Patent Application
20230330277
The contents of the electronic sequence listing (EN8.xml; Size; 2,908 bytes; and Date of Creation: May 2, 2023 is herein incorporated by reference in its entirety.
The claimed invention relates to novel anti-infective therapeutic compounds and methods. With greater particularity, the claimed invention discloses aptamer-radioisotope therapeutic agents used against microbiological infections. With still greater particularity, the claimed invention specifically targets S. aureus and E. coli infections with different energy levels of radioisotopes.
Antimicrobial resistance is currently recognized as one of the greatest threats to human health worldwide. According to the World Health Organization (WHO), antimicrobial-resistant strains are present in all parts of the world, and new resistance mechanisms continue to emerge and spread globally. Rising rates of AMR will make it increasingly difficult and expensive to control and treat infections and could affect the sustainability of some modern healthcare interventions.
In the United States alone, at least 2 million people become infected with antibiotic-resistant bacteria, with approximately 23,000 associated deaths each year. Infection with these microorganisms is further complicated by the limited number of effective therapeutic options.
Without proper handling, it is projected that there will be 10,000,000 deaths by 2050 attributed to antimicrobial resistant infections. Therefore, a new approach to treating these infections is highly needed.
Current therapeutics for microbial infections are rapidly losing their efficacy owing to anti-microbial drug resistance. A key problem with over administration of antibiotics is the ability of bacteria to adapt and become resistant to chemical antibiotics resulting in greatly diminishing or even no effect against bacterial replication.
New strategies against bacterial infection require not just new chemical entities, but entirely new anti-bacterial solutions.
By linking novel aptamers targeting specific bacteria to radioisotopes using a linker, new therapeutic and diagnostic modalities are enabled. While radioisotope therapy can be used to treat even the most dangerous forms of antimicrobial-resistant bacterial infections with limited side effects, target specificity remains lacking absent Applicant's novel claimed aptamer linked conjugate.
When used according to the claimed system, disclosed radioisotopes, which are very powerful, have multiple energies that are available for both therapeutic and diagnostic use. By utilizing highly specific novel aptamers binding to surface proteins of bacteria together with radioisotopes, the claimed invention kills bacterial targets with minimal side effects to healthy tissue without creating the increased risk of antibiotic resistance.
The claimed invention provides a novel system and related method for treatment of bacterial infections in general and offers new solutions for therapeutic resolution of antibiotic resistant bacteria strains.
Moreover, radiotherapeutics are unlike chemical compounds in that increased resistance to treatment should not be a consequence with the claimed system and method.
By combining the novel S. aureus and E. coli aptamers providing infection target specificity conjugated with therapeutic radioisotopes, a novel, broad and robust anti-microbial health and wellness system is placed in the hands of patients and medical health care providers.
The accompanying drawings are included to better illustrate exemplary embodiments of the claimed invention.
Examples: Example 1 Novel Engineered Aptamer for E. coli. Aptamers are short single stranded DNA/RNA molecules that can selectively bind to a specific target. The claimed invention utilizes Applicant's novel engineered aptamers isolated from E. coli in the first illustrative embodiment and S. aureus in the second illustrative embodiment that have specific binding towards two types of bacteria, E. coli and S. aureus. In the first illustrative embodiment, the affinity of the engineered aptamer for E. coli targets is demonstrated.
Example 2: Novel Engineered Aptamer for S. aureus. In the second illustrative embodiment, the affinity of the engineered aptamer for S. aureus targets is demonstrated.
The fairly low binding of the aptamer to the background (healthy cell) demonstrates that the aptamer has good binding strength to the target (pathogen) and is highly specific. The specificity of the disclosed aptamer enables high target affinity which enables the killing of specific pathogens with minimal side effects.
Radioisotopes: To complement the specific aptamer affinity for individual bacterial strains, aptamers are linked to radioisotopes for novel anti-bacterial effects.
Examples of linker (301) include DOTA, ethylenediaminetetraacetic acid (EDTA), and diethylenetriaminepentaacetic acid (DTPA). While all the chelators complexed thallium rapidly and efficiently, DOTA performed better than both EDTA and DTPA. This is shown in the
Due to EDTA and DTPA being acyclic chelators, having 6 and 8 donor atoms respectively, they are unstable and this results in their complexes being thermodynamically favorable but not kinetically stable. DOTA, on the other hand, enables a more stable chelation of thallium than DTPA, at least in vitro as the thallium ion directly coordinated to all the 8 donor atoms in a twisted square antiprismatic coordination as can be seen in the
In a preferred embodiment of the claimed invention, DOTA-NHS dissolved in dry DMSO (3.85 μL) is added to a solution of the aptamer with a mixture containing equal volumes of sodium phosphate buffer (0.1M, pH 8.5, 200 μL) and sodium carbonate (0,1M, pH 8.5, 200 μL). This reaction mixture is stirred in the dark at room temperature for 2 hours and then desalted using a spin-filter dialysis using a 10 kDa molecular weight cut-off centrifugal filter to stop the reaction by buffer exchange with ultrapure water. Afterwards, the solution is purified by peak collection using semi-preparatory HPLC system. Collected fractions can then be frozen at −80° C., lyophilized overnight, dissolved in ultrapure water and stored at −20° C. or −80° C.
Following the binding of the linker and aptamer, the conjugate can then be radiolabelled by adding [201TI]TICl3 (40 μL, 3 MBq) to the conjugate solution in an Eppendorf tube with ammonium acetate buffer (0.25 M, pH 5, 100 μL). To obtain [201TI]TICl3, [201TI]TI+ must first be oxidized to [201TI]TI3+, which can be done in a few methods shown in
Radioisotope variations: Further embodiments of the claimed invention include five specific types of radioisotopes, which are categorized as Alpha, Auger, Beta, Gamma and Positron. Alpha, Auger and Beta are higher energy radioisotopes compared to Gamma and Positron, which consequently makes them better and more effective for therapeutic use. In the case of bacterial infection, however, gamma and positron can also be utilized therapeutically as the amount of energy required to kill bacteria are not as high as opposed to other diseases such as treating cancer.
Therapeutic Use: According to the claimed invention, a good range of radioisotopes with different energies are hereby provided. In terms of anti-microbial embodiments, there are various options available to treat different severity of infections. Gamma/positron can be first applied to see whether the radioisotope is strong enough to kill the target. If not, stronger energy radioisotopes such as alpha, auger or beta may be subsequently deployed. Some examples of radioisotopes according to the claimed invention are Thallium-201 and Copper-64.
Thallium-201 is an isotope of Thallium that is widely used for medical imaging, such as myocardial imaging, tumor diagnosis, coronary artery diagnosis, etc. It has been used for imaging heart function under stress and rest conditions since about 1975, which makes it one of the oldest and best studied of the present-day agents.
Cu-64 is a positron and beta emitting isotope of copper that have been previously used for molecular radiotherapy and positron emission tomography (PET). As a positron emitting isotope, which is generally used for diagnostic purposes such as imaging, the 12.7 hours half-life of Cu-64 provides the flexibility to image both smaller molecules and larger, slower clearing nanoparticles.
Diagnostic Use: when radioisotopes are used for diagnostic tests, it is desirable to reduce to a minimum the radiation dose delivered to the patient during the test. As gamma and positron have lower energy, they can also be used for diagnostic purposes. Before treatment, we can use the aptamer+radioisotope (gamma/positron) conjugate to diagnose the patient (to check the location, severity of infection, etc.). After diagnosis, we can follow up with treatment using stronger radioisotopes (alpha/auger/beta).
Radiopharmaceutical Applications: With the prolonged usage of antibiotics, they are becoming less and less effective for treatment due to increasing bacteria resistance. As a result, it would require the use of second- and third-line treatments which can seriously harm patients; while some have no treatment options at all. On the other hand, radioisotope therapy can be used to treat even the most dangerous forms of antimicrobial-resistant bacterial infections with limited side effects. Thus, according to the claimed invention, radioisotopes are combined with novel aptamers to produce a drug with high specificity and energy to kill the target bacteria, for both therapeutic and diagnostic uses.
Advantages of the presently claimed system: When used according to the claimed system, disclosed radioisotopes, which are very powerful, have multiple energies that are available for both therapeutic and diagnostic use. Their escalating energy levels allow them to be utilized for adaptive radiotherapy for different severities of infections. Moreover, the claimed embodiments usage of radiation, which is harder to develop resistance to, to kill the bacteria minimizes the probability of resistance. This, combined with multiple disclosed embodiments of radioisotopes that allows different options for treatment in case pathogens develop resistance, can provide effective treatments for the ever-increasing antimicrobial resistance. With the highly specific binding to surface proteins of bacteria, the claimed invention also kills targets with minimal side effects to our body/our healthy cells.
In the description, numerous specific details are set forth in order to provide a thorough understanding of the present embodiments. It will be apparent, however, to one having ordinary skill in the art that the specific detail need not be employed to practice the present embodiments. In other instances, well-known materials or methods have not been described in detail in order to avoid obscuring the present embodiments.
Reference throughout this specification to “one embodiment”, “an embodiment”, “one example” or “an example” means that a particular feature, structure or characteristic described in connection with the embodiment or example is included in at least one embodiment of the present embodiments. Thus, appearances of the phrases “in one embodiment”, “in an embodiment”, “one example” or “an example” in various places throughout this specification are not necessarily all referring to the same embodiment or example. Furthermore, the particular features, structures or characteristics may be combined in any suitable combinations and/or sub-combinations in one or more embodiments or examples. In addition, it is appreciated that the figures provided herewith are for explanation purposes to persons ordinarily skilled in the art and that the drawings are not necessarily drawn to scale.
As used herein, the terms “comprises,” “comprising,” “includes,” “including,” “has,” “having,” or any other variation thereof, are intended to cover a non-exclusive inclusion. For example, a process, article, or apparatus that comprises a list of elements is not necessarily limited to only those elements but may include other elements not expressly listed or inherent to such process, article, or apparatus. Additionally, any examples or illustrations given herein are not to be regarded in any way as restrictions on, limits to, or express definitions of any term or terms with which they are utilized. Instead, these examples or illustrations are to be regarded as being described with respect to one particular embodiment and as being illustrative only. Those of ordinary skill in the art will appreciate that any term or terms with which these examples or illustrations are utilized will encompass other embodiments which may or may not be given therewith or elsewhere in the specification and all such embodiments are intended to be included within the scope of that term or terms. Language designating such nonlimiting examples and illustrations includes, but is not limited to: “for example,” “for instance,” “e.g.,” and “in one embodiment.”
The claimed invention has industrial applicability in the biomedical arts. In particular, the claimed invention is directly relevant to cardiac health and related therapeutic administration of pharmaceuticals for mitigation of and therapeutic effects against cardiac diseases.
This patent application is a continuation-in-part and claims priority to U.S. patent application Ser. No. 16/374,838 filed Apr. 4, 2019 to Patrick Shau-park Leung entitled “Personalized Healthcare P4 Alzheimer's Detection System and Method” which claims priority to provisional patent application 62/653,547 filed Apr. 5, 2018. Furthermore this patent application is a continuation-in-part and claims priority to U.S. patent application Ser. No. 15/666,699 filed Aug. 2, 2017 to Patrick Shau-park Leung entitled “Personalized Glucose and Insulin Monitoring System.” In addition, this patent application is a continuation-in-part and claims priority to U.S. patent application Ser. No. 15/469,138 filed Mar. 24, 2017 to Patrick Shau-park Leung entitled “Public personalized mobile health sensing system, method and device” which is a continuation of U.S. patent application Ser. No. 15/056,163 filed Feb. 29, 2016 to Patrick Shau-park Leung entitled “Mobile automated health sensing system, method and device”.
The contents of the sequence listing submitted in XML, format identified as “EN8.XML” created on 5/2/2023 and is 2,908 bytes in length is herein incorporated by reference in its entirety.
| Number | Date | Country | |
|---|---|---|---|
| Parent | 15469138 | Mar 2017 | US |
| Child | 17933525 | US | |
| Parent | 15056163 | Feb 2016 | US |
| Child | 15469138 | US | |
| Parent | 16374838 | Apr 2019 | US |
| Child | 15056163 | US |
