ALKYL-SUBSTITUTED HYDROXAMATE RESIN FOR USE IN A GENERATOR SYSTEM
20250312770 ยท 2025-10-09
Inventors
- Vanessa A. Sanders (Sound Beach, NY, US)
- Leah Marie Gajecki (New York, NY, US)
- Melissa Ann Deri (New York, NY, US)
Cpc classification
B01J45/00
PERFORMING OPERATIONS; TRANSPORTING
B01D59/26
PERFORMING OPERATIONS; TRANSPORTING
B01J2220/52
PERFORMING OPERATIONS; TRANSPORTING
B01J20/265
PERFORMING OPERATIONS; TRANSPORTING
International classification
Abstract
In one aspect, the disclosure relates to a hydroxamate-based resin for use in a .sup.44Ti/.sup.44Sc generator system. In an aspect, the carboxylate groups of a commercially available resin can be synthetically modified to produce an alkyl-substituted hydroxamate resin. In one aspect, the carboxylate resin can be a commercial resin. The disclosure also relates to a .sup.44Ti/.sup.44Sc generator system comprising an alkyl-substituted hydroxamate resin of Formula I and a method of producing .sup.44Sc, the method comprising decay of .sup.44Ti in a .sup.44Ti/.sup.44Sc generator system using an alkyl-substituted hydroxamate resin of Formula I. In an aspect, the alkyl can be methyl. The disclosure further relates to a .sup.172Hf/.sup.172Lu generator system comprising the use of an alkyl-substituted hydroxamate resin of Formula I and a method of producing .sup.172Lu comprising decay of .sup.172Hf in a .sup.172Hf/.sup.172Lu generator system using an alkyl-substituted hydroxamate resin. In an aspect, the alkyl can be methyl.
Claims
1. An alkyl-substituted hydroxamate resin of Formula I: ##STR00008## wherein R comprises a resin backbone that is a polymer-coated silica based media and wherein R comprises an alkyl group, and wherein the alkyl group is branched or unbranched, is saturated, and has from about 1 to about 12 carbon atoms in its longest chain.
2. The alkyl-substituted hydroxamate resin of claim 1, wherein R is methyl.
3. The alkyl-substituted hydroxamate resin of claim 1, wherein the resin preferentially binds a parent isotope over a daughter isotope.
4. The alkyl-substituted hydroxamate resin of claim 3, wherein the resin has a distribution coefficient of greater than or equal to 5000 for the parent isotope and a distribution coefficient of less than or equal to 5 for the daughter isotope.
5. The alkyl-substituted hydroxamate resin of claim 3, wherein the parent isotope comprises .sup.44Ti and the daughter isotope comprises .sup.44Sc.
6. The alkyl-substituted hydroxamate resin of claim 3, wherein the parent isotope comprises .sup.172Hf and the daughter isotope comprises .sup.172Lu.
7. A radionuclide generator system comprising an elution bed, wherein the elution bed comprises an alkyl-substituted hydroxamate resin of Formula I: ##STR00009## wherein R comprises a resin backbone that is a polymer-coated silica based media and wherein R comprises an alkyl group, and wherein the alkyl group is branched or unbranched, is saturated, and has from about 1 to about 12 carbon atoms in its longest chain.
8. The radionuclide generator system of claim 7, wherein R is methyl.
9. The radionuclide generator system of claim 7, wherein the radionuclide generator system preferentially retains a parent isotope in contact with the alkyl-substituted hydroxamate resin while allowing a daughter isotope to be eluted.
10. The radionuclide generator system of claim 9, wherein the resin has a distribution coefficient of greater than or equal to 5000 for the parent isotope and a distribution coefficient of less than or equal to 5 for the daughter isotope.
11. The radionuclide generator system of claim 9, wherein the parent isotope comprises .sup.44Ti and the daughter isotope comprises .sup.44Sc.
12. The radionuclide generator system of claim 9, wherein the parent isotope comprises .sup.172Hf and the daughter isotope comprises .sup.172Lu.
13. The radionuclide generator system of claim 7, wherein the elution bed has a bed volume of from about 0.3 mL to about 2 mL.
14. The radionuclide generator system of claim 9, wherein the system is loaded with from about 20 Ci to about 10 mCi of the parent isotope.
15. A method for producing .sup.44Sc, the method comprising: (a) contacting .sup.44Ti with an alkyl-substituted hydroxamate resin of Formula I: ##STR00010## wherein R comprises a resin backbone that is a polymer-coated silica based media and wherein R comprises an alkyl group, and wherein the alkyl group is branched or unbranched, is saturated, and has from about 1 to about 12 carbon atoms in its longest chain; (b) allowing at least a portion of the .sup.44Ti to decay to .sup.44Sc; and (c) eluting the .sup.44Sc.
16. The method of producing .sup.44Sc according to claim 15, wherein R is methyl.
17. The method of claim 15, wherein the .sup.44Sc is eluted with a dilute aqueous solution.
18. The method of claim 15, wherein the dilute aqueous solution comprises HCl, saline, or any combination thereof.
19. (canceled)
20. The method of claim 18, wherein the .sup.44Sc is eluted with at least one bed volume of the HCl acid.
21. The method of claim 15, wherein step (c) can be repeated one or more times before the resin releases any .sup.44Ti.
Description
BRIEF DESCRIPTION OF THE DRAWINGS
[0012] Many aspects of the present disclosure can be better understood with reference to the following drawings. The components in the drawings are not necessarily to scale, emphasis instead being placed upon clearly illustrating the principles of the present disclosure. Moreover, in the drawings, like reference numerals designate corresponding parts throughout the several views.
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[0028] Additional advantages of the invention will be set forth in part in the description which follows, and in part will be obvious from the description, or can be learned by practice of the invention. The advantages of the invention will be realized and attained by means of the elements and combinations particularly pointed out in the appended claims. It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the invention, as claimed.
DETAILED DESCRIPTION
[0029] In one aspect, in the methods disclosed herein, the carboxylate groups of commercially available Waters Accell Plus CM resin and other commercially available resins can be synthetically altered to produce alkyl-substituted hydroxamate functional groups, such as, for example, methyl-substituted hydroxamate functional groups. In an alternative aspect, a commercially available hydroxamate resin can be used in the methods disclosed herein. In a further aspect, the silica-based media of the Accell resin allows for a chemically robust resin backbone, and, without wishing to be bound by theory, the alkyl substituent on the hydroxamate functional group lowers the pK.sub.a of the modified resin, compared to the protonated/unsubstituted form which can allow for stronger metal binding.
[0030] In an aspect, R represents the alkyl substituent on the hydroxamate functional group. In a further aspect, alkyl groups can branched or unbranched, can be saturated, and can have from 1-12 carbon atoms in their longest chains, or can have 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 carbon atoms. Further in this aspect, non-limiting examples of suitable straight-chained, saturated alkyl groups include methyl, ethyl, n-propyl, n-butyl, n-pentyl, n-hexyl groups and dodecyl. In a preferred embodiment, the straight chain, saturated alkyl group can be a methyl group.
[0031] In another aspect, non-limiting examples of suitable branched, saturated alkyl groups include isopropyl, isobutyl, sec-butyl, t-butyl, 1-methylbutyl, 2-methylbutyl, 3-methylbutyl (isopentyl), 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl (neopentyl), 1-methylpentyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl groups, and 2-methyl-5-ethyldecyl. In a preferred embodiment, the branched, saturated alkyl groups include isopropyl and/or t-butyl.
[0032] In one aspect, the disclosed alkyl-substituted hydroxamate resin can be synthesized in two steps using commercially available reagents under mild conditions (Scheme 1). Further in this aspect, R is a resin backbone that can be a polymer-coated silica based media. In a still further aspect, RCOOH can be preferably a Waters Accell Plus CM resin. In one aspect, R is an alkyl group that can be branched or unbranched, can be saturated, and can have from 1-12 carbon atoms in its longest chain.
##STR00003##
[0033] In a preferred embodiment, R is methyl. In one aspect, a methyl-substituted hydroxamate resin can have Formula II, shown below:
##STR00004##
[0034] In one aspect, a methyl-substituted hydroxamate resin is synthesized as follows. In an aspect, Accell Plus CM resin (1.0 g) is suspended in water (8.0 mL) in a Falcon tube and a solution of 2,3,5,6-tetrafluorophenol (TFP) in acetonitrile (1.0 mL, 1.2 M) and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDAC) (0.385 g) are added. In a further aspect, this is allowed to mix by inversion at room temperature for 1 hour after which an additional molar equivalent of TFP solution (1.0 mL, 1.2 M) and EDAC (0.385 g) are added to the Falcon tube. In one aspect, the solution is then mixed by inversion at room temperature for 2 hours. In a further aspect, the resin can then be isolated by vacuum filtration, washed with water and acetonitrile, and dried by continuous suction. In another aspect, the ester resin can then be converted to the N-methyl-substituted hydroxamate resin by reacting with N-methylhydroxylamine hydrochloride (0.837 g) in a methanolic 1 M NaOH solution at room temperature for 18 hours and mixing by inversion. In a still further aspect, the final resin can be isolated by vacuum filtration, washed with water and acetonitrile, and dried by continuous suction. In any of these aspects, the resins can be characterized using ATR-IR spectroscopy.
[0035] In any of these aspects, the resin can preferentially bind a parent isotope over a daughter isotope. In an aspect, the resin can have a distribution coefficient of greater than or equal to 5000 for the parent isotope and a distribution coefficient of less than or equal to 5 for the daughter isotope. In one aspect, the parent isotope can be .sup.44Ti and the daughter isotope can be .sup.44Sc. In an alternative aspect, the parent isotope can be .sup.172Hf and the daughter isotope can be .sup.172Lu.
[0036] Also disclosed are radionuclide generator systems having an elution bed, wherein the elution bed contains an alkyl-substituted hydroxamate resin as disclosed herein. In a further aspect, the radionuclide generator systems preferentially retain a parent isotope in contact with the alkyl-substituted hydroxamate resin while allowing a daughter isotope to be eluted. In some aspects, the elution bed can have a bed volume of from about 0.3 mL to about 2 mL, or of about 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, or about 2 mL, or a combination of any of the foregoing values, or a range encompassing any of the foregoing values. In another aspect, the radionuclide generator system can be loaded with from about 20 Ci to about 10 mCi of the parent isotope, or about 20, 50, 75, 100, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 850, 900, or 950 Ci, or about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 mCi of the parent isotope, or a combination of any of the foregoing values, or a range encompassing any of the foregoing values. In an aspect, the parent isotope can decay while in contact with the alkyl-substituted hydroxamate resin, releasing the daughter isotope.
[0037] Also disclosed herein is a method for producing .sup.44Sc, the method including contacting .sup.44Ti with the disclosed alkyl-substituted hydroxamate resin, allowing at least a portion of the .sup.44Ti to decay to .sup.44Sc, and eluting the .sup.44Sc. In an aspect, the .sup.44Sc can be eluted with a dilute aqueous solution. In one aspect, the dilute solution can be an acid such as, for example, HCl, or can be saline, or any combination thereof. In another aspect, the HCl can have a concentration of from about 0.01 M to about 10.8 M, or from about 0.5 M to about 8 M, or of about 0.01, 0.05, 0.1, 0.25, 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, or about 10.8 M, or a combination of any of the foregoing values, or a range encompassing any of the foregoing values. In one aspect, the .sup.44Sc can be eluted with at least one bed volume of the acid, or can be eluted with two, three, four, or more bed volumes of the acid. In still another aspect, the elution step can be repeated at least once before the resin releases any .sup.44Ti. In some aspects, depending on bed volume, .sup.44Ti purity, and other factors, step (c) can be conducted at least 75 times before the resin releases any .sup.44Ti.
[0038] In one aspect, the methyl-substituted hydroxamate resin can be tested in comparison to the commercially available ZR resin in side-by-side generator evaluations. In a further aspect, the robustness of the generator can be investigated by monitoring: the time it takes until .sup.44Ti breakthrough, and the yield of .sup.44Sc. Further in this aspect, generators were loaded with a 20 Ci .sup.44Ti sample onto a 300 L bed volume column in 2 M HCl and eluted daily with four bed volumes of 0.5 M HCl. In one aspect, each eluted fraction was monitored for .sup.44Ti breakthrough and the total amount of eluted .sup.44Sc activity was calculated using high purity germanium (HPGe) spectroscopy. In another aspect, once .sup.44Ti breakthrough was observed the generator was discarded, the elution profile for the .sup.44Sc comparison is shown below (
[0039] In an aspect, the lifetime of the generator using the methyl hydroxamate resin is almost seven times longer than the ZR resin generator, and overall showed increased .sup.44Sc activity elution under the same conditions. In a further aspect, initial optimization studies also show drastically increased .sup.44Sc activity elution by varying the concentration of the HCl eluent. In one aspect, on a 300 L bed volume generator using the disclosed AM resin loaded with 100 Ci .sup.44Ti, only 9% .sup.44Sc was eluted in 4 bed volumes of 0.5 M HCl, but 86% .sup.44Sc was eluted in the same volume using 2 M HCl with no observable .sup.44Ti breakthrough.
[0040] Many modifications and other embodiments disclosed herein will come to mind to one skilled in the art to which the disclosed compositions and methods pertain having the benefit of the teachings presented in the foregoing descriptions and the associated drawings. Therefore, it is to be understood that the disclosures are not to be limited to the specific embodiments disclosed and that modifications and other embodiments are intended to be included within the scope of the appended claims. The skilled artisan will recognize many variants and adaptations of the aspects described herein. These variants and adaptations are intended to be included in the teachings of this disclosure and to be encompassed by the claims herein.
[0041] Although specific terms are employed herein, they are used in a generic and descriptive sense only and not for purposes of limitation.
[0042] As will be apparent to those of skill in the art upon reading this disclosure, each of the individual embodiments described and illustrated herein has discrete components and features which may be readily separated from or combined with the features of any of the other several embodiments without departing from the scope or spirit of the present disclosure.
[0043] Any recited method can be carried out in the order of events recited or in any other order that is logically possible. That is, unless otherwise expressly stated, it is in no way intended that any method or aspect set forth herein be construed as requiring that its steps be performed in a specific order. Accordingly, where a method claim does not specifically state in the claims or descriptions that the steps are to be limited to a specific order, it is no way intended that an order be inferred, in any respect. This holds for any possible non-express basis for interpretation, including matters of logic with respect to arrangement of steps or operational flow, plain meaning derived from grammatical organization or punctuation, or the number or type of aspects described in the specification.
[0044] All publications mentioned herein are incorporated herein by reference to disclose and describe the methods and/or materials in connection with which the publications are cited. The publications discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication provided herein can be different from the actual publication dates, which can require independent confirmation.
[0045] While aspects of the present disclosure can be described and claimed in a particular statutory class, such as the system statutory class, this is for convenience only and one of skill in the art will understand that each aspect of the present disclosure can be described and claimed in any statutory class.
[0046] It is also to be understood that the terminology used herein is for the purpose of describing particular aspects only and is not intended to be limiting. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the disclosed compositions and methods belong. It will be further understood that terms, such as those defined in commonly used dictionaries, should be interpreted as having a meaning that is consistent with their meaning in the context of the specification and relevant art and should not be interpreted in an idealized or overly formal sense unless expressly defined herein.
[0047] Prior to describing the various aspects of the present disclosure, the following definitions are provided and should be used unless otherwise indicated. Additional terms may be defined elsewhere in the present disclosure.
Definitions
[0048] As used herein, comprising is to be interpreted as specifying the presence of the stated features, integers, steps, or components as referred to, but does not preclude the presence or addition of one or more features, integers, steps, or components, or groups thereof. Moreover, each of the terms by, comprising, comprises, comprised of, including, includes, included, involving, involves, involved, and such as are used in their open, non-limiting sense and may be used interchangeably. Further, the term comprising is intended to include examples and aspects encompassed by the terms consisting essentially of and consisting of. Similarly, the term consisting essentially of is intended to include examples encompassed by the term consisting of.
[0049] As used in the specification and the appended claims, the singular forms a, an and the include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to a radionuclide, a resin, or an eluent, includes, but is not limited to, mixtures or combinations of two or more such radionuclides, resins, or eluents, and the like.
[0050] It should be noted that ratios, concentrations, amounts, and other numerical data can be expressed herein in a range format. It will be further understood that the endpoints of each of the ranges are significant both in relation to the other endpoint, and independently of the other endpoint. It is also understood that there are a number of values disclosed herein, and that each value is also herein disclosed as about that particular value in addition to the value itself. For example, if the value 10 is disclosed, then about 10 is also disclosed. Ranges can be expressed herein as from about one particular value, and/or to about another particular value. Similarly, when values are expressed as approximations, by use of the antecedent about, it will be understood that the particular value forms a further aspect. For example, if the value about 10 is disclosed, then 10 is also disclosed.
[0051] When a range is expressed, a further aspect includes from the one particular value and/or to the other particular value. For example, where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure, e.g. the phrase x to y includes the range from x to y as well as the range greater than x and less than y. The range can also be expressed as an upper limit, e.g. about x, y, z, or less and should be interpreted to include the specific ranges of about x, about y, and about z as well as the ranges of less than x, less than y, and less than z. Likewise, the phrase about x, y, z, or greater should be interpreted to include the specific ranges of about x, about y, and about z as well as the ranges of greater than x, greater than y, and greater than z. In addition, the phrase about x to y, where x and y are numerical values, includes about x to about y.
[0052] It is to be understood that such a range format is used for convenience and brevity, and thus, should be interpreted in a flexible manner to include not only the numerical values explicitly recited as the limits of the range, but also to include all the individual numerical values or sub-ranges encompassed within that range as if each numerical value and sub-range is explicitly recited. To illustrate, a numerical range of about 0.1% to 5% should be interpreted to include not only the explicitly recited values of about 0.1% to about 5%, but also include individual values (e.g., about 1%, about 2%, about 3%, and about 4%) and the sub-ranges (e.g., about 0.5% to about 1.1%; about 5% to about 2.4%; about 0.5% to about 3.2%, and about 0.5% to about 4.4%, and other possible sub-ranges) within the indicated range.
[0053] As used herein, the terms about, approximate, at or about, and substantially mean that the amount or value in question can be the exact value or a value that provides equivalent results or effects as recited in the claims or taught herein. That is, it is understood that amounts, sizes, formulations, parameters, and other quantities and characteristics are not and need not be exact, but may be approximate and/or larger or smaller, as desired, reflecting tolerances, conversion factors, rounding off, measurement error and the like, and other factors known to those of skill in the art such that equivalent results or effects are obtained. In some circumstances, the value that provides equivalent results or effects cannot be reasonably determined. In such cases, it is generally understood, as used herein, that about and at or about mean the nominal value indicated 10% variation unless otherwise indicated or inferred. In general, an amount, size, formulation, parameter or other quantity or characteristic is about, approximate, or at or about whether or not expressly stated to be such. It is understood that where about, approximate, or at or about is used before a quantitative value, the parameter also includes the specific quantitative value itself, unless specifically stated otherwise.
[0054] As used herein, the term effective amount refers to an amount that is sufficient to achieve the desired modification of a physical property of the composition or material. For example, an effective amount of a resin refers to an amount that is sufficient to achieve the desired improvement in the property modulated by the formulation component, e.g. achieving the desired level of separation of .sup.44Ti from .sup.44Sc over the desired time period. The specific level in terms of wt % in a resin required as an effective amount will depend upon a variety of factors including the amount of resin, chemical identity of the resin including any substituents, initial .sup.44Ti to .sup.44Sc ratio, and eluent identity and concentration.
[0055] As used herein, the terms optional or optionally means that the subsequently described event or circumstance can or cannot occur, and that the description includes instances where said event or circumstance occurs and instances where it does not.
[0056] Cold as used herein refers to a reaction or molecule that is not radioactive. In an aspect, reactions and processes can be optimized using cold reagents and materials prior to performing the same reactions and processing with radioactive reagents and/or materials.
[0057] Unless otherwise specified, temperatures referred to herein are based on atmospheric pressure (i.e. one atmosphere).
[0058] Now having described the aspects of the present disclosure, in general, the following Examples describe some additional aspects of the present disclosure. While aspects of the present disclosure are described in connection with the following examples and the corresponding text and figures, there is no intent to limit aspects of the present disclosure to this description. On the contrary, the intent is to cover all alternatives, modifications, and equivalents included within the spirit and scope of the present disclosure.
EXAMPLES
[0059] The following examples are put forth so as to provide those of ordinary skill in the art with a complete disclosure and description of how the compounds, compositions, articles, devices and/or methods claimed herein are made and evaluated, and are intended to be purely exemplary of the disclosure and are not intended to limit the scope of what the inventors regard as their disclosure. Efforts have been made to ensure accuracy with respect to numbers (e.g., amounts, temperature, etc.), but some errors and deviations should be accounted for. Unless indicated otherwise, parts are parts by weight, temperature is in C. or is at ambient temperature, and pressure is at or near atmospheric.
Example 1: Materials and Methods
Chemicals and Reagents
[0060] All chemicals were used without further purification. Hydroxylamine hydrochloride, N-methylhydroxylamine hydrochloride, N-phenylhydroxylamine, 2,3,5,6-tetrafluorophenol (TFP), 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC), potassium hydroxide, SOCl.sub.2, anhydrous diethylether, acetonitrile (ACN), and HPLC grade methanol (MeOH) were purchased from Sigma Aldrich. Optima grade hydrochloric acid (HCl) was purchased from Fisher Scientific (Pittsburgh, PA, USA) and diluted to suitable concentrations (6 M, 4 M, 2 M, 0.5 M, 0.1 M) with 18 M water (25 C., Milli-Q, Millipore, Burlington, MA, USA). ZR resin was purchased from Triskem Intl. (Brunz, France), Amberlite IRC50 resin and Amberlite CG50 resin were purchased from Sigma Aldrich and Accell Plus CM resin was purchased from Waters (Milford, MA, USA). Deionized Milli-Q water (18 M, Millipore) which had been purified by passing through a 10 cm column of Chelex 100 resin (Bio-Rad Laboratories, Hercules, CA, USA) was used in all reactions and solution preparations.
Measurement of Radioactivity
[0061] The absolute radioactivity of .sup.44Sc and .sup.44Ti was measured by -spectrometry using a high-purity germanium (HPGe) detector, samples were counted for 10 minutes. .sup.44Ti was detected directly via the 67.87 keV (93.0%) and 78.32 keV (96.4%) -lines and .sup.44Sc was detected via the 1157.02 keV (99.9%) -line.
Synthesis of Resins
[0062] Accell Resins: Ester Resin. Accell Plus CM resin (1.0 g) was suspended in Chelex water (8.0 mL) in a 15 mL Falcon tube. TFP solution (1.0 mL, 1.2 M in ACN, 1.20 mmol) and EDAC (0.385 g, 2.48 mmol) were added to the Falcon tube. The reaction was mixed by inversion at room temperature for 1 hour, after which an additional 1.0 mL of TFP solution (1.2 M in ACN, 1.20 mmol) and EDAC (0.385 g, 2.48 mmol) were added to the reaction to ensure complete conversion of the carboxylate groups to the ester resin. The reaction was mixed by inversion at room temperature for a further 2 hours after which the final resin was isolated by vacuum filtration and washed with 315 mL water and 315 mL ACN and dried by continuous suction. This resin can be stored dry under ambient conditions without any apparent degradation or hydrolysis. IR (ATR, selected bands, v.sub.max): 3384, 1781, 1670, 1065, 956, 794, 452 cm.sup.1.
[0063] Accell Resins: UH Resin (AU). Hydroxylamine hydrochloride (0.695 g, 10.0 mmol) was dissolved in 1 M NaOH (1.0 mL) and MeOH (2.0 mL) to form the free base hydroxylamine. The ester functionalized resin (1.00 g) was added to this solution in a 15 mL Falcon tube and was mixed by inversion at room temperature for 18 hours. The hydroxamate resin was then isolated by vacuum filtration and washed with 315 mL water and 315 mL ACN. This resin can be stored dry under ambient conditions without any apparent degradation or hydrolysis. IR (ATR, selected bands, v.sub.max): 3347, 1731, 1660, 1451, 1062, 795, 451 cm.sup.1. Formation of the hydroxamate functional group was also verified by the visual formation of a dark red complex upon addition of Fe(III) in dilute acid to the resin.
[0064] Accell Resins: MH Resin (AM). The N-methylhydroxamate Accell resin can be synthesized by substituting N-methylhydroxylamine hydrochloride (0.837 g, 10.0 mmol) for the hydroxylamine hydrochloride in the above procedure. IR (ATR, selected bands, v.sub.max): 3361, 1733, 1654, 1449, 1062, 795, 452 cm.sup.1. Formation of the hydroxamate functional group was also verified by the visual formation of a dark red complex upon addition of Fe(III) in dilute acid to the resin.
[0065] Accell Resins: PH Resin (AP). N-phenylhydroxylamine (1.09 g, 10.0 mmol) was dissolved in MeOH (3.00 mL) and the ester functionalized resin (1.00 g) was added to this solution in a 15 mL Falcon tube. The reaction was mixed by inversion at room temperature for 18 hours and isolated and washed in accordance with the above procedure. IR (ATR, selected bands, v.sub.max): 3355, 1724, 1662, 1451, 1061, 795, 452 cm.sup.1. Formation of the hydroxamate functional group was also verified by the visual formation of a dark red complex upon addition of Fe(III) in dilute acid to the resin.
[0066] IRC50 Resins: Anhydride Resin. IRC50 resin (2.00 g) was suspended in SOCl.sub.2 (6.00 mL) in a 25 mL round bottom flask fitted with a condenser, and the solution was refluxed for 4 hours. The resin was isolated by vacuum filtration and washed excessively with anhydrous diethyl ether to remove all unreacted SOCl.sub.2 and dried by continuous suction. IR (ATR, selected bands, v.sub.max): 1800, 1745, 1456, 1390, 1258, 1111, 1007, 960, 853, 738, 713 cm.sup.1.
[0067] IRC50 Resins: UH Resin. Hydroxylamine hydrochloride (0.500 g, 7.20 mmol) was dissolved in MeOH (5.0 mL) and added to a solution of KOH (0.404 g, 7.20 mmol) in MeOH (5.0 mL). The KCl salt precipitated from solution immediately, but the solution was stored in the fridge for an hour to ensure complete precipitation of the salt. The salt was removed by filtration and the MeOH solution added to the anhydride resin (1.00 g) in a 15 mL Falcon tube. The reaction was mixed by inversion at room temperature for 1 hour, after which 3 M KOH (2.4 mL in MeOH, 7.20 mmol) was added to the Falcon tube. The reaction was mixed by inversion at room temperature for 5 minutes after which the resin was isolated by vacuum filtration and washed with MeOH. The potassium salt resin is then suspended in 0.1 M HCl (5.0 mL) and mixed by inversion at room temperature for a few minutes to protonate. The final resin was isolated by vacuum filtration and washed with water and MeOH and dried by continuous suction. This resin undergoes slow hydrolysis under ambient conditions over several days. IR (ATR, selected bands, v.sub.max): 3300, 1724, 1670, 1547, 1477, 1447, 1387, 1346, 1200, 1015, 969 cm.sup.1. Formation of the hydroxamate functional group was also verified by the visual formation of a dark red complex upon addition of Fe(III) in dilute acid to the resin.
[0068] IRC50 Resins: MH Resin. The N-methylhydroxamate IRC50 resin can be synthesized by substituting N-methylhydroxylamine hydrochloride (0.601 g, 7.20 mmol) for the hydroxylamine hydrochloride in the above procedure. This resin undergoes slow hydrolysis under ambient conditions over several days. IR (ATR, selected bands, v.sub.max): 3366, 1719, 1642, 1548, 1473, 1445, 1408, 1384, 1350, 1238, 1167, 1131, 1017, 966 cm.sup.1. Formation of the hydroxamate functional group was also verified by the visual formation of a dark red complex upon addition of Fe(III) in dilute acid to the resin.
[0069] IRC50 Resins: PH Resin. N-phenylhydroxylamine (0.786 g, 7.20 mmol) was dissolved in MeOH (6.0 mL) and added to the anhydride resin (1.00 g) in a 15 mL Falcon tube. The reaction was mixed by inversion at room temperature for 1 hour after which 3 M KOH (2.4 mL in MeOH, 7.20 mmol) was added to the reaction and mixed by inversion for 5 minutes at room temperature. The phenylhydroxamate resin was then isolated and washed in accordance with the above procedure. This resin undergoes slow hydrolysis under ambient conditions over several days. IR (ATR, selected bands, v.sub.max): 3361, 1692, 1652, 1540, 1473, 1446, 1403, 1343, 1256, 1181, 1019 cm.sup.1. Formation of the hydroxamate functional group was also verified by the visual formation of a dark red complex upon addition of Fe(III) in dilute acid to the resin.
[0070] CG50 Resins: Anhydride Resin. CG50 resin (2.00 g) was suspended in SOCl.sub.2 (6.00 mL) in a 25 mL round bottom flask fitted with a condenser, and the solution was refluxed for 4 hours. The IR of the resulting resin showed incomplete conversion, so the resin was refluxed for a further 4 hr in fresh SOCl.sub.2 (6.0 mL). The resin was isolated by vacuum filtration and washed excessively with anhydrous diethyl ether to remove all unreacted SOCl.sub.2 and dried by continuous suction. IR (ATR, selected bands, v.sub.max): 1800, 1754, 1455, 1389, 1255, 1112, 1007, 960. 845, 737, 603 cm.sup.1.
[0071] CG50 Resins: UH Resin. The unsubstituted hydroxamate CG50 resin can be synthesized by substituting the CG50 anhydride resin (1.00 g) for the IRC50 anhydride resin in the above IRC50 UH resin procedure. IR (ATR, selected bands, v.sub.max): 3294, 1722, 1674, 1547, 1475, 1445, 1386, 1342, 1250, 1191, 1007, 965 cm.sup.1. Formation of the hydroxamate functional group was also verified by the visual formation of a dark red complex upon addition of Fe(III) in dilute acid to the resin.
[0072] CG50 Resins: MH Resin. The N-methylhydroxamate CG50 resin can be synthesized by substituting the CG50 anhydride resin (1.00 g) for the IRC50 anhydride resin in the above IRC50 MH resin procedure. IR (ATR, selected bands, v.sub.max): 3382, 1720, 1649, 1548, 1472, 1444, 1406, 1386, 1344, 1245, 1166, 1130, 1019, 966 cm.sup.1. Formation of the hydroxamate functional group was also verified by the visual formation of a dark red complex upon addition of Fe(III) in dilute acid to the resin.
[0073] CG50 Resins: PH Resin. The N-phenylhydroxamate CG50 resin can be synthesized by substituting the CG50 anhydride resin (1.00 g) for the IRC50 anhydride resin in the above IRC50 PH resin procedure. IR (ATR, selected bands, v.sub.max): 3366, 1698, 1647, 1485, 1446, 1387, 1340, 1253, 1172, 1018, 965 cm.sup.1. Formation of the hydroxamate functional group was also verified by the visual formation of a dark red complex upon addition of Fe(III) in dilute acid to the resin.
IR Measurement
[0074] Attenuated total reflectance infrared (ATR-IR) spectroscopy was performed by using a Thermo Scientific Nicolet iS50 FTIR Spectrometer. The ATR-IR data were collected on dry solid samples and processed by using OMNIC version 9.3.32 software.
Preparation and Evaluation of .sup.44Ti/.sup.44Sc Generators
[0075] Production of .sup.44Ti used in the radionuclide generators was conducted by irradiation of a scandium sputtering target (American Elements) at the Brookhaven Linac Isotope Producer (BLIP).
[0076] 10 mL tapered Bio-Rad columns were rinsed with Chelex water and filled to the bed volume (BV) with the corresponding resin that had been weighed and slurried in Chelex water. A small piece of glass wool was added to the top of the resin for protection, and the column washed with Chelex water and 2 M HCl or 0.1 M HCl. A sample of .sup.44Ti was evaporated to dryness and taken up in either 2 M HCl or 0.1 M HCl and loaded onto the columns by pipette. The solutions were allowed to elute by gravity. The generators were eluted 3 to 5 times per week with 4 BV of dilute HCl and collected and counted by fraction.
Total Organic Content (TOC) Studies
[0077] 10 mL tapered Bio-Rad columns were rinsed with Chelex purified water and filled to a bed volume (BV) of 300 L with the corresponding resin that had been weighed and slurried in Chelex purified water. Two columns were prepared with the ZR resin and two were prepared with the synthesized AM resin. One of each type of resin column was rinsed with 50 BV of 0.5 M HCl and the other two remaining columns with each type of resin was eluted with 50 BV of 2 M HCl. Samples were collected after 10, 20, 30, 40, and 50 BV of eluent and sent for total organic content (TOC) analysis at Long Island Analytical Laboratories Inc. (Holbrook, New York 11741). A blank of each acid used (0.5 and 2 M HCl) was also sent for analysis for comparison.
Radiolabeling Experiments
[0078] Dry Down Method. In order to obtain the .sup.44Sc in a form suitable for radiolabeling, dry down processing was implemented where the generator eluted .sup.44Sc was evaporated dry on a hot plate in an open beaker at 140 C. and re-dissolved in 0.25 M NH.sub.4OAc pH 4 buffer. Radiolabeling of generator eluted .sup.44Sc with DOTA, HOPO and NOTA was performed by mixing the ligand stock solution (0.04 mg/mL, 0.25 M NH.sub.4OAc pH 4 buffer) with the post-processed .sup.44Sc (i.e., dried and re-dissolved .sup.44Sc) in a 1.5 mL Eppendorf tube and heating the solution at 90 C. for one hour in a thermomixer. The only parameter varied was the ligand concentration, which was pre-determined as a ligand molar excess over the .sup.44Sc molar amount (calculated by activity). The activity of .sup.44Sc used in radiolabeling varied from 25-50 Ci for the AM generator experiments, and 5-10 Ci for the ZR generator experiments due to varying generator and post-processing behavior.
[0079] Direct Radiolabeling Method. The .sup.44Sc generator eluent (in 0.5 M HCl) was obtained and neutralized with dilute NaOH solution and then used directly (e.g., without dry down processing) in subsequent labeling reactions. Radiolabeling of generator eluted .sup.44Sc with DOTA was performed by mixing the ligand stock solution (0.04 mg/mL, 0.25 M NH.sub.4OAc pH 4 buffer) with the neutralized .sup.44Sc eluent in a 1.5 mL Eppendorf tube and heating the solution at 90 C. for one hour in a thermomixer. The only parameter varied was the ligand concentration, which was pre-determined as a ligand molar excess over the .sup.44Sc molar amount (calculated by activity). The activity of .sup.44Sc used in radiolabeling varied from 25-50 Ci.
[0080] For both methods, the dry down method and the direct radiolabeling method the radiolabeling yield was determined using thin-layer chromatography (iTLC-SG strips, Agilent). The plates were developed in 0.04 M NH.sub.4OAc/MeOH (50/50) pH 5 buffer and were counted for 1 minute on a BIOSCAN AR 2000. The R.sub.f values for the free .sup.44Sc and labeled .sup.44Sc (with all three ligands) were 0 and 0.9 respectively.
Example 2: Results and Discussion
Resin Synthesis and Characterization
[0081] The unsubstituted hydroxamate Accell resin (AU) was prepared following modified literature procedures by functionalizing the carboxylate groups of the Accell resin first to an ester, followed by conversion to the hydroxamate by reaction with hydroxylamine hydrochloride. The N-methylhydroxamate Accell resin (AM) can be synthesized using the same procedure substituting hydroxylamine hydrochloride for N-methylhydroxylamine hydrochloride. The N-phenylhydroxamate Accell resin (AP) can be synthesized from the ester functionalized resin using a similar procedure, but without the use of NaOH, as the N-phenylhydroxylamine is already in the free base form (Scheme 2). Addition of 1.0 mL of NaOH to the N-phenylhydroxylamine resin reaction results in hydrolysis of the ester resin back to the carboxylate form.
##STR00005##
[0082] The characterization of the Accell resins by ATR-IR was difficult due to weak signal intensities caused by low ligand density and the overpowering silica peaks of the resin backbone. Because of this only the carbonyl region of the IR spectra was used in characterization. The starting Accell carboxylate resin shows asymmetric (v.sub.as) and symmetric (v.sub.s) CO stretching frequencies at 1562 cm.sup.1 and 1408 cm.sup.1 respectively. In the intermediate ester resin this CO stretching frequency shifts to higher energy at 1781 cm.sup.1. All three hydroxamate resins show a diagnostic CO stretching frequency signal around 1730 cm.sup.1 as well as a second peak at around 1450 cm.sup.1, there is also evidence for the v.sub.as and v.sub.s carboxylate CO peaks at approximately 1550 cm.sup.1 and 1400 cm.sup.1 respectively. There is no evidence of the carboxylate peaks in the ester resin, suggesting that there is some hydrolysis of the ester in the hydroxylamine reaction. The peaks seen at approximately 3400, 1650, 1065, 960, 800 and 450 cm.sup.1 in all of the Accell resin spectra are assigned as silica peaks, this was confirmed by direct comparison with the IR spectrum of crude silica gel. Formation of the hydroxamate functional group was also verified by the visual formation of a dark red complex upon addition of Fe(III) in dilute acid to the resin.
[0083] The procedure for the synthesis of the IRC50 and CG50 hydroxamate resins are the same but differ from the Accell resin synthesis. Attempts to use the same procedure as described above using IRC50 or CG50 resin did not result in formation of the ester functionalized resin in the first step, but rather formation of an anhydride. This was determined by characterization of the intermediate product using ATR-IR, which showed two CO stretching frequencies at 1800 cm.sup.1 and 1750 cm.sup.1, this was true for both the IRC50 and CG50 resins. An alternate route to hydroxamate synthesis is conversion of carboxylates into acid chlorides which are then reacted with hydroxylamine, this has been suggested to work in the literature for similar resins. Reaction of IRC50 and CG50 with SOCl.sub.2 under reflux resulted in the same anhydride product that was observed in the ester formation attempt (Scheme 3), this was evident by the identical IR spectrum of the product. Due to the high ligand density of the resins and the close contact of the neighboring carboxylate groups, all attempts at formation of the acid chlorides or esters (as in the Accell resin synthesis) always resulted in formation of the anhydride derivative without any apparent formation of the ester or acid chloride in any appreciable amounts.
[0084] There is literature evidence that it is possible to convert anhydrides into a 1:1 mix of hydroxamates and carboxylic acids by reaction with the free-based hydroxylamine and base. Based on this the hydroxamate IRC50 and CG50 resins were synthesized using modified literature procedures using the anhydride resin (Scheme 3). The hydroxylamine hydrochloride and N-methylhydroxylamine hydrochloride were free-based by reaction with one equivalent of KOH in MeOH, followed by removal of the KCl salt. This MeOH hydroxylamine solution was then added to the dry anhydride resin and allowed to react for 1 hour at room temperature. An additional molar equivalent of KOH in MeOH was then added briefly to catalyze the reaction to completion, but it was found that prolonged exposure of the hydroxamate resin to base resulted in hydrolysis back to the starting carboxylate resin. This product is presumably the potassium salt of the resin, so to protonate the hydroxamate groups the resin was washed in 0.1 M HCl. Exposure of this resin to 1 M HCl solutions resulted in hydrolysis of the hydroxamates back to carboxylates. Since the N-phenylhydroxylamine is already in the free base form this can be dissolved in MeOH and reacted directly with the anhydride resin, although addition of the catalytic KOH after the initial 1-hour reaction seemed to aid in the product formation.
##STR00006##
[0085] All six of these resins form dark red complexes in dilute acid solution with Fe(III) suggesting successful formation of the hydroxamate group. The IR spectra of resins show peaks due to both the carboxylate groups as well as the hydroxamate groups, with the CO stretching frequency of the hydroxamate found around 1720 cm.sup.1 in all six resins. With the unsubstituted hydroxamate IRC50 and CG50 resins there was also sometimes an IR peak around 1680 cm.sup.1 (and no peak at 1720 cm.sup.1) that was not seen in the N-methyl or N-phenyl derivatives. This is likely due to the CN stretching frequency that is present in the resonance form of the unsubstituted hydroxamate (Scheme 4).
##STR00007##
K.SUB.D .Studies of AM Resin
[0086] Results of the distribution coefficient studies of Sc and Ti on the AM resin in varying concentration of HCl are shown in
Generator Performance
[0087] Study 1: Load in 2 M HCl, Elute in 0.5 M HCl. All generators had 300 L BV and were loaded on the same day with 20 Ci of .sup.44Ti as well as three 300 UL BV blank generators loaded with 13 Ci of .sup.44Ti using the same procedure with unmodified IRC50, CG50 and Accell resins to ensure the carboxylate groups of the parent resin were not contributing to any metal binding. All nine synthesized resins were tested as well as commercially available ZR resin, the mass of resin used in each generator is given in Table 1. All generators were then subsequently eluted with four bed volumes of 0.5 M HCl daily (or semi-daily for the longer lasting generators) to monitor the amount of .sup.44Sc eluted as well as any .sup.44Ti breakthrough.
TABLE-US-00001 TABLE 1 Masses of Resins Used for 300 L BV Generator in a 10 mL Tapered Column Mass (mg) Resin Blank Unsubstituted Methyl Phenyl Accell 115 125 120 110 IRC50 110 170 160 150 CG50 80 100 105 110 ZR 80
[0088] Unsurprisingly all unfunctionalized resins (in the carboxylate form) showed complete .sup.44Ti breakthrough during the load and failed to bind appreciable amounts of either metal (Table 2). For the remaining resins, during the 2 M HCl load experiment it was found that all functionalized IRC50 and CG50 resins failed to bind the Ti and displayed complete .sup.44Ti breakthrough either during the load or in the subsequent elution (Elute 1). This is likely due to hydrolysis of the hydroxamate groups in the 2 M HCl as described previously. Similar results were found by Holland et al with the unsubstituted Accell based hydroxamate resin, where the loading .sup.89Zr(IV) efficiency was found to be >99.9% at HCl concentrations of 2 M, but at higher concentrations the binding affinity of the resin decreases drastically. The AU and AM resins, however, displayed 100% .sup.44Ti loading efficiency, and the ZR resin showed a small amount of .sup.44Ti breakthrough during the load leading to 93.5% .sup.44Ti loading efficiency.
TABLE-US-00002 TABLE 2 2M HCl Load Generator Experiment .sup.44TI Breakthrough % Ti Breakthrough Resin Load Elute 1 Accell Blank 100 IRC50 Blank 97 CG50 Blank 100 ZR 6.5 Accell UH (AU) 0 0 Accell MH (AM) 0 0 Accell PH (AP) 72 18 IRC50 UH 96 3 IRC50 MH 100 IRC50 PH 72 26 CG50 UH 93 4 CG50 MH 100 CG50 PH 100
[0089] The ZR, AU and AM generators were eluted daily with 4 bed volumes of 0.5 M HCl. The resulting .sup.44Sc activity eluted as well as any potential .sup.44Ti breakthrough was monitored through HPGe spectroscopy. The AU generator displayed the highest eluted .sup.44Sc activity, almost double that of the ZR and AM generators, but also was the first to display .sup.44Ti breakthrough. The AU generator was eluted seven times (total 28 BV) over seven days before 0.45% of the load activity .sup.44Ti breakthrough was observed on the 8th day, due to the .sup.44Ti breakthrough this generator was no longer eluted. The ZR generator was eluted 10 times (total 40 BV) over 10 days before 0.16% of the load activity .sup.44Ti breakthrough was observed. The AM generator was eluted 75 times (total 300 BV) over several months before 0.3% of the load activity .sup.44Ti breakthrough was observed. The elution profile of these three generators is displayed in
[0090] Interestingly, the activity of eluted .sup.44Sc was initially highest for the AU generator (around 20%), while the ZR and AM generators eluted lower activities in the same volume of 0.5 M HCl (both around 10%). However, over time the elution activity for .sup.44Sc from the AM generator increased to a maximum of 50% and consistently eluted 45% for the remainder of its lifetime. The reason for this increased activity over time is still unclear, but it may be a result of the protonation of hydroxamate groups in the resin by the dilute HCl over time leading to fewer open coordination sites for the free .sup.44Sc to bind.
[0091] A scaled-up generator experiment was similarly conducted using the AM resin to test the system in a more clinically relevant scale as it showed extremely promising results in the small scale 7 40 kBq (20 Ci) generator study. A 1 mL BV column (0.400 g AM resin) was loaded with 37 MBq (1 mCi) of .sup.44Ti in 2 M HCl. The generator was eluted daily with 4 BV of 2 M HCl and the fractions counted to monitor the .sup.44Sc elution activity as well as any potential .sup.44Ti breakthrough. The elution profile of this generator is shown in
[0092] Attempts to further improve this generator towards direct radiolabeling conditions were conducted by eluting in more dilute HCl. A 0.5 mL BV column (0.200 g AM resin) was loaded in 2 M HCl with 37 MBq (1 mCi) of .sup.44Ti and then eluted daily with 4 BV of 0.5 M HCl. The elution profile of this generator is shown in
[0093] From the generator studies that have been conducted to date using the AM resin, there has not been any observation of resin degradation or structural damage from radiation from the .sup.44Ti or .sup.44Sc, even in the higher activity 1 mCi generators. There is a risk that with higher activity generators the hydroxamate-based resin may undergo radiolytic damage from the high energy gammas from .sup.44Sc or the positron emission, but further studies are needed.
[0094] Study 2: Load in 0.1 M HCl, Elute in 0.5 M HCl. The blank generators were only tested in the 2 M HCl loading experiment as all three showed 100% .sup.44Ti breakthrough during the load in the first fraction. The IRC50 resins were also not tested as they had undergone hydrolysis and degraded prior to this study. All generators had 300 L BV and were loaded on the same day with 20 Ci of .sup.44Ti and the same mass of resin was used as in study 1 (Table 1). All generators were then subsequently eluted with four bed volumes of 0.5 M HCl daily (or semi-daily for the longer lasting generators) to monitor the amount of .sup.44Sc eluted as well as any .sup.44Ti breakthrough.
[0095] Similar results were found in the 0.1 M HCl loading experiments, with the ZR, AU and AM resins giving the longest lifetimes without .sup.44Ti breakthrough. The IRC50 resins were not tested in the 0.1 M HCl loading experiments as all three of the functionalized resins had degraded in air overtime (few weeks). The slow hydrolysis of the IRC50 resins was evident by the changing IR spectra as well as failed Fe(III) binding tests. The lower molarity HCl loading solution did slow down hydrolysis of the resins evident by the lower .sup.44Ti breakthrough (Table 3), but all three CG50 resins as well as the Accell phenylhydroxamate resin (AP) showed consistent breakthrough over the load and two subsequent elutions, so they were abandoned and no longer eluted daily.
TABLE-US-00003 TABLE 3 0.1M HCl Load Generator Experiment .sup.44Ti Breakthrough % Ti Breakthrough Resin Load Elute 1 Elute 2 ZR 3 2 0 Accell UH (AU) 0 0 0 Accell MH (AM) 0 0 0 Accell PH (AP) 20 10 6 CG50 UH 42 7 4 CG50 MH 64 19 2 CG50 PH 69 15 3
[0096] Similar to study 1, the AU resin was the first to display .sup.44Ti breakthrough after 5 elutions (total 20 BV) and the ZR resin after 13 elutions (total 52 BV) (
[0097] Study 3: Load in 2 M HCl, Elute in 2 M HCl. Due to the strong performance of the AM resin compared to all other synthesized resins, only the AM resin was used in further studies in direct comparison to commercially available ZR resin. The low elution activity of .sup.44Sc in the two previous studies prompted the use of higher molarity HCl to elute the generators in attempts to increase the amount of .sup.44Sc being eluted in the same volume of eluent. Both the ZR and AM generators had 300 L BV and were loaded on the same day with 20 Ci of .sup.44Ti and used the same mass of resin that was used in the previous studies. Both generators were loaded in 2 M HCl and were subsequently eluted using 4 bed volumes of 2 M HCl daily to monitor the amount of .sup.44Sc eluted as well as any .sup.44Ti breakthrough. The elution profile of both generators, along with an AU (V$LG) generator, is shown in
[0098] It is immediately evident that the use of higher concentrated acid did indeed increase the amount of .sup.44Sc being eluted with values of around 70% for the AM generator compared to 45% (study 1) and 30% (study 2). It is also evident that the lifetime of both the ZR and AM generators was decreased with the use of 2 M HCl, this was more drastic for the AM generator that displayed .sup.44Ti breakthrough (0.45%) during the 20th elution (after 76 BV). The lifetime of the AM generator under these conditions was still more than twice that of the ZR generator, but it appears that the use of more concentrated HCl as eluent may be less desired as it possibly expedites the .sup.44Ti breakthrough. Further studies are needed varying the concentration of HCl eluent to better ascertain the behavior of the AM resin under varying pHs.
[0099] Another AM generator was prepared around the same time both loading and eluting in 2 M HCl on a much bigger scale to assess the viability of this resin in a clinically relevant generator system. A 1 mL BV column (0.400 g AM resin) was loaded with 1 mCi of .sup.44Ti, this generator study was conducted in a hotcell due the high activity. This generator was eluted daily with 4 BV of 2 M HCl and the fractions counted to monitor the .sup.44Sc elution activity as well as any potential .sup.44Ti breakthrough. The elution profile of this generator is shown below in
[0100] Study 4: Large Scale Load in 2 M, HCl Elute in 0.5 M HCl. The long lifetime of the small-scale (20 Ci, low activity) generators eluted in 0.5 M HCl and the high activity elution of the 1 mCi generator eluted in 2 M HCl led to the study of larger scale (high activity) generators eluting in 0.5 M HCl to see if higher activities of .sup.44Sc could be eluted in 0.5 M HCl. A 500 UL BV AM resin (0.200 g) generator was loaded with 720 Ci of .sup.44Ti in 2 M HCl and this generator was eluted daily with 0.5 M HCl fractions. Initially this (720 Ci, high activity) generator was eluted with 40.5 mL fractions (total of 2 mL), but the eluted activity decreased drastically over the first 13 elutions as can be seen in
[0101] Since the eluted activity had increased and 90% of the activity was present in the first two fractions, the fraction volume was decreased back to 0.5 mL for elutions 23-25. The eluted activity immediately decreased as was observed before, so elutions 26-34 were again conducted with 1 mL fractions and the eluted activity again increased. As was observed previously, 90-95% of the eluted activity in these elutions were found in the first 2 fractions (first 2 mL) which suggests that a 2 mL volume should be sufficient to extract the majority of the eluted .sup.44Sc, but when the generator is eluted in 40.5 mL fractions (2 mL) this activity decreases drastically. This trend suggests that the elution activity dependence may not be volume driven but rather kinetically driven, since 21 mL fractions elutes 70% more activity than 40.5 mL fractions. Both of these methods use the same volume of 0.5 M HCl to elute, but 1 mL fractions are collected more rapidly than 20.5 mL fractions. To test this the generator was eluted using 21 mL fractions (2 mL total) for elutions 35-47, and the eluted activity remained around 70% as was seen in elutions 26-34 which used 4 mL total volume. .sup.44Ti breakthrough was observed in elution 48 so this generator was abandoned, but this study did prove that a higher activity: resin ratio does increase the amount of activity eluted from the generator (similar to what was observed in the 1 mCi generator) and that kinetics of the elution may impact the eluted activity more than the volume of eluent used. The small-scale generator study that was loaded and eluted in the same solutions only gave 40-45% .sup.44Sc eluted in 4 BV of 0.5 M HCl while this generator eluted 70% in 4 BV of 0.5 M HCl.
[0102] Another AM resin generator of this type was prepared for ongoing studies, but a smaller 5 cm length0.5 cm ID column was used to increase the length of the resin bed by using a column with a smaller inner diameter. This should increase the lifetime of the generator by increasing the path length that the .sup.44Ti must travel before breakthrough is observed. For a 500 L BV, the length of the resin bed in the 10 mL tapered BioRad columns is 1 cm while in the smaller 5 cm columns this length increases to 2 cm. This increased length should increase the lifetime of this generator before .sup.44Ti breakthrough is observed, ideally by double. This strategy of increasing the length of the resin bed by use of a column with a small inner diameter has been used in the literature, where an HPLC type 150 mm2.1 mm ID PEEK column is employed. Future studies using this column will be conducted with the disclosed generator system, which should allow for enhanced optimization and longer lifetimes as the resin bed will be 10 longer than previous generators that used the 10 mL BioRad columns which have a 1 cm ID.
[0103] Using the smaller 5 cm column, a 500 UL BV AM resin (0.200 g) generator was prepared loading 1 mCi of .sup.44Ti in 2 M HCl. This generator was eluted daily with 21 mL fractions of 0.5 M HCl and the elution profile is shown in
[0104] Since both this 1 mCi AM generator and the 720 Ci AM generator are able to elute high activities of .sup.44Sc in small volumes of dilute (0.5 M) HCl, the eluent from these generators were used for direct radiolabeling studies. The results of these studies are summarized in the radiolabeling section. Direct radiolabeling was not possible with any of the other generators mentioned in this study since 2 M HCl is too acidic to be used in this sort of labeling reaction, and the smaller scale generators did not elute high enough activities in small volumes to be of use. This is why the dry down method was used prior to radiolabeling studies with .sup.44Sc eluted from these generators. It is also worth mentioning that none of the ZR generators prepared in this study eluted sufficient activities of .sup.44Sc in appropriate volumes of dilute HCl, so this resin is not conducive to produce .sup.44Sc for use in direct radiolabeling reactions.
Eluent Variability Study
[0105] The use of 0.5 M and 2 M HCl as generator eluent with AM resin has already been verified, but the use of other eluents such as buffers was also of interest. If the generator can be eluted successfully using a mild buffer there would be no need for post-elution processing prior to radiolabeling of the .sup.44Sc, which would be the most ideal for use in clinical studies. In order to be adequate for direct radiolabeling, the .sup.44Sc needs to be eluted in small volumes of slightly acidic (pH 2-7) media without any competitive binding ligands such as oxalates. To test the elution behavior of the AM resin using different eluents, a 300 UL BV 100 Ci .sup.44Ti generator was loaded in 2 M HCl and eluted using varying solvents. Between each change of eluent, the column was flushed with water to ensure no residual solvent was left. Each solvent was tested only a few times to ascertain the viability of its use to elute large quantities of .sup.44Sc in small volumes without .sup.44Ti breakthrough. The generator was eluted with 4 BV of each solvent and the results of the elution are shown in Table 4.
TABLE-US-00004 TABLE 4 Elution Behavior of 100 Ci AM Generator with Varying Eluents Eluent % .sup.44Sc Eluted % .sup.44Ti Breakthrough 0.25M NH.sub.4OAc pH 4 2.1 0 0.25M NH.sub.4OAc pH 4 0.6 0 0.1M Na Citrate pH 4 31.8 0.8 4M HCl 59.6 0 4M HCl 36.8 0 4M HCl 58.0 0 6M HCl 67.2 0 6M HCl 77.1 0 6M HCl 80.7 0
[0106] The use of sodium citrate buffer as an eluent was only attempted once as there was evident .sup.44Ti breakthrough observed, and the use of NH.sub.4OAc buffer was only conducted twice as the activity of .sup.44Sc was negligible. From these results it is evident that the use of a radiolabeling buffer as an eluent is not viable, as the NH.sub.4OAc is not able to efficiently remove the .sup.44Sc, and while the sodium citrate was successful in eluting modest activities of .sup.44Sc, the citrate also appears to be a competitive binder for the .sup.44Ti and leads to breakthrough. Similar to what was seen in the smaller generator studies, by increasing the molarity of HCl eluent the amount of .sup.44Sc eluted is also increased. This is demonstrated by the gradual increase of eluted activity from 0.5, 2, 4 and 6 M HCl. This strongly suggests that pH is a strong contributing factor in the elution of .sup.44Sc from the AM resin. While the percentage of .sup.44Sc eluted in small volumes of 4 and 6 M HCl is high, it is far too acidic to be used in subsequent radiolabeling experiments without diluting or neutralizing (which is difficult at these high molarities), so it is a less ideal option.
Distribution Coefficient (K.SUB.D.) Studies
[0107] K.sub.D values were obtained using the batch contact method. 20 mg of AM resin was prepared in 1.5 mL Eppendorf vials. Stock solutions for each metal were prepared by diluting High Purity ICP standards (100 ppm for Sc and 1000 ppm for Ti, both in 2% HNO.sub.3) in HCl at various concentrations (0.5, 1, 2, 4, 8 and 10.8 M) to a final concentration of 10 ppm. Initial studies were conducted by first drying down the ICP standards and re-dissolving in the appropriate HCl concentration, but the results did not vary statistically from those obtained by the direct dilution method, so it was determined that the small amount of HNO.sub.3 present in the initial metal solutions did not affect or contribute to the resulting distribution coefficients. 1 mL of these standard solutions was then added to the vials containing the resin which were then sealed and mixed by inversion at room temperature for 24 h to allow for equilibration. Following mixing, the tubes were centrifuged and a 500 l aliquot obtained, diluted in 2% HNO.sub.3 and submitted for ICP-OES analysis to determine the concentration of analyte remaining in the aqueous phase. This was compared to the concentration of the initial cold metal standard and the distribution coefficient calculated using the following equation, where C8 and CA are the elemental concentrations of the analyte in solution before and after equilibrium respectively (Equation (1)). Each experiment was done in triplicate at each concentration range.
Total Organic Content
[0108] To determine and compare the amount of organic impurities that are potentially being eluted from the extraction chromatographic ZR resin and the covalently bound synthesized AM resin, small columns containing the resin were prepared and eluted with 0.5 M and 2 M HCl. The eluent was collected and sent for total organic content analysis. The results of the analysis arc shown below, the 0.5 M HCl in
Specific Activity of Generator Produced .SUP.44.Sc
[0109] To determine the specific activity of the AM resin generator produced .sup.44Sc, the eluted fractions from the 1 mCi 2 M HCl eluted generator were collected and analyzed by ICP-OES (Supplementary Materials) to determine the amount of cold Sc in the collected eluent. All four fractions from each elution 1, 5, 10, 15, 20, and 25 were combined, diluted to 8 mL in 2% HNO.sub.3 and each analyzed by ICP-OES. The analyzed Sc peaks from elution 20 had an RSD of 43% so no reliable data was obtained, but all remaining elution samples had RSD values below 10% and the resulting Sc concentration and amount are given in Table 5.
TABLE-US-00005 TABLE 5 Resulting Scandium ICP-OES Data from 1 mCi 2M HCl Generator Elutions Combined Amount of Sc Combined ICP Sample Sc Fractions Sc in Combined Fractions Concentration Concentration Fractions Elution Volume (mL) (ppm) (ppm) (mg) % RSD 1 3.885 0.001 0.0021 0.008 1.03 5 3.195 0.001 0.0025 0.008 1.09 10 3.374 0 0.sup.a 0.sup.a 9.14 15 4.148 0 0.sup.a 0.sup.a 8.11 20 3.129 0 43.41 25 3.633 0.001 0.0022 0.008 2.45 .sup.aBelow detection limit.
[0110] These results demonstrate that there is very little cold scandium in the eluted fractions from the AM resin generator, suggesting that the eluted .sup.44Sc is carrier free, or at the very least that the amount of cold scandium in the elutions is below the detection limit of the ICP-OES.
Stripping of .sup.44Ti from AM Resin
[0111] Stripping of the .sup.44Ti from the AM resin was tested using several different solvents. Ideally the stripping solution should be able to isolate the full amount of .sup.44Ti in relatively small volumes in a form that is not chemically detrimental to the .sup.44Ti. Initially, concentrated HCl and HNO.sub.3 were tested; four bed volumes of each concentrated acid were passed through study 3 generator (300 L BV, 20 Ci) in attempts to remove the .sup.44Ti (Table 6). Remarkably neither acid eluted appreciable amount of .sup.44Ti. This suggests that once the Ti is bound, the AM resin tightly retains Ti- and is robust and fairly resistant to harsh low pH conditions.
TABLE-US-00006 TABLE 6 Stripping Attempts Using 4 BV of Concentrated Acids Acid % .sup.44Sc Eluted % .sup.44Ti Eluted HCl 95.5 0.5 HNO.sub.3 88.0 2.6
[0112] The use of dilute H.sub.2O.sub.2 in HCl has been used to efficiently strip .sup.44Ti from the ZR resin so that was also tested. 2% H.sub.2O.sub.2 in 2 M HCl was used to strip the 1 mCi generator, and 99.8% .sup.44Ti was recovered in 24 BV, although 70% was recovered in the first 8 BV (
Radiolabeling Experiments
[0113] To assess the purity of the eluted .sup.44Sc, radiolabeling experiments were conducted on .sup.44Sc eluted from both the 1 mCi AM generator eluted in 2 M HCl, as well as a 100 Ci 300 L BV ZR generator as direct comparison. All of these radiolabeling studies using .sup.44Sc eluted from these generators were conducted using the dry down method mentioned previously. Both of these generators were eluted using 4 BV of 2 M HCl, and a portion of the eluent was evaporated to dryness at 140 C. on a hotplate, and then re-dissolved in 0.25 M NH.sub.4OAc pH 4 buffer in order to be radiolabeled. Both eluents were processed and radiolabeled in the exact same manner to ensure a direct comparison between the two different generator-produced .sup.44Sc samples. The radiolabeling experiments were done using three different chelators (DOTA, NOTA and HOPO), but in each case the experiment conditions were the same. The appropriate amount of ligand was added to the .sup.44Sc solution and heated at 90 C. for one hour in a thermomixer. The radiolabeling yield was then determined using radio TLC conditions as described above. The DOTA and HOPO reactions were done using a 50,000 ligand molar excess, and the NOTA using a 100,000 ligand molar excess due to the lower thermodynamic stability of Sc-NOTA complexes. Each radiolabeling experiment was conducted in triplicate using different generator eluents (done on different days) and the TLCs were also each done in triplicate to ensure consistent results. The results of these radiolabeling experiments are summarized in Table 7.
TABLE-US-00007 TABLE 7 Radiolabeling Yields Determined by RadioTLC AM ZR DOTA 98.75% 0.68 38.19% 7.74 NOTA 97.42% 0.59 10.71% 12.19 HOPO 86.86% 7.43 38.77% 12.61
[0114] The error associated with the ZR generator experiments is slightly higher due to the lower activity used, and the separation of peaks in the HOPO and NOTA experiments were not as clean as with DOTA leading to slightly higher errors in those experiments. Overall, it is clear that the AM generator produced .sup.44Sc labeled much better than the ZR .sup.44Sc under the same conditions, suggesting that it is of higher purity. It is worth noting that the ZR .sup.44Sc solution after processing is yellow in color, where the AM solution is completely colorless. This color is likely due to an organic-based impurity, possibly from the ZR resin, as the .sup.44Sc or radiolabeled complexes should be completely colorless. The characterization of this impurity is currently unknown but seems to hinder the radiolabeling of the ZR generator eluted radioisotope.
[0115] The .sup.44Sc eluent from the 720 Ci and 1 mCi generators that were loaded in 2 M HCl and eluted in 0.5 M HCl were used in direct radiolabeling experiments since the eluent was in a form conducive to this method of labeling. The .sup.44Sc generator eluent in 0.5 M HCl was initially neutralized with equal amounts of 0.5 M NaOH and then added to the appropriate amount of ligand stock solution in 0.25 M NH.sub.4OAc pH 4 buffer. This solution had an overall pH of 4. This solution was then heated at 90 C. for 1 hour in a thermomixer and the radiolabeling yield was then determined using radio TLC conditions as described previously. An initial experiment was conducted with 50,000 molar equivalents of DOTA (similar to what was conducted in the dry down method) and the radioyield was 99.46%+0.05%. This excess was then decreased to 25,000 molar equivalents of DOTA and the resulting radioyield remained high at 99.28%+0.08%. This was then decreased further to 10,000 molar equivalents of DOTA, and the average radioyield from four separate reactions was 85.10%+10.57%. A comparison labeling study was done with 10,000 molar equivalents of DOTA using the dry down method which gave a radioyield of 9.36%+0.16%. The direct radiolabeling method not only allows for more quick and facile labeling studies as post-elution processing is not needed, but it also showed drastically higher yields at lower ligand amounts compared to the dry down method. Further studies are needed to optimize these labeling studies, but the use of AM resin generator produced .sup.44Sc obtained in 0.5 M HCl shows extremely promising direct radiolabeling results which is currently not possible with .sup.44Ti/.sup.44Sc generators in the literature.
[0116] One of the biggest drawbacks to currently employed .sup.44Sc generators is that the eluent needs post-elution purification in order to use the .sup.44Sc in subsequent radiolabeling reactions. This is due to the presence of competing chelators in the eluent such as the oxalates present in oxalic acid. To overcome this, current methods employ the use of a second column to purify the .sup.44Sc by removal of the oxalates and isolation of the .sup.44Sc in a radiolabeling conducive buffer. To avoid post-elution processing of the generator eluent and allow for direct radio labeling of the .sup.44Sc several criteria need to be met: the eluent must be free of competing chelators, the eluent must not be highly acidic, and the volume: activity ratio must be low in order to keep radiolabeling reaction total volumes reasonable. The generator presented in this work meets all of these criteria and should allow for use of the eluted .sup.44Sc directly from the generator without the need of time-consuming post-elution processing and loss of activity.
[0117] To test this, preliminary radiolabeling studies were conducted on the generator eluent from the 37 MBq (1 mCi) generator eluted with 0.5 M HCl with DOTA under standard radiolabeling conditions. The .sup.44Sc was obtained in 0.5 M HCl and then brought to a pH of 4 with the addition of 0.5 M NaOH. This solution was then added directly to an Eppendorf vial containing DOTA stock solution in 0.25 M NH.sub.4OAc at a pH of 4, with a resulting pH of 4 for the reaction solution. The solution was heated at 90 C. for 60 min and the radioyield determined by iTLC. This reaction was conducted using 925 kBq (25 Ci) of .sup.44Sc and 1.59 nmol of DOTA resulting in a 99.240.21% yield, which is consistent with other generator produced .sup.44Sc samples that have been processed using chromatography post-elution. More studies are needed to further optimize and study the radiolabeling efficiencies of this generator produced .sup.44Sc, but these preliminary results show extremely promising evidence for the ability to directly radiolabel .sup.44Sc from this generator system.
Example 3: Alternative Radionuclide Systems
[0118] In another embodiment, the present generator may be a .sup.172Hf/.sup.172Lu generator system comprising the use of an alkyl-substituted hydroxamate resin and a method of producing .sup.172Lu comprising decay of .sup.172Hf in a .sup.172Hf/.sup.172Lu generator system using an alkyl-substituted hydroxamate resin.
[0119] .sup.172Hf has a half life of 1.87 years and decays by electron capture (EC) to .sup.172Lu (t.sub.1/2=6.7 days) which then decays by electron capture to stable .sup.172Yb (
[0120] In the case that Hf is most stable in the (IV) oxidation state and Lu is only stable in the (III) oxidation state, this is similar to Ti(IV) and Sc(III). Although the ionic radii of Hf and Lu are larger than that of Ti and Sc, they possess similarities in bonding, so the hydroxamate groups of the resin will likely selectively bind Hf over Lu and allow for retention of .sup.172Hf on the resin matrix and elution of .sup.172Lu.
[0121] Preliminary cold studies were conducted using the methyl hydroxamate resin to ascertain the distribution coefficient (K.sub.D) for both Hf and Lu in varying concentrations of HCl to determine the extent of this generator system (
[0122] It should be emphasized that the above-described embodiments of the present disclosure are merely possible examples of implementations set forth for a clear understanding of the principles of the disclosure. Many variations and modifications may be made to the above-described embodiment(s) without departing substantially from the spirit and principles of the disclosure. All such modifications and variations are intended to be included herein within the scope of this disclosure and protected by the following claims.
REFERENCES
[0123] 1. Agrawal, Y. K., et al, 1999. Poly (styrene-p-hydroxamic acids): synthesis, and ion exchange separation of rare earths. React. Funct. Polym. 39. 155-164. [0124] 2. Antimicrobial Resistance C. Global burden of bacterial antimicrobial resistance in 2019: a systematic analysis. Lancet 2022; 399 (10325): 629-655. [0125] 3. Bae T et al. Generating a collection of insertion mutations in the Staphylococcus aureus genome using bursa aurealis. Methods Mol Biol 2008; 416:103-116. [0126] 4. Bafaro E et al. The emerging role of zinc transporters in cellular homeostasis and cancer. Signal Transduct Target Ther 2017; 2. [0127] 5. Bilinskaya A et al. Variability in Zinc Concentration among Mueller-Hinton Broth Brands: Impact on Antimicrobial Susceptibility Testing of Metallo-beta-Lactamase-Producing Enterobacteriaceae. J Clin Microbiol 2020; 58 (12). [0128] 6. Bohlmann L et al. Chemical Synergy between Ionophore PBT2 and Zinc Reverses Antibiotic Resistance. mBio 2018; 9 (6). [0129] 7. Brandenburg K S et al. Novel murine model for delayed wound healing using a biological wound dressing with Pseudomonas aeruginosa biofilms. Microb Pathog 2018; 122:30-38. [0130] 8. Brazel E B et al. Dysregulation of Streptococcus pneumoniae zinc homeostasis breaks ampicillin resistance in a pneumonia infection model. Cell Rep 2022; 38 (2): 110202. [0131] 9. Brennan, B. J., et al, 2016. Molecular titanium-hydroxamate complexes as models for TiO.sub.2 surface binding. Chem. Commun. 52, 2972-2975. [0132] 10. Brown E D et al. Antibacterial drug discovery in the resistance era. Nature 2016; 529 (7586): 336-343. [0133] 11. Cornaz, J. P., et al, 1954. Selektive lonenaustauscher fur Fe3+ 1. Experientia 10, 137-138. [0134] 12. Cornaz. J.-P., et al. 1957. S3urechloride und Hydroxamsauren von Carboxyl-lonenaustauschem. 10. Mitteilung Uber lonenaustauscher. Helv. Chim. Acta 40, 2015-2019. [0135] 13. Crawford C L et al. A copper-dependent compound restores ampicillin sensitivity in multidrug-resistant Staphylococcus aureus. Sci Rep 2020; 10 (1): 8955. [0136] 14. Crawford C L et al. Pyrazolopyrimidinones, a novel class of copper-dependent bactericidal antibiotics against multi-drug resistant S. aureus. Metallomics 2019; 11 (4): 784-798. [0137] 15. Dalecki A G et al. Combinatorial phenotypic screen uncovers unrecognized family of extended thiourea inhibitors with copper-dependent anti-staphylococcal activity. Metallomics 2016; 8 (4): 412-421. [0138] 16. Dalecki A G et al. Targeting Biofilm Associated Staphylococcus aureus Using Resazurin Based Drug-susceptibility Assay. J Vis Exp 2016; (111). [0139] 17. Davidson, D., 1940. Hydroxamic acids in qualitative organic analysis. J. Chem. Educ. 17, 81. [0140] 18. De Oliveira D M P et al. Rescuing Tetracycline Class Antibiotics for the Treatment of Multidrug-Resistant Acinetobacter baumannii Pulmonary Infection. mBio 2022: e0351721. [0141] 19. Dirks, C. B. A., et al, 2015. On the Development and Characterisa tion of an Hydroxamate Based Extraction Chromatographic Resin. 6 1st Radiobioassay and Radiochemistry Measurements Conference. Iowa City, IA, USA. [0142] 20. Domnanich, K. A., et al, 201 7. (44)Sc for labeling of DOTA- and NODAGA-functionalized peptides: preclinical in vitro and in vivo investigations. EJNMMI Radiopharm Chem 1, 8. [0143] 21. Eigner, S., et al, 2013. Imaging of protein synthesis: in vitro and in vivo evaluation of 44Sc-DOTA-Puromycin. Mal. Imag. Biol. 15, 79-86. [0144] 22. Eskenazi A et al. Combination of pre-adapted bacteriophage therapy and antibiotics for treatment of fracture-related infection due to pandrug-resistant Klebsiella pneumoniae. Nat Commun 2022; 13 (1): 302. [0145] 23. Fey P D et al. A genetic resource for rapid and comprehensive phenotype screening of nonessential Staphylococcus aureus genes. mBio 2013; 4 (1): e00537-00512. [0146] 24. Filosofov, D. V., et al, 2010. A 44TV44Sc radionuclide generator for potential application of 44Sc-based PET-radiopharmaceuticals. rca-Radiochimica Acta 98, 149-156. [0147] 25. Gao W et al. Nanomaterials arising amid antibiotic resistance. Nat Rev Microbiol 2021; 19 (1): 5-6. [0148] 26. Hemansez, R., et al, 2014. 44Sc: an attractive isotope for peptide-based PET imaging. Mal. Pharm. 11, 2954-2961. [0149] 27. Herscheid, J. D., et al, 1983. Manganese-52m for direct application: a new 52Fe/52mMn generator based on a hydroxamate resin. Int. J. Appl. Radiat. Isot. 34, 883-886. [0150] 28. Higgins, F. S., et al, 2006. Infrared and Raman spectroscopy study of alkyl hydroxamic acid and alkyl hydroxamate isomers. Appl. Spectrosc. 60, 279-287. [0151] 29. Holland, J. P., et al, 2009. Standardized methods for the production of high specific-activity zirconium-89. Nucl. Med. Biol. 36, 729-739. [0152] 30. Honarvar, H., et al, 2017. Evaluation of the first 44Sc-labeled Affibody molecule for imaging of HER2-expressing tumors. Nucl. Med. Biol. 45, 15-21. [0153] 31. Hood M I et al. Nutritional immunity: transition metals at the pathogen-host interface. Nat Rev Microbiol 2012; 10 (8): 525-537. [0154] 32. Johnson B A et al. Bacitracin: A New Antibiotic Produced by a Member of the B. Subtilis Group. Science 1945; 102 (2650): 376-377. [0155] 33. Jones, K. E., et al, 2017. Ti(IV) and the siderophore desferrioxamine B: a tight complex has biological and environmental implications. Inorg. Chem. 56, 1264-1272. [0156] 34. Kehl-Fie T E et al. Nutritional immunity beyond iron: a role for manganese and zinc. Curr Opin Chem Biol 2010; 14 (2): 218-224. [0157] 35. Kennedy A D et al. Epidemic community-associated methicillin-resistant Staphylococcus aureus: recent clonal expansion and diversification. Proc Natl Acad Sci USA 2008; 105 (4): 1327-1332. [0158] 36. Kennedy A D, et al. Complete nucleotide sequence analysis of plasmids in strains of Staphylococcus aureus clone USA300 reveals a high level of identity among isolates with closely related core genome sequences. J Clin Microbiol 2010; 48 (12): 4504-4511. [0159] 37. Kerdjoudj, R., et al, 2016. Scandium (iii) complexes of monophosphorus acid DOTA analogues: a thermodynamic and radiolabelling study with 44Sc from cyclotron and from a 44Ti/44Sc generator. Dalton Trans. 45, 1398-1409. [0160] 38. Khodadadi, R., et al, 1995. Poly (hydroxamic Acid) Chelating Resin: the Synthesis and Uses. Iran. J. Polymer. Sci. Tech. 4, 248-255. [0161] 39. Koumarianou, E., et al, 201 2. 44Sc-DOTA-BN[2-14]NH2 in comparison to 6 8Ga-DOTA-BN[2-14]NH2 in pre-clinical investigation. Is 44Sc a potential radionuclide for PET? Appl. Radiat.! sot. 70, 2669-2676. [0162] 40. Larenkov, A. A., et al, 2021. Separation of 44Sc from 44Ti in the context of A generator system for radiopharmaceutical purposes with the example of [44Sc]Sc-PSMA-6 17 and [44Sc]Sc-PSMA-I& T synthesis. Molecules 26, 6371. [0163] 41. Lelievre P et al. The Multifaceted Roles of Copper in Cancer: A Trace Metal Element with Dysregulated Metabolism, but Also a Target or a Bullet for Therapy. Cancers (Basel) 2020; 12 (12). [0164] 42. Lewis K. Antibiotics: Recover the lost art of drug discovery. Nature 2012; 485 (7399): 439-440. [0165] 43. Ling L L et al. A new antibiotic kills pathogens without detectable resistance. Nature 2015; 517 (7535): 455-459. [0166] 44. Liu, C. Y., et al, 1992. Synthesis and coordination behavior of hydroxamate resin with varying spacer groups. Polyhedron 11, 551-558. [0167] 45. Mawji, E., 2008. Hydroxamate siderophores: occurrence and importance in the Atlantic ocean. Environ. Sci. Technol. 42, 8675-8680. [0168] 46. Meijs, W. E., et al, 1994. Production of highly pure no-carrier added 89Zr for the labelling of antibodies with a positron emitter. Appl. Radiat. Isot. 45, 1143-1147. [0169] 47. Ming L J et al. Metal binding and structure-activity relationship of the metalloantibiotic peptide bacitracin. J Inorg Biochem 2002; 91 (1): 46-58. [0170] 48. Muller, C. et al. 2013. Promises of cyclotron-produced 44Sc as a diagnostic match for trivalent -emitters: in vitro and in vivo study of a 44Sc-DOTA-Folate conjugate. J. Nucl. Med. 54, 2168-2174. [0171] 49. Orlowska, E., et al, 2016. Benzoic hydroxamate-based iron complexes as model compounds for humic substances: synthesis, characterization and algal growth experiments. RSC Adv. 6, 40238-40249. [0172] 50. Padial, N. M., et al, 2019. Hydroxamate titanium-organic frameworks and the effect of siderophore-type linkers over their photocatalytic activity. J. Am. Chem. Soc. 141, 13124-13133. [0173] 51. Passadis, S. S., Hadjithoma, S., Siafarika, P., Kalampounias, A. G., Keramidas, A. O., Miras, H. N., Kabanos, T. A., 2021. Synthesis, structural and physicochemical characterization of a titanium (IV) compo und with the hydroxamate ligand N,2-Dihydroxybenzamide. Molecules 26, 5588. [0174] 52. Petrie, G., Locke, D., Meloan, C. E., 1965. Hydroxamic acid chelate ion exchange resin. Anal. Chem. 37, 919-&. [0175] 53. Phillips, R. J., Fritz, J. S., 1980. Synthesis and analytical properties of an n-phenylhydroxamic acid resin. Anal. Chim. Acta 121, 225-232. [0176] 54. Phillips, R. J., 1980. Chelating ion exchange with macroreticular hydroxamic acid resins. Doctor of Philosophy, Iowa State University. [0177] 55. Phillips, R. J., Fritz, J. S., 1982. Extraction of metal ions by N-phenyl-, N-methyl-, and Nunsubstituted hydroxamic acid resins. Anal. Chim. Acta 139, 237-246. [0178] 56. Pruszyn ki, M., Loktionova, N. S., Filosofov, D. V., Reisch, F., 2010. Post-elution processing of 44Ti/44Sc generator-derived 44Sc for clinical application. Appl. Radiat.! sot. 68, 1636-1641. [0179] 57. Radchenko, V., Meyer, C. A. L., Engle, J. W., Naranjo, C. M., Unc, G. A., Mastren, T., Brugh, M., Birnbaum, E. R., John, K. D., Nortier, F. M., Fassbender, M. E., 2016. Separation of 44Ti from proton irradiated scandium by using solid-phase extraction chromatography and design of 44Ti/44Sc generator system. J. Chromatogr. A 1477, 39-46. [0180] 58. Radford R J et al. Chelators for investigating zinc metalloneurochemistry. Curr Opin Chem Biol 2013; 17 (2): 129-136. [0181] 59. Ramchandani D et al. Copper depletion modulates mitochondrial oxidative phosphorylation to impair triple negative breast cancer metastasis. Nat Commun 2021; 12 (1): 7311. [0182] 60. Reddy, A. S., Kumar, M. S., Reddy, G. R., 2000. A convenient method for the preparation of hydroxamic acids. Tetrahedron Lett. 41, 6285-6288. [0183] 61. Renfrow, W. B., Hauser, C. R., 1937. The relative rates of decomposition of the potassium salts of certain meta and para substituted dibenzhydroxamic acids. A study of the lossen Rearrangement, J. Am. Chem. Soc.!: >9, :I3U8-:I314. [0184] 62. Roesch, F., 2012. Scandium-44: benefits of a long-lived PET radionuclide available from the (44)Ti/(44)Sc generator system. Curr. Rad. 5, 187-201. [0185] 63. Saxena, M., Loza-Rosas, S. A., Gaur, K., Sharma, S., Perez Otero, S. C., Tinoco, A. D., 2018. Exploring titanium ([V) chemical proximity to iron (HI) to elucidate a function for Ti(IV) in the human body. Coord. Chem. Rev. 363, 109-125. [0186] 64. Shukla R et al. Mode of action of teixobactins in cellular membranes. Nat Commun 2020; 11 (1): 2848. [0187] 65. Silver L L. Challenges of antibacterial discovery. Clin Microbiol Rev 2011; 24 (1): 71-109. [0188] 66. Speer A et al. Copper-boosting compounds: a novel concept for antimycobacterial drug discovery. Antimicrob Agents Chemother 2013; 57 (2): 1089-1091. [0189] 67. Szaniszlo, P. J., Powell, P. E., Reid, C. P. P., Cline, G. R., 1981. Production of hydroxamate siderophore iron chelators by ectomycorrhizal Fungi. Mycologia 73, 1 158-1174. [0190] 68. Wang F et al. Turning tumor-promoting copper into an anti-cancer weapon via high-throughput chemistry. Curr Med Chem 2010; 17 (25): 2685-2698. [0191] 69. Wong, P. T., Bhattacharjee, S., Cannon, J., Tang, S., Yang, K., Bowden, S., Vamau, V., O'Konek, J. J., Choi, S. K., 2019. Reactivity and mechanism of a-nucleophile scaffolds as catalytic organophosphate scavengers. Org. Biomol. Chem. 17, 3951-3963.