsive bacterial cells typically exhibit long, unidirectional run movements interrupted by short reversal events at a specific average non-periodic frequency, resulting in forward displacement.

[0184] When the transport therapy complex operates within the oxygen-attracting magnetic field strength range, it will perform run-roll and run-reverse movements within the target oxygen level region to maintain its position at that oxygen concentration, thus forming a transport therapy complex band diffused within or near the target hypoxic zone. The volume occupied by the transport therapy complex within or near the hypoxic zone corresponds to the OATZ (Oxic-Anoxic Transition Zone). The duration for which the transport therapy complex remains within the target oxygen level region (OATZ), such as a tumor hypoxic zone, is called the TOLRD (Target Oxygen Level Retention Duration). TOLRD is the average or median duration for which the transport therapy complex maintains its position within the target oxygen level region (such as a tumor hypoxic zone). For the oxygen-attracting magnetic field strength range, a target oxygen level retention duration of 1.0 means that the transport therapy complex will remain within the target oxygen level region after reaching it until the post-injection velocity = 0 (the post-injection velocity is defined in the next section). For the range of magnetotactic magnetic field strength, a target oxygen level retention duration of 0 means that the transport therapy complex will pass through the target oxygen level region without significant lingering (manifested as no significant tumbling, only brief reversal motion, and a longer running phase). For the range of magnetotactic magnetic field strength, the target oxygen level retention duration ranges from slightly above 0 to slightly below 1.0, corresponding to the lower limit (MIN-MAFIR) and upper limit (MAX-MAFIR) of the magnetotactic magnetic field strength range.

[0185] Velocity Post-Injection (VPI) Post-injection velocity (VPI) typically declines upon exposure to physiological environmental factors that reduce the motility of the magneto-oxygen-responsive bacterial transport therapeutic complex. A physiological temperature of 37 °C is a potential contributing factor. To compensate for the effects of specific physiological or tumor microenvironment conditions, the VPI obtained at a specific time post-injection can be further adjusted by multiplying by an adjustment factor. The VPI can be expressed using an equation (e.g., VPI = 0.4 (0.1 t)). 2 -8.1 t + 188), where t is the time after injection in minutes, and the unit of the post-injection velocity is µm. s-1The velocity after injection is represented in the form of graphs, tables, etc., and is the translational velocity, calculated as the straight-line distance traveled between two points. Since magneto-oxygen-responsive bacterial cells are not usually passively pushed or pulled by flagella located on one side of the cell, but rather the cell itself moves in a helical motion due to the angle between the longitudinal axis of the magnetosome chain and the axis of the flagellar propulsion structure, thus contributing to the displacement efficiency and further increasing the displacement velocity through the fluid environment, the actual or instantaneous velocity of the transport therapeutic carrier or transport therapeutic complex will be related to the total helical distance. Therefore, the instantaneous (lateral or rotational) velocity is different from the translational (longitudinal) velocity; the latter (translational velocity) is what is actually considered when estimating the total distance traveled.

[0186] Velocity Transpotherapeutic Complex Standard Deviation (VTCSD) The velocity of the transpotherapeutic complexes (VTC) is defined here as the velocity at t = 0 when the post-injection velocity equals the velocity of the transpotherapeutic complexes, where the velocity of the transpotherapeutic complexes is the velocity recorded before transport from the manufacturing site, and VDI (Velocity During Injection) is the velocity during injection. The standard deviation (SD) of the velocity of the transpotherapeutic complexes, referred to here as VTCSD, is a measure of the change in the velocity of the transpotherapeutic complexes. A low standard deviation of the transpotherapeutic complex velocity indicates that the velocity values ​​of the transpotherapeutic complexes in the injected transpotherapeutic complexes tend to approach the average velocity of the ensemble of transpotherapeutic complexes, while a high standard deviation of the transpotherapeutic complex velocity indicates a wider distribution of the velocity of the transpotherapeutic complexes.

[0187] Total Bolus Escape Time (TBET) Total plasma escape time (TPS) refers to the total dissolution time of an injected transpotherapeutic complex plasma, i.e., the total time required for all motile transpotherapeutic complexes to escape from the injected plasma. For a given plasma, TPS increases with increasing transpotherapeutic complex concentration (TCC), leading to a decrease in the injection volume (VOL) per given therapeutic dose (especially for larger volumes) and an increase in plasma viscosity, which improves retention within tissues. For a given transpotherapeutic complex, TPS also depends on the plasma escape surface (BES). The plasma escape surface is defined as the north-facing surface of the injected transpotherapeutic complex plasma oriented in a north-seeking transpotherapeutic complex configuration with respect to the applied magnetic field. The transpotherapeutic complexes on the plasma escape surface are typically the first to escape from the plasma. Depending on the plasma geometry, the plasma escape surface may remain constant or vary with decreasing plasma volume. Therefore, the escape surface of the injection bolus can be adjusted by using various injection techniques to properly shape the injection bolus.

[0188] Major Magneto-Aerotactic Targeting Methods (MATM) Magneto-Aerotactic targeting methods can be single magneto-aerotactic targeting methods (SMATM) or combined magneto-aerotactic targeting methods (CMATM). Single magneto-aerotactic targeting methods include: magnetic field strength range-hypoxic region targeting, post-injection velocity-hypoxic region targeting, standard deviation of transport-therapeutic complex velocity-hypoxic region targeting, and total drug escape time-hypoxic region targeting. Combined magneto-aerotactic targeting methods combine two or more single magneto-aerotactic targeting methods.

[0189] Magnetic field strength range - Targeting in hypoxic areas The magnetic field strength range-anoxic zone targeting method adjusts the magnetic field strength range along the displacement path generated by the magnetotactic targeting mode or its derivative mode to target the anoxic zone. Figure 3 A simple example is shown. Figure 3 The target area is hypoxic region 2. The drug is applied from the injection (INJ) site. Figure 3The arrows in the diagram indicate the directional magnetic field configuration. To target hypoxic region 2, hypoxic region 1 is bypassed by first applying a directional magnetic field configuration within the range of magnetotactic magnetic field strength, and then applying a directional magnetic field configuration within the range of oxygenattractant magnetic field strength when the transport therapy complex is sufficiently close to or within hypoxic region 2.

[0190] While magnetic field strength range-hypoxia targeting offers good control over the selection of the target hypoxic region, it still requires adequate information about the distribution and location of the hypoxic region, typically obtained and estimated beforehand from suitable medical imaging modalities, as well as a relatively accurate estimate of the distance the transport therapy complex travels from the injection site to the target hypoxic region. To fully utilize magnetic field strength range-hypoxia targeting, the standard deviation of the transport therapy complex velocity should be relatively low, and the mean should be sufficiently high to provide adequate HBPIT (Hypoxic Bypass Post Injection Time). HBPIT is defined here as the maximum elapsed time after injection, at which point the transport therapy complex can escape the hypoxic region while traveling within the magnetotactic magnetic field strength range, primarily due to the decay (degradation) of the velocity after injection (and thus the shortening of the distance traveled to the hypoxic region), especially in magnetotactic bacteria suitable for use as hypoxic targeting transport therapy complexes. This, in turn, limits the time during which magnetic field strength range-hypoxia targeting can be applied and the total distance from the injection site to the bypassable hypoxic region, although this distance can be relatively large due to the post-injection velocity. It should also be noted that the magnetic field strength range-hypoxia zone targeting can also be used to target different hypoxia zones with different concentrations of transport therapy complexes, for example in... Figure 3 The example shown allows for targeting hypoxic regions 2 and 3 with different ratios of transport therapy complexes, wherein a magnetotactic magnetic field strength range is set for hypoxic region 1, a magnetotactic-oxygenatactic magnetic field strength range is set for hypoxic region 2, and an oxygenatactic magnetic field strength range is set for hypoxic region 3. This is only a simple example, and other scenarios are possible.

[0191] Post-injection speed - targeted to hypoxic areas Velocity Post Injection – Hypoxic Zone Targeting (VPI-HZT) methods typically operate within the range of magnetotactic magnetic field strength (although the magnetotactic-oxygen-attracting magnetic field strength range can also be used), but rely on post-injection velocity decay (post-injection velocity decline) to target hypoxic zones. This is in Figure 4 The diagram is simply shown, where t is the time elapsed after injection.

[0192] exist Figure 4In the simplified example shown, the transport therapy complex has a sufficiently high post-injection velocity to bypass the hypoxic region 1 with a directional magnetic field configuration within the range of magnetotactic magnetic field strength. As the transport therapy complex continues to travel along the displacement path derived from the magnetic field configuration, the decaying post-injection velocity continuously decreases until it reaches a value that is no longer sufficient to traverse the hypoxic region. At this stage, the run-state distance derived from the decaying post-injection velocity is insufficient to avoid the oxygen gradient associated with the hypoxic region. Therefore, the transport therapy complex remains stationary. Figure 4 The hypoxic zone 2 in the example shown represents a specific hypoxic zone.

[0193] When targeting specific hypoxic regions, post-injection velocity-hypoxic zone targeting requires accurate estimation of the post-injection velocity after decay and the corresponding distance traveled along the displacement path. Because the post-injection velocity-hypoxic zone targeting method is highly dependent on the post-injection velocity after decay, the range of control that can be applied by the operator is limited. Furthermore, the targeting range from the injection site is limited and determined by the decaying post-injection velocity.

[0194] Standard deviation of transport therapy complex velocity - hypoxic zone targeting The Velocity Transpotherapeutic Complex Standard Deviation – Hypoxic Zone Targeting (VTCSD-HZT) method utilizes the velocity distribution of the transport therapy complex and, therefore, the post-injection velocity change within the injection volume to target hypoxic zones. The basic principle of the VTCSD-HZT method is illustrated in [the diagram / example]. Figure 5 and Figure 6 In, and primarily depends on the standard deviation of the transport-therapeutic complex velocity, the average velocity of the transport-therapeutic complex during injection, and the factors arising from... Figure 5 and Figure 6 The left arrow in the figure indicates the rate of decay during the injection.

[0195] Figure 5An example of the standard deviation of the transport therapy complex velocity for a batch of transport therapy complexes measured at the factory is shown. In this example, only the fastest transport therapy complex is selected by rejecting the slowest one. Assuming the velocity during injection (which is the post-injection velocity at t = 0) is equal to the velocity of the transport therapy complex, if the transport therapy complex operates within the range of magnetotactic magnetic field strength, the first hypoxic zone near the injection site may be bypassed by this transport therapy complex. This is because the velocity during injection and the post-injection velocity shortly after injection for all transport therapy complexes are faster than the velocity threshold during injection within the range of magnetotactic magnetic field strength (defined here as the minimum velocity during injection required to bypass the hypoxic zone). As time progresses after injection, the post-injection velocity decreases while maintaining the range of magnetotactic magnetic field strength, which helps in targeting the hypoxic zone. The rate and distribution of targeted hypoxic zones along the displacement path in the hypoxic zone will depend on the standard deviation of the transport therapy complex velocity. A high standard deviation of the transport therapy complex velocity will increase the diffusion of magneto-oxygen-responsive bacteria across the hypoxic zone along the displacement path and result in a lower transport therapy complex density per hypoxic zone. Conversely, a low standard deviation of the transport therapy complex velocity will result in a more localized (less diffused) hypoxic zone but with a higher transport therapy complex density per hypoxic zone.

[0196] Figure 6 The diagram illustrates a velocity threshold during injection within a magnetotactic magnetic field strength range, located at the average injection velocity of the transport therapy complex. This allows for targeting of hypoxic regions closer to the injection site while maintaining the magnetotactic magnetic field strength range, and also enables targeting of other hypoxic regions farther from the injection site. Therefore, the same transport therapy complex batch can be designed with different percentages to the left or right of the velocity threshold during injection within the magnetotactic magnetic field strength range of the transport therapy complex velocity standard deviation to optimize hypoxic region targeting relative to the distribution and location of hypoxic regions within the tumor volume. This approach forms the basis of the transport therapy complex velocity standard deviation-hypoxic region targeting method. Generally, to target numerous hypoxic regions that may be geographically dispersed, a high transport therapy complex velocity standard deviation, indicating a wider velocity range distribution, may be more suitable when relying solely on the transport therapy complex velocity standard deviation-hypoxic region targeting method.

[0197] Total drug escape time - hypoxia zone targeting The Total Bolus Escape Time – Hypoxic Zone Targeting (TBET-HZT) approach relies on a combination of the dissolution rate of the injected transport therapeutic complex drug particle and post-injection velocity decay to achieve hypoxic zone targeting within the tumor volume using a directional vector that typically operates within a range of magnetotactic magnetic field strengths (although a magnetotactic-oxygen-attracting magnetic field strength range can also be used). Considering the surface area to volume (S / V) ratio of a spherical injected drug particle, where surface area = 12.57r... 2 And the volume = 4.189 r 3 Let r be the radius of the spherical drug delivery system. We can conclude that dividing a single injection drug delivery system into smaller ones results in a larger effective drug delivery escape surface, thus shortening the total drug delivery escape time. The same principle can be applied: modifying the volume shape of the drug delivery system (e.g., orienting the longitudinal axis of a cylindrical drug delivery system perpendicular to the direction vector) will result in a larger surface area to volume ratio, leading to a shorter total drug delivery escape time. Typically, when a magnetic field configuration within the range of magnetotactic magnetic field strength is applied, the total drug delivery escape time is equal to or shorter than the velocity threshold during injection within that range to target hypoxic areas in the tumor volume outside the injection site. This is because a total drug delivery escape time longer than the velocity threshold during injection within the range of magnetotactic magnetic field strength will result in a certain percentage of the transport therapy complex remaining at the injection site, although this may be appropriate in specific circumstances. Besides the drug delivery escape surface, other factors can also influence and be used to adjust the total drug delivery escape time; transport therapy complex concentration is one example.

[0198] For the transport therapy complex located on the escape surface of the drug bolus that escapes from the drug bolus and travels toward the hypoxic zone, the transport therapy complex within the drug bolus is also affected by physiological environmental factors that reduce the mobility of the transport therapy complex. This results in a slower injection rate during which the transport therapy complex that escapes from the drug bolus decays over time after injection, making the injection rate during which the last transport therapy complex to escape from the drug bolus is lower than the injection rate during which the first transport therapy complex to escape from the drug bolus is injected.

[0199] If the total puff escape time equals the motion time, where the direction vector, within the magnetotactic magnetic field strength range, has an average injection velocity significantly higher than the injection velocity threshold within that range, and the standard deviation of the transport therapy complex velocity is very low, then we can expect that: the first transtherapeutic complex to escape from the injected puff will bypass the first hypoxic zone, and when the post-injection velocity decays to below the injection velocity threshold within the magnetotactic magnetic field strength range, the target hypoxic zone is farther from the injection site. Subsequently, the transtherapeutic complex escaping from the injected puff will reach a hypoxic zone closer to the injection site, thus bringing the target hypoxic zone closer to the injection site. Finally, the transtherapeutic complex escaping from the injected puff will not have sufficient post-injection velocity to escape from the hypoxic zone closest to the injection site, resulting in the target hypoxic zone also being targeted. If the total puff escape time is shortened further, targeting hypoxic zones closer to the injection site may be impossible, while targeting hypoxic zones farther from the injection site may be increased.

[0200] Combined magnetic-oxygen targeting method Combined Magneto-Aerotactic Targeting Methods (CMATM) combines two or more individual magneto-aerotactic targeting methods described in previous chapters.

[0201] Although the invention has been described with reference to preferred embodiments, it should be understood that various modifications can be made by those skilled in the art. Such modifications and variations are considered to be within the scope and rights of the invention.

[0202] The foregoing description, taken in conjunction with the accompanying drawings, provides a representative, non-limiting example of the invention. This detailed description is intended only to teach those skilled in the art further details of practicing preferred aspects of this teaching, and is not intended to limit the scope of the invention. Furthermore, each additional feature and teaching disclosed in the context may be used alone or in combination with other features and teachings.

[0203] Furthermore, the combination of features and steps disclosed in the detailed description and experimental examples above is not essential for practicing the invention in the broadest sense, but is instead taught only to specifically describe representative examples of the invention. Moreover, various features of the above representative examples, as well as the following independent and dependent claims, can be combined in ways not specifically and expressly enumerated to provide additional useful embodiments of this teaching.

Claims

1. A method for improving the targeted delivery of at least one of a therapeutic agent, an imaging agent, and a diagnostic agent attached to magnetotactic-responsive bacteria, said magnetotactic-responsive bacteria being adapted to move spontaneously in the presence of a magnetic field, said magnetotactic-responsive bacteria causing at least one of the therapeutic agent, the imaging agent, and the diagnostic agent to deposit in a hypoxic area within a patient, said method comprising: The total drug escape time of the solution of the magnetotactic oxytocinoids when injected into the patient is adjusted to affect targeting of target areas with hypoxia in the patient's body. Shortening the total drug escape time facilitates the targeting of the magnetotactic oxytocinoids to one or more hypoxia areas far from the injection site of the magnetotactic oxytocinoids, while taking into account the reduction in the post-injection velocity of the magnetotactic oxytocinoids when exposed to the patient's environment.

2. The method according to claim 1, wherein, The total pneumatic mass escape time is adjusted by adjusting one or more of the following items: The concentration of the magneto-oxygen-responsive bacteria in the solution; The volume of the drug pellet; The viscosity of the solution; as well as When the magnetotactic bacteria are north-seeking, the drug cluster is exposed to a surface facing north of the magnetic field; or when the magnetotactic bacteria are south-seeking, the drug cluster is exposed to a surface facing south of the magnetic field.

3. The method according to claim 1 or 2, wherein, The total drug escape time is adjusted by dividing the drug bolus into smaller boluses and injecting them into the patient during injection, thereby reducing the bolus volume because the individual volume of each smaller bolus is smaller than the volume of the bolus.

4. The method according to any one of claims 1 to 3, wherein, The total drug escape time is extended by heating the solution in the injection device before injecting the solution into the patient.

5. The method according to any one of claims 1 to 4, wherein, The total drug escape time is prolonged by reducing the volume rate at which the drug is injected into the patient.

6. The method according to any one of claims 1 to 5, wherein, The total escape time of the drug bolus can be adjusted by changing the shape of the bolus.

7. The method according to any one of claims 1 to 6, wherein, Shortening the total drug escape time is necessary to reach tumor portions that the injection device cannot access, requiring the magnetotactic bacteria to travel from the injection point to the tumor portion.

8. The method according to any one of claims 1 to 7, wherein, The total escape time of the drug bolus can be adjusted by influencing the shape of the bolus through the selection of an appropriate syringe.

9. The method according to any one of claims 1 to 8, wherein, The escape time of the total drug mass is shortened by increasing the surface area to volume ratio of the injected drug mass.

10. The method according to any one of claims 1 to 9, wherein, When it is desired to target hypoxic areas closer to the injection site, there is a tendency to prolong the total drug escape time.

11. A method for influencing the total drug escape time of a solution of magneto-oxygen-responsive bacteria attached to at least one of a therapeutic agent, an imaging agent, and a diagnostic agent injected into a patient, wherein, The magnetotactic-oxygen-responsive bacteria are adapted to move spontaneously when exposed to a magnetic field, and the magnetotactic-oxygen-responsive bacteria cause at least one of the therapeutic agent, the imaging agent, and the diagnostic agent to deposit in a target area with a hypoxic region within the patient's body, the method comprising: The total drug escape time of the magnetotactic responsive bacteria can be shortened by at least one of the following: increasing the surface area to volume ratio of the drug flotation, decreasing the concentration of the magnetotactic responsive bacteria in the solution, decreasing the viscosity of the solution, and, for north-seeking magnetotactic responsive bacteria, increasing the surface of the drug flotation exposed to the north of the magnetic field, and for south-seeking magnetotactic responsive bacteria, increasing the surface of the drug flotation exposed to the south of the magnetic field; or The total drug escape time of the magnetotactic responsive bacteria is prolonged by at least one of the following: reducing the surface area to volume ratio of the drug flotation, increasing the concentration of the magnetotactic responsive bacteria in the solution, increasing the viscosity of the solution, and for north-seeking magnetotactic responsive bacteria, reducing the exposure of the drug flotation to the north-facing surface of the magnetic field, and for south-seeking magnetotactic responsive bacteria, reducing the exposure of the drug flotation to the south-facing surface of the magnetic field. The shorter total drug escape time of the magnetotactic responsive bacteria is used to target one or more hypoxic areas in the target region of the patient's body that are far from the injection site.

12. The method according to claim 11, wherein, When the injection device cannot reach one or more hypoxic areas within the tumor that are further from the injection site, and the magnetotactic responsive bacteria need to travel further from the injection site to the hypoxic area, targeting the hypoxic area is considered. The method includes shortening the total drug escape time of the magnetotactic responsive bacteria by at least one of the following: increasing the surface area to volume ratio of the drug mass, reducing the concentration of the magnetotactic responsive bacteria in the drug mass, reducing the viscosity of the drug mass, and increasing the surface of the drug mass exposed to the north of the magnetic field for north-seeking magnetotactic responsive bacteria, and increasing the surface of the drug mass exposed to the south of the magnetic field for south-seeking magnetotactic responsive bacteria.

13. The method according to claim 11 or 12, further comprising increasing the surface area to volume ratio of the drug mass by dividing the drug mass into smaller drug masses.

14. The method according to any one of claims 11 to 13, wherein, The magnetotactic bacteria attach to the therapeutic agent.

15. The method according to any one of claims 11 to 14, wherein, The drug pellet was prepared by further considering the velocity distribution of the magnetotactic bacteria in the pellet.

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