Patent Application
20240199882
This invention relates generally to the field of dye materials and, more particularly, to novel shortwave infrared (SWIR) dye materials and compositions that absorb light at wavelengths of 900 nm to 1400 nm, which is ideal for deep tissue imaging owing to minimized light scattering and interference from endogenous pigments when using light in this wavelength range. An approach to access such molecules is to tune the photophysical properties of known near-infrared dyes. A series of easily accessible (three steps) SWIR xanthene dyes has been developed based on a dibenzazepine donor conjugated to thiophene (SCR-1), thienothiophene (SCR-2), or bithiophene (SCR-3). Since SCR-1 undergoes a bathochromic shift when aggregated for in vivo studies, we developed a ratiometric nanoparticle for nitric acid (NO) (rNP-NO), which was employed to visualize pathological levels of nitric oxide in a drug-induced liver injury model via deep tissue SWIR photoacoustic (PA) imaging. This dye series can be utilized as a component in nanosensor designs for imaging studies.
The “biological window” is traditionally considered to span light wavelengths of 650 nm to 900 nm in the near infrared (NIR), a spectral region where interference due to auto-fluorescence is minimal compared to ultraviolet (UV) and visible light. Moreover, less optical absorbers are present that may intercept and scatter light wavelengths of 650 nm to 900 nm, allowing for deeper tissue imaging. These unique properties of NIR light have enabled a variety of applications including photodynamic therapy, image-guided surgery, and photothermal ablation. In vivo molecular imaging via fluorescence and photoacoustic (PA) modes are additional applications that have attracted significant interest owing to their ability to detect biomarkers via activity-based sensing. However, a paradigm shift is currently underway whereby the preferred incident light window employs even longer wavelengths. This so-called shortwave infrared (SWIR) window includes light from 900 nm to 1400 nm, which overlaps less with interfering endogenous pigments like hemoglobin found in blood or melanin in skin. In the context of PA imaging, the use of SWIR imaging agents enables depths (˜10 cm) several times greater than comparable NIR systems.
PA imaging is a modality that relies on the use of light to stimulate the production of acoustic waves (known as the “PA effect”). After a chromophore is excited by a pulsed laser, a portion of the absorbed energy is released as heat, which causes rapid thermoelastic expansion within the sample being imaged. This increase in temperature leads to pressure fluctuations that generate a detectable ultrasound signal, which can be reconstructed to create high-resolution 3D images of the region of interest. The ability of SWIR light to penetrate far into the body results in a more robust PA response in deep tissue. This improvement is necessary for biomedical applications such as the assessment of liver injury from drug overdose. However, most small-molecule dyes operate in the far red to NIR window with only a handful of known SWIR dyes, presumably because they are difficult to prepare, exhibit low chemical stability, and are difficult to formulate (poor solubility or hypsochromic shift) for in vivo studies.
Of the existing SWIR-absorbing dyes, most are based on the polymethine structure (e.g., commercially available IR-1061); however, examples from the aza-BODIPY, benzo[1,2-c:4,5-c]bis([1,2,5]-thiadiazole) (BBTD) core, and xanthene families are also known.
There is a need to improve the case of formation, case of modification and the physical properties of tSWIR-absorbing dyes.
The present xanthene based dyes fulfill this need. The wavelength of maximum absorbance of xanthene dyes is moved into the SWIR window using thiophene-dibenzazepine type donor ligands. These xanthene dyes are easy to make, can be readily modified, and exhibit remarkable photophysical properties, i.e., the xanthene dyes can be used for PA imaging, and have high molar absorptivity, low quantum yields, and SWIR absorbance.
A series of three xanthene-based SWIR dyes have been synthesized using a donor-acceptor-donor (D-A-D) design. One example (SCR-1) has been used to prepare a nitric oxide (NO)-responsive probe. The SCR-1 dye has been co-encapsulated together with a second SWIR dye (non-responsive reference) within a biocompatible polymer matrix to generate a robust nanosensor, which can detect NO in deep tissue using a drug-induced liver injury model via ratiometric SWIR PA imaging.
In an aspect, the disclosure relates to a shortwave infrared (SWIR) dye comprising a counterion and a structure of Formula I
wherein X is selected from O, Si and P;
R1 is hydrogen and R2 is a donor which is substituted or unsubstituted and is selected from a dibenzazepinyl, a thienyldibenzazepine, a bithienyldibenzazepine, a thienodibenzazepine, a dihydrodibenzazepinyl, a thienyldihydrodibenzazepine, a bithienyldihydrodibenzazepine, and a thienodihydrodibenzazepine;
Both R1 and R2 are the same donors, are substituted or unsubstituted, and are selected from a dibenzazepinyl, a thienyldibenzazepine, a bithienyldibenzazepine, a thienodibenzazepine, a dihydrodibenzazepinyl, a thienyldihydrodibenzazepine, a bithienyldihydrodibenzazepine, and a thienodihydrodibenzazepine; and
R1 and R2 are different donors and R1 is a donor, which is substituted or unsubstituted and is selected from a dibenzazepinyl, a thienyldibenzazepine, a bithienyldibenzazepine, a thienodibenzazepine, a dihydrodibenzazepinyl, a thienyldihydrodibenzazepine, a bithienyldihydrodibenzazepine, and a thienodihydrodibenzazepine and R2 is a donor which is substituted or unsubstituted and is selected from a dibenzazepinyl, a thienyldibenzazepine, a bithienyldibenzazepine, a thienodibenzazepine, a dihydrodibenzazepinyl, a thienyldihydrodibenzazepine, a bithienyldihydrodibenzazepine, a thienodihydrodibenzazepine, 1-(thiophen-2-yl)piperidine, 1-(thieno[3,2-b]thiophen-2-yl)piperidine, 1-([2,2′-bithiophen]-5-yl)piperidine, C2-C12 dialkyl amino, indolizine-3-yl, diphenylamino, and julolidinyl. In Group C, the dyes are unsymmetrical. For instance, the donor at R1 could be thienyldibenzazepine and at R2 could be bithienyldibenzazepine; or both R1 and R2 can be thienyldibenzazepine but one of R1 and R2 is substituted and the other is not.
In the foregoing embodiment, X can be O, and R3 and R4 can be absent;
In each of the foregoing embodiments, X can be O, and R3 and R4 can be absent;
R1 is hydrogen and R2 is a donor which is substituted with 0 to 3 substituents and is selected from a dibenzazepinyl, a thienyldibenzazepine, a bithienyldibenzazepine, a thienodibenzazepine, a dihydrodibenzazepinyl, a thienyldihydrodibenzazepine, a bithienyldihydrodibenzazepine, and a thienodihydrodibenzazepine; or
Both R1 and R2 are the same donors and are substituted with 0 to 3 substituents, and can be selected from dibenzazepinyl, thienyldibenzazepine, bithienyldibenzazepine, thienodibenzazepine, dihydrodibenzazepinyl, thienyldihydrodibenzazepine, bithienyldihydrodibenzazepine, or thienodihydrodibenzazepine;
In each of the foregoing embodiments, X can be O, and R3 and R4 can be absent;
R1 and R2 are different donors and R1 is a donor which is substituted with 0 to 3 substituents and is selected from a dibenzazepinyl, a thienyldibenzazepine, a bithienyldibenzazepine, a thienodibenzazepine, a dihydrodibenzazepinyl, a thienyldihydrodibenzazepine, a bithienyldihydrodibenzazepine, and a thienodihydrodibenzazepine and R2 is a donor which is substituted with 0 to 3 substituents, and is selected from a dibenzazepinyl, a thienyldibenzazepine, a bithienyldibenzazepine, a thienodibenzazepine, a dihydrodibenzazepinyl, a thienyldihydrodibenzazepine, a bithienyldihydrodibenzazepine, a thienodihydrodibenzazepine, 1-(thiophen-2-yl)piperidine, 1-(thieno[3,2-b]thiophen-2-yl)piperidine, 1-([2,2′-bithiophen]-5-yl)piperidine, C2-C12 dialkyl amino, indolizine-3-yl, diphenylamino, and julolidinyl;
In each of the foregoing embodiments, X can be O, and R3 and R4 can be absent;
R1 and R2 are different donors and R1 is a donor which is substituted with 0 to 3 substituents and selected from a dibenzazepinyl, a thienyldibenzazepine, a bithienyldibenzazepine, a thienodibenzazepine, a dihydrodibenzazepinyl, a thienyldihydrodibenzazepine, a bithienyldihydrodibenzazepine, and a thienodihydrodibenzazepine and R2 is a donor which is substituted with 0 to 3 substituents and is selected from a dibenzazepinyl, a thienyldibenzazepine, a bithienyldibenzazepine, a thienodibenzazepine, a dihydrodibenzazepinyl, a thienyldihydrodibenzazepine, a bithienyldihydrodibenzazepine, a thienodihydrodibenzazepine, 1-(thiophen-2-yl)piperidine, 1-(thieno[3,2-b]thiophen-2-yl)piperidine, 1-([2,2′-bithiophen]-5-yl)piperidine, diethyl amino, and julolidinyl;
In each of the foregoing embodiments, the counterion is selected from nitrite, sulfate, phosphate, bicarbonate, trifluoroacetate, pentafluoropropanoate, chloride, bromide, iodide, perchlorate, nitrate, benzenesulfonate, p-toluenesulfonate, methylsulfate, ethylsulfate, propylsulfate, tetrafluoroborate, tetraphenylborate, hexafluorophosphate, benzenesulfinate, acetate, trifluoroacetate, propionacetate, benzoate, oxalate, succinate, malonate, oleate, stearate, citrate, monohydrogen diphosphate, dihydrogen monophosphate, pentachlorostannate, chlorosulfonate, fluorosulfonate, trifluoromethansulfonate, hexafluoroarsenate, hexafluoroantimonate, molybdenite, tungstate, titanate, zirconate ions, and any combination thereof.
In each of the foregoing embodiments, the counterion can be selected from trifluoroacetate, pentafluoropropanoate, chloride, bromide, iodide, fluorosulfonate, and trifluoromethansulfonate.
In each of the foregoing embodiments, R19, R20, R22, R23, R24, R25, R26, R27, R28, and R29 can be each independently selected from hydrogen, sulfonate, halogen, hydroxy, amino, nitro, cyano, carboxy, an alkyl group having 1 to 6 carbon atoms, an alkenyl group having 2 to 6 carbon atoms, an alkylether group having 2 to 20 carbon atoms and 1 to 5 oxygen atoms, an aryl group having 6 to 10 carbon atoms, and a heterocyclic group having 6 to 10 carbon atoms, and wherein 0 or 1 pair of R22 and R23, R24 and R25, R25 and R26, R26 and R27, R27 and R28, R28 and R29, together with the carbons to which they are attached can form a saturated or unsaturated six membered ring.
In each of the foregoing embodiments, R19, R20, R22, R23, R24, R25, R26, R27, R28, and R29 can be each independently selected from hydrogen, sulfonate, halogen, hydroxy, an alkyl group having 1 to 6 carbon atoms, and a phenyl group.
In each of the foregoing embodiments, R19, R20, R22, R23, R24, R25, R26, R27, R28, and R29 can be hydrogen.
In each of the foregoing embodiments, the SWIR dye can have one of the following structures:
In each of the foregoing embodiments, the SWIR dye can absorb light having a wavelength of 800 nm to 1400 nm.
In another aspect, the disclosure relates to a composite comprising the SWIR dye of any one of foregoing embodiments in a polymer matrix. Preferably, the polymer matrix is solid at room temperature (20° C.).
In another aspect, the disclosure relates to a composite comprising the SWIR dye of any one of the foregoing embodiments encapsulated in a phospholipid-polymer conjugate. In this embodiment, the phospholipid-polymer conjugate may be 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-poly(ethylene glycol) (DSPE-PEG).
Yet another aspect of the disclosure is a method for making a SWIR dye, the method comprising:
wherein R18 and R21 are independently selected from Cl, Br, I and OSO2R52, wherein R52 is a hydrogen or C1-C4 alkyl;
Both R1 and R2 are the same donors, are substituted or unsubstituted, and are selected from a dibenzazepinyl, a thienyldibenzazepine, a bithienyldibenzazepine, a thienodibenzazepine, a dihydrodibenzazepinyl, a thienyldihydrodibenzazepine, a bithienyldihydrodibenzazepine, and a thienodihydrodibenzazepine; and
R1 and R2 are different donors and R1 is a donor which is substituted or unsubstituted and is selected from a dibenzazepinyl, a thienyldibenzazepine, a bithienyldibenzazepine, a thienodibenzazepine, a dihydrodibenzazepinyl, a thienyldihydrodibenzazepine, a bithienyldihydrodibenzazepine, and a thienodihydrodibenzazepine and R2 is a donor which is substituted or unsubstituted and is selected from a dibenzazepinyl, a thienyldibenzazepine, a bithienyldibenzazepine, a thienodibenzazepine, a dihydrodibenzazepinyl, a thienyldihydrodibenzazepine, a bithienyldihydrodibenzazepine, a thienodihydrodibenzazepine, 1-(thiophen-2-yl)piperidine, 1-(thieno[3,2-b]thiophen-2-yl)piperidine, 1-([2,2′-bithiophen]-5-yl)piperidine, C2-C12 dialkyl amino, indolizine-3-yl, diphenylamino, and julolidinyl. The Group C dyes are unsymmetrical. For instance, the donor at R1 could be a thienyldibenzazepine and at R2 the donor could be a thienodibenzazepine; or both R1 and R2 can be thienyldibenzazepine but one is substituted and the other is not.
In the foregoing embodiment, the catalyst in step (a) can be a palladium compound.
In each of the foregoing embodiments, the compound of Formula II can be as follows:
In each of the foregoing embodiments, the compound formed in step (a) can have the following structure:
In yet another aspect, the disclosure relates to a composition comprising the SWIR dyes of any one of the foregoing embodiments and a pharmaceutically acceptable carrier or a solid polymer matrix.
In yet another aspect, the disclosure relates to a method for imaging a biological sample, the method comprising:
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. The transitional phrase “consisting essentially of” limits the scope of a claim to the specified materials or steps and those that do not materially affect the basic and novel characteristic(s) of the claimed invention.
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 solvent,” “a linear alkyl group,” or “an alcohol,” include, but are not limited to, mixtures or combinations of two or more such solvents, linear alkyl groups, or alcohols, and the like.
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.
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’ to less than ‘y’.
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.
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, ± a 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.
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 SWIR dye for imaging of a biological sample refers to an amount that is sufficient to achieve the desired image quality. The specific level in terms of wt % or mol % in a composition required as an effective amount will depend upon a variety of factors including the absorption maxima of the dye, whether the biological sample is an isolated sample or is part of an organism in vivo, the identity of any pharmaceutically acceptable carrier, and the capabilities of the device used to measure the photoacoustic signal produced by the dye via non-radiative decay.
As used herein, the term “donor” refers to an electron donor and the term “acceptor” refers to an electron acceptor in a donor-acceptor or donor-acceptor-donor molecule or dye. The terms “donor” and “acceptor” may be used to refer to the radicals in a donor-acceptor molecule or dye or may be used to refer to separate chemical compounds that will form a donor-acceptor molecule once reacted together.
As used herein, the terms “optional” or “optionally” mean 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.
Unless otherwise specified, pressures referred to herein are based on atmospheric pressure (i.e. one atmosphere).
“SWIR” as used herein refers to the region of the electromagnetic spectrum having wavelengths of from about 800 nm to about 1400 nm, or from about 900 nm to about 1400 nm. In one aspect, the compounds disclosed herein emit fluorescence and/or absorb radiation in the SWIR region.
As used herein, “fluorescence quantum yield” (ϕ) refers to the ratio of photons absorbed to photons emitted through fluorescence.
“Molar absorptivity,” “molar absorption coefficient,” and “extinction coefficient” refer to how strongly a chemical compound absorbs light at a given wavelength. Molar absorptivity is an intrinsic property of the compound; however, this coefficient varies with wavelength and solvent. Molar absorptivity is typically expressed in terms of absorption at a particular wavelength, such as the maximum position in the absorption band. Units are typically given as L/mol·cm or M−1·cm−1. In one aspect, the disclosed SWIR dyes have a high & in the SWIR spectral region.
In one aspect, the SWIR dye has an absorption maximum in dimethyl sulfoxide (DMSO) at from about 800 nm to about 1400 nm, or about 810 nm to about 1200 nm, or from about 830 nm to about 1100 nm.
In another aspect, the SWIR dye has a molar absorption coefficient of about 70,000 M−1·cm−1 or greater, or of about 75,500 M−1·cm−1, 76,000 M−1·cm−1, 76,500 M−1·cm−1, 77,000 M−1·cm−1, 77,500 M−1·cm−1, 78,000 M−1·cm−1, 78,500 M−1·cm−1, or about 78,900 M−1·cm−1 or greater; or up to about 140,500 M−1·cm−1, 138,000 M−1·cm−1, 130,500 M−1·cm−1, or about 122,900 M−1·cm−1 or a combination of any of the foregoing values, or a range encompassing any of the foregoing values. In still another aspect, the SWIR dye has a fluorescence quantum yield of less than 5%, or less than 4%.
As used herein, the term “substituted” is contemplated to include all permissible substituents of organic compounds while not materially affecting the basic and novel characteristic(s) of the claimed invention. Also, the terms “substitution” or “substituted with” include the implicit proviso that such substitution is in accordance with permitted valence of the substituted atom and the substituent, and that the substitution results in a stable compound, e.g., a compound that does not spontaneously undergo transformation such as by rearrangement, cyclization, elimination, etc.
The term “alkyl” as used herein is a linear, branched, or cyclic saturated hydrocarbon group of 1 to 20 carbon atoms, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, s-butyl, t-butyl, n-pentyl, isopentyl, s-pentyl, neopentyl, hexyl, heptyl, octyl, nonyl, decyl, dodecyl, tetradecyl, hexadecyl, and the like. The alkyl group can be cyclic (also referred to as “carbocyclic”) or acyclic. The alkyl group can be branched or unbranched. The alkyl group can also be substituted or unsubstituted. For example, the alkyl group can be substituted with 1 to 3 substituents independently selected from a halogen, sulfonate, hydroxy, amino, nitro, cyano, carboxy, a heterocyclic group having 3 to 16 carbon atoms, an alkylether having 2 to 20 carbon atoms and 1 to 5 oxygen atoms, and an alkoxy group having 1 to 20 carbon atoms. A “lower alkyl” group is an alkyl group containing from one to six (e.g., from one to four) carbon atoms. The term alkyl group can also be a C1 alkyl, C1-C2 alkyl, C1-C3 alkyl, C1-C4 alkyl, C1-C5 alkyl, C1-C6 alkyl, C1-C7 alkyl, C1-C8 alkyl, C1-C9 alkyl, C1-C10 alkyl, and the like up to and including a C1-C18 alkyl.
The term “cycloalkyl” as used herein is a non-aromatic carbon-based ring composed of three carbon atoms to ten carbon atoms. Examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, norbornyl, and the like.
The terms “alkoxy” and “alkoxyl” as used herein to refer to an alkyl or cycloalkyl group bonded through an ether linkage; that is, an “alkoxy” group can be defined as -OA1 where A1 is alkyl, alkenyl or cycloalkyl as defined above.
The term “aromatic” as used herein refers to a ring structure having cyclic clouds of delocalized π electrons above and below the plane of the molecule, where the π clouds contain (4n+2) π electrons. A further discussion of aromaticity is found in Morrison and Boyd, Organic Chemistry, (5th Ed., 1987), Chapter 13, entitled “Aromaticity,” pages 477-497, incorporated herein by reference. The term “aromatic group” is inclusive of both aryl and heteroaryl groups.
The term “aryl” as used herein is a group that contains any carbon-based aromatic group including, but not limited to, benzene, naphthalene, phenyl, biphenyl, anthracene, pyrene, and the like. The aryl group can be substituted or unsubstituted. The aryl group can be substituted with one or more groups including, but not limited to, alkyl, cycloalkyl, alkoxy, alkenyl, cycloalkenyl, alkynyl, cycloalkynyl, aryl, heteroaryl, aldehyde, —NH2, carboxylic acid, ester, ether, halide, hydroxy, ketone, azide, nitro, silyl, sulfo-oxo, or thiol as described herein.
The term “aldehyde” as used herein is represented by the formula: —C(O)H. Throughout this specification “C(O)” is a shorthand notation for a carbonyl group, i.e., C═O.
The terms “amine” or “amino” as used herein are represented by the formula: —NA1A2, where A1 and A2 can be, independently, hydrogen or alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, cycloalkynyl, aryl, or heteroaryl group as described herein. A specific example of amino is —NH2.
The term “carboxylic acid” as used herein is represented by the formula: —C(O)OH.
The term “ester” as used herein is represented by the formulae: —OC(O)A1 and —C(O)OA1, where A1 is selected from alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, cycloalkynyl, aryl, and heteroaryl.
The term “amide” as used herein is represented by the formulae: —N(A1)C(O)A2 and —C(O)N(A2)2, where A1 and A2 are independently selected from hydrogen, alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, cycloalkynyl, aryl, and heteroaryl.
The term “ether” as used herein is represented by the formula: -A1OA2, where A1 and A2 are independently selected from alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, cycloalkynyl, aryl, and heteroaryl.
The terms “halo,” “halogen” or “halide,” as used herein can be used interchangeably and refer to a group selected from —F, —Cl, —Br, and —I.
The term “heteroalkyl” as used herein refers to an alkyl group containing at least one heteroatom. Suitable heteroatoms include, but are not limited to, O, N, Si, P and S, wherein the nitrogen, phosphorous and sulfur atoms are optionally oxidized, and the nitrogen heteroatom is optionally quaternized. Heteroalkyl groups can be substituted as defined above for alkyl groups.
The terms “heterocycle”, “heterocyclic” or “heterocyclyl,” as used herein can be used interchangeably and refer to single and multi-cyclic aromatic (heteroaromatic) or non-aromatic ring (heteroalkyl) systems in which at least one of the ring members is other than carbon. Heterocycle includes pyridine, pyrimidine, furan, thiophene, pyrrole, isoxazole, isothiazole, pyrazole, oxazole, thiazole, imidazole, oxazole, including, 1,2,3-oxadiazole, 1,2,5-oxadiazole and 1,3,4-oxadiazole, thiadiazole, including, 1,2,3-thiadiazole, 1,2,5-thiadiazole, and 1,3,4-thiadiazole, triazole, including, 1,2,3-triazole, 1,3,4-triazole, tetrazole, including 1,2,3,4-tetrazole and 1,2,4,5-tetrazole, pyridazine, pyrazine, triazine, including 1,2,4-triazine and 1,3,5-triazine, tetrazine, including 1,2,4,5-tetrazine, pyrrolidine, piperidine, piperazine, morpholine, azetidine, tetrahydropyran, tetrahydrofuran, dioxane, and the like. The term heterocyclyl group can also be a C2 heterocyclyl, C2-C3 heterocyclyl, C2-C4 heterocyclyl, C2-C5 heterocyclyl, C2-C6 heterocyclyl, C2-C7 heterocyclyl, C2-C8 heterocyclyl, C2-C9 heterocyclyl, C2-C10 heterocyclyl, C2-C11 heterocyclyl, and the like up to and including a C2-C18 heterocyclyl. For example, a C2 heterocyclyl comprises a group which has two carbon atoms and at least one heteroatom, including, but not limited to, aziridinyl, diazetidinyl, dihydrodiazetyl, oxiranyl, thiiranyl, and the like. Alternatively, for example, a C5 heterocyclyl comprises a group which has five carbon atoms and at least one heteroatom, including, but not limited to, piperidinyl, tetrahydropyranyl, tetrahydrothiopyranyl, diazepanyl, pyridinyl, and the like. It is understood that a heterocyclyl group may be bound either through a heteroatom in the ring, where chemically possible, or one of carbons comprising the heterocyclyl ring.
The term “hydroxyl” or “hydroxy” as used herein is represented by the formula —OH.
The term “nitro” as used herein is represented by the formula —NO2.
The term “nitrile” or “cyano” as used herein is represented by the formula —CN.
As described herein, compounds of the invention may contain “optionally substituted” moieties. In general, the term “substituted,” whether preceded by the term “optionally” or not, means that one or more hydrogens of the designated moiety are replaced with a suitable substituent. Unless otherwise indicated, an “optionally substituted” group may have a suitable substituent at each substitutable position of the group, and when more than one position in any given structure may be substituted with more than one substituent selected from a specified group, the substituent may be either the same or different at every position. Combinations of substituents envisioned by this invention are preferably those that result in the formation of stable or chemically feasible compounds. It is also contemplated that, in certain aspects, unless expressly indicated to the contrary, individual substituents can be further optionally substituted (i.e., further substituted or unsubstituted).
The term “radical,” refers to a fragment, group, or substructure of a molecule described herein, regardless of how the molecule is prepared. For example, a 2,4-thiazolidinedione radical in a particular compound has the structure:
regardless of whether thiazolidinedione is used to prepare the compound. In some embodiments the radical (for example an alkyl) can be further modified (i.e., substituted alkyl) by having bonded thereto one or more “substituent radicals.” The number of atoms in a given radical is not critical to the present invention unless it is indicated to the contrary elsewhere herein.
The patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Office upon request and payment of the necessary fee.
Small molecule organic dyes may be attractive for clinical applications because of their tendency to metabolize in the cells and their potential for low toxicity. Disclosed herein is a method for making SWIR organic dyes by combining donor and acceptor groups. In one aspect, the choice of a good donor—acceptor pair can significantly lower the optical bandgap of a dye due to the promotion of charge transfer events.
The novel SWIR dyes can be used in biological imaging such as in vivo photoacoustic or fluorescence imaging of tumor angiogenesis monitoring, blood oxygenation mapping, functional brain imaging, skin melanoma detection, methemoglobin measuring, etc. In one aspect, the disclosure provides new materials that absorb light in the SWIR region where biological tissues are most transparent. In a further aspect, the compositions allow for direct, real-time laser imaging of biological samples at a faster, more affordable rate than an MRI, while also potentially allowing real time analysis during surgery.
The xanthene-based dyes of the disclosure have outstanding photophysical properties and stimuli responses. For instance, the new SWIR xanthene-based PA imaging agents SCR-1, SCR-2 and SCR-3 have absorption maxima at 840, 950 and 1040, respectively.
These three dyes were made using the donor-acceptor-donor (D-A-D) design. These dyes were based on two factors: (i) a good overlap of the thiophene donor and xanthene acceptor to lower the bandgap of the dye due to charge transfer events, and (ii) an amino group connected to the thiophene to increase donor strength in the push-pull mechanism of xanthene-based dyes.
The xanthene dyes can be in a donor-acceptor (D-A) design or a donor-acceptor-donor (D-A-D) design. For the D-A-D design, the donors can be symmetrical or unsymmetrical. For symmetrical dyes, R1 and R2 are identical donors. For unsymmetrical dyes, R1 and R2 are not identical.
Although the structures described and drawn herein show the cationic charge on a single atom, the donors can be in resonance with one another such that the cationic charge can move from the donor at R1 to the donor at R2. R1 and R2 can be selected from one of the following Groups A, B and C:
R1 is hydrogen and R2 is a donor which is selected from a substituted or unsubstituted dibenzazepinyl, thienyldibenzazepine, bithienyldibenzazepine, thienodibenzazepine, dihydrodibenzazepinyl, thienyldihydrodibenzazepine, bithienyldihydrodibenzazepine, and thienodihydrodibenzazepine;
Both R1 and R2 are the same donors, are substituted or unsubstituted and are selected from dibenzazepinyl, thienyldibenzazepine, bithienyldibenzazepine, thienodibenzazepine, dihydrodibenzazepinyl, thienyldihydrodibenzazepine, bithienyldihydrodibenzazepine, and thienodihydrodibenzazepine; and
R2 is a different donor from R1, and R1 is a donor which is selected from a substituted or unsubstituted dibenzazepinyl, thienyldibenzazepine, bithienyldibenzazepine, thienodibenzazepine, dihydrodibenzazepinyl, thienyldihydrodibenzazepine, bithienyldihydrodibenzazepine, and thienodihydrodibenzazepine, and R2 is a donor which is substituted or unsubstituted, and is selected from dibenzazepinyl, thienyldibenzazepine, bithienyldibenzazepine, thienodibenzazepine, dihydrodibenzazepinyl, thienyldihydrodibenzazepine, bithienyldihydrodibenzazepine, thienodihydrodibenzazepine, 1-(thiophen-2-yl)piperidine, 1-(thieno[3,2-b]thiophen-2-yl)piperidine, 1-([2,2′-bithiophen]-5-yl)piperidine, a C2-C12 dialkyl amine, indolizine derivative, diphenylamine, and julolidine.
In Group B, the dyes are symmetrical, such as XanthCR-880. In Group C, the dyes are unsymmetrical. For instance, the donor at R1 could be thienyldibenzazepine and at R2 could be bithienyldibenzazepine; or both R1 and R2 can be thienyldibenzazepine but one is substituted and the other is not.
The preferred structures of the donors are shown below.
wherein R34, R35, R36, R37, R38, R39, R40, R41, R42, and R43 are independently selected from hydrogen, halogen, hydroxy, amino, nitro, cyano, carboxy, an alkyl group having 1 to 20 carbon atoms, an alkenyl group having 2 to 20 carbon atoms, an alkynyl group having 2 to 20 carbon atoms, an alkoxy group having 1 to 20 carbon atoms, an alkylether group having 2 to 20 carbon atoms and 1 to 5 oxygen atoms, an aryl group having 6 to 10 carbon atoms, and a heterocyclic group having 3 to 16 carbon atoms. Preferably the dibenzazepine has 3 or fewer substituents, more preferably 1 substituent and most preferably no substituents. These dibenzazepine donors are derived from the following ring structure:
The disclosure also includes dihydrogenated dibenzazepines as optional donors, wherein the above structures containing the R34, R35, R36, R37, R38, R39, R40, R41, R42, and R43 ligands are modified such that the carbons bonded to the R38 and R39 ligands each have an additional hydrogen attached thereto and the double bond between the carbons bonded to the R38 and R39 ligands is a single bond. These dihydrogenated dibenzazepine donors are sometimes referred to herein as dihydrodibenzazepinyl groups and are derived from a 10,11-dihydro-5H-dibenzo[b,f]azepine ring.
wherein R34, R35, R36, R37, R38, R39, R40, R41, R42, and R43 are as described above. R44 and R45 are each independently selected from hydrogen, halogen, hydroxy, amino, nitro, cyano, carboxy, an alkyl group having 1 to 20 carbon atoms, an alkenyl group having 2 to 20 carbon atoms, an alkynyl group having 2 to 20 carbon atoms, an aryl group having 6 to 10 carbon atoms, a heterocyclic group having 3 to 16 carbon atoms, an alkoxy group having 1 to 20 carbon atoms, and an alkylether group having 2 to 20 carbon atoms and 1 to 5 oxygen atoms. Preferably, the thienyldibenzazepine has 3 or fewer substituents, more preferably 1 substituent and most preferably no substituents. These thienyldibenzazepine donors are derived from the following ring structure:
The disclosure also includes dihydrogenated thienyldibenzazepines as optional donors, wherein the above-structures containing the R34-R45 ligands are modified such that the carbons bonded to the R38 and R39 ligands each have an additional hydrogen attached thereto and the double bond between the carbons bonded to the R38 and R39 ligands is, instead, a single bond. These dihydrogenated thienyldibenzazepine donors are sometimes referred to herein as thienyldihydrodibenzazepine groups and are derived from the following ring structure:
wherein R34, R35, R36, R37, R38, R39, R40, R41, R42, and R43 are as described above. R46, R47, R48 and R49 are each independently selected from hydrogen, halogen, hydroxy, amino, nitro, cyano, carboxy, an alkyl group having 1 to 20 carbon atoms, an alkenyl group having 2 to 20 carbon atoms, an alkynyl group having 2 to 20 carbon atoms, an aryl group having 6 to 10 carbon atoms, a heterocyclic group having 3 to 16 carbon atoms, an alkoxy group having 1 to 20 carbon atoms, and an alkylether group having 2 to 20 carbon atoms and 1 to 5 oxygen atoms. Preferably, the bithienyldibenzazepine has 3 or fewer substituents, more preferably 1 substituent and most preferably no substituents. These bithienyldibenzazepine donors are derived from the following ring structure:
The disclosure also includes dihydrogenated bithienyldibenzazepines as optional donors, wherein the above-structures containing the R34-R43 and R46-R49 ligands are modified such that the carbons bonded to the R38 and R39 ligands each have an additional hydrogen attached thereto and the double bond between the carbons bonded to R38 and R39 ligands is instead a single bond. These dihydrogenated bithienyldibenzazepine donors are sometimes referred to herein as bithienyldihydrodibenzazepine groups and are derived from the following ring structure:
wherein R34, R35, R36, R37, R38, R39, R40, R41, R42, and R43 are as described above. R50 and R51 are each independently selected from hydrogen, halogen, hydroxy, amino, nitro, cyano, carboxy, an alkyl group having 1 to 20 carbon atoms, an alkenyl group having 2 to 20 carbon atoms, an alkynyl group having 2 to 20 carbon atoms, an aryl group having 6 to 10 carbon atoms, a heterocyclic group having 3 to 16 carbons, an alkoxy group having 1 to 20 carbon atoms, and an alkylether group having 2 to 20 carbon atoms and 1 to 5 oxygen atoms. Preferably, the thienodibenzazepine has 3 or fewer substituents, more preferably 1 substituent and most preferably no substituents. These thienodibenzazepine donors are derived from the following ring structure:
The disclosure also includes dihydrogenated thienodibenzazepines as optional donors, wherein the above-structures containing the R34-R43 and R50-R51 ligands are modified such that the carbons bonded to the R38 and R39 ligands each have an additional hydrogen attached thereto and the double bond between the carbons bonded to R38 and R39 ligands is instead a single bond. These dihydrogenated thienodibenzazepine donors are sometimes referred to herein as thienodihydrodibenzazepine groups and are derived from the following ring structure:
wherein R5, R6, R7, R8, R9, R10, and R11 are each independently selected from hydrogen, sulfonate, halogen, hydroxy, amino, nitro, cyano, carboxy, an alkyl group having 1 to 20 carbons, an alkenyl group having 2 to 20 carbon atoms, an alkynyl group having 2 to 20 carbon atoms, an aryl group having 6 to 10 carbon atoms, a heterocyclic group having 3 to 16 carbon atoms, an alkoxy group having 1 to 20 carbon atoms, and an alkylether group having 2-20 carbon atoms and 1 to 5 oxygen atoms. Preferably, the 1-(thiophen-2-yl)piperidine has 3 or fewer substituents, more preferably 1 substituent and most preferably no substituents, in other words, most preferably all of R5, R6, R7, R8, R9, R10, and R11 are hydrogen.
The wavy line is globally used herein to refer to the location of the bond between the donor and the xanthene core structure.
wherein R7, R8, R9, R10, and R11 are as described above. R12 and R13 are each independently selected from hydrogen, sulfonate, halogen, hydroxy, amino, nitro, cyano, carboxy, an alkyl group having 1 to 20 carbon atoms, an alkenyl group having 2 to 20 carbon atoms, an alkynyl group having 2 to 20 carbon atoms, an aryl group having 6 to 10 carbon atoms, a heterocyclic group having 3 to 16 carbon atoms, an alkoxy group having 1 to 20 carbon atoms, and an alkylether group having 2 to 20 carbon atoms and 1 to 5 oxygen atoms. Preferably, the 1-(thieno[3,2-b]thiophen-2-yl)piperidine has 3 or fewer substituents, more preferably 1 substituent and most preferably no substituents.
wherein R7, R8, R9, R10, and R11 are as described above. R14, R15, R16, and R17 are each independently selected from hydrogen, sulfonate, halogen, hydroxy, amino, nitro, cyano, carboxy, an alkyl group having 1 to 20 carbon atoms, an alkenyl group having 2 to 20 carbon atoms, an alkynyl group having 2 to 20 carbon atoms, an aryl group having 6 to 10 carbon atoms, a heterocyclic group having 3 to 16 carbon atoms, an alkoxy group having 1 to 20 carbon atoms, and an alkylether group having 2 to 20 carbon atoms and 1 to 5 oxygen atoms. Preferably the 1-([2,2′-bithiophen]-5-yl)piperidine has 3 or fewer substituents, more preferably 1 substituent and most preferably no substituents.
A C2-C12 dialkyl amine which is bonded to the xanthene core at the amine nitrogen, and is optionally substituted with 1 to 3 substituents selected from an alkenyl group having 2 to 10 carbon atoms, an alkynyl group having 2 to 10 carbon atoms, an alkoxy group having 1 to 20 carbon atoms, an alkylether group having 2 to 20 carbon atoms and 1 to 5 oxygen atoms, an aryl group having 6 to 10 carbon atoms, and a heterocyclic group having 3 to 16 carbon atoms. Preferably, the C2-C12 dialkyl amine has 1 substituent and most preferably no substituents.
Indolizin-3-yl which is bonded to the xanthene core at the 3-position and is substituted or unsubstituted. Preferably, the indolizine-3-yl donor is substituted with 1 to 3 substituents which are independently selected from halogen, sulfonate, hydroxy, amino, nitro, cyano, carboxy, an alkyl group having 1 to 20 carbon atoms, an alkenyl group having 2 to 20 carbon atoms, an alkynyl group having 2 to 20 carbon atoms, an aryl group having 6 to 10 carbon atoms, a heterocyclic group having 3 to 16 carbon atoms, an alkoxy group having 1 to 20 carbon atoms, and an alkylether group having 2 to 20 carbon atoms and 1 to 5 oxygen atoms. Preferably, the indolizine-3-yl donor has 3 or fewer substituents; even more preferably the indolizine-3-yl donor is substituted at the 1- and 2-positions with an alkyl group having 1 to 5 carbons and an aryl group having 6 to 10 carbon atoms; and even more preferably, the indolizine-3-yl donor is substituted at the 1-position with an alkyl group having 1 to 4 carbon atoms and at the 2-position with an aryl group having 6 to 8 carbon atoms; and most preferably, the indolizine donor is 1-methyl-2-phenylindolizin-3-yl.
The diphenylamine preferably has the following cationic structure (the corresponding nonionic structure is not shown):
wherein R30, R31, R32, and R33 are independently selected from hydrogen, an alkyl group having 1 to 20 carbons, an alkenyl group having 2 to 20 carbon atoms, an alkynyl group having 2 to 20 carbon atoms, an alkoxy group having 1 to 20 carbon atoms, an alkylether group having 2 to 20 carbon atoms and 1 to 5 oxygen atoms, and an aryl group having 6 to 10 carbon atoms, at least one combination of (R30 and R31) and (R32 and R33) together with N forming a substituted or unsubstituted pyrrolidine ring, a substituted or unsubstituted piperidine ring, a substituted or unsubstituted morpholine ring, a substituted or unsubstituted tetrahydropyridine ring or a substituted or unsubstituted cyclohexylamine ring.
Julolidine preferably has the following cationic structure (the corresponding nonionic structure is not shown):
Julolidine is optionally substituted with 1 to 3 substituents independently selected from a halogen, sulfonate, hydroxy, amino, nitro, cyano, carboxy, an alkyl group having 1 to 20 carbon atoms, an alkenyl group having 2 to 20 carbon atoms, an alkynyl group having 2 to 20 carbon atoms, an alkoxy group having 1 to 20 carbon atoms, an alkylether group having 2 to 20 carbon atoms and 1 to 5 oxygen atoms, an aryl group having 6 to 10 carbon atoms, and a heterocyclic group having 3 to 16 carbon atoms. Preferably, the julolidine donor is not substituted or has one substituent.
The wavy line is refers to the location of the bond between the donor and the xanthene core structure.
In various aspects, the dyes of the present disclosure can be given to a patient in a biocompatible composition in an amount effective to allow for PA or Fl analysis of a particular tissue, organ or system. As used herein, “biocompatible” refers to a material or composition that does not cause harm to living tissue. In one aspect, the SWIR dyes disclosed herein are biocompatible. Herein, the term “biocompatible” is used interchangeably with “pharmaceutical” or “pharmaceutically acceptable”. As used herein, “pharmaceutically acceptable carriers” means one or more of a pharmaceutically acceptable diluents, preservatives, antioxidants, solubilizers, emulsifiers, coloring agents, releasing agents, coating agents, sweetening, flavoring and perfuming agents, and adjuvants.
In another aspect, the present disclosure relates to pharmaceutical compositions suitable for parenteral administration, such as intravenous administration, although the most suitable route in any given case will depend on the particular host, and nature and severity of the conditions for which the SWIR dyes are being administered for imaging.
In various aspects, the present disclosure also relates to a pharmaceutical composition comprising a pharmaceutically acceptable carrier or diluent and an effective amount of the SWIR dye compounds of the present disclosure for bioimaging, a product of the method of the disclosure, a pharmaceutically acceptable salt thereof, a hydrate thereof, a solvate thereof, a polymorph thereof, or a stereochemically isomeric form thereof. In a further aspect, the compounds of the present disclosure, a product of the method of the disclosure, a pharmaceutically acceptable salt thereof, a hydrate thereof, a solvate thereof, a polymorph thereof, or a stereochemically isomeric form thereof, or any subgroup or combination thereof may be formulated into various pharmaceutical forms for administration purposes.
Pharmaceutical compositions of the present disclosure suitable for parenteral administration can include sterile aqueous or oleaginous solutions, suspensions, or dispersions. Furthermore, the compositions can be in the form of sterile powders for the extemporaneous preparation of such sterile injectable solutions or dispersions. In some aspects, the final injectable form is sterile and must be effectively fluid for use in a syringe. The pharmaceutical compositions should be stable under the conditions of manufacture and storage; thus, preferably should be preserved against the contaminating action of microorganisms such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (e.g., glycerol, propylene glycol and liquid polyethylene glycol), vegetable oils, and suitable mixtures thereof.
Injectable solutions, for example, can be prepared in which the carrier comprises saline solution, such as a HEPES Buffered Saline or similar solutions.
As used herein, “nontoxic” refers to a material or composition that does not kill cells or organisms. In a further aspect, the SWIR dyes disclosed herein are nontoxic.
In terms of brightness, luminosity is dependent on the extinction coefficient (molar absorptivity) of the fluorophores or their ability to absorb light, and the quantum efficiency or effectiveness at transforming absorbed light into emitted luminescence. The SWIR dyes themselves are not very fluorescent, but they are sufficiently fluorescent for brightness imaging. For instance, when the SWIR dye binds to proteins, the protein becomes more easily detectable.
In an aspect, the disclosure relates to a method for making SWIR dyes, the method comprising:
wherein R18 and R21 are individually selected from Cl, Br, I and OSO2R52 and R52 is a hydrogen or lower alkyl;
Both R1 and R2 are the same donors and both are substituted or unsubstituted, and are selected from dibenzazepinyl, thienyldibenzazepine, bithienyldibenzazepine, thienodibenzazepine, dihydrodibenzazepinyl, thienyldihydrodibenzazepine, bithienyldihydrodibenzazepine, and thienodihydrodibenzazepine; and
R1 and R2 are different donors and R1 is a donor which is substituted or unsubstituted and is selected from dibenzazepinyl, thienyldibenzazepine, bithienyldibenzazepine, thienodibenzazepine, dihydrodibenzazepinyl, thienyldihydrodibenzazepine, bithienyldihydrodibenzazepine, and thienodihydrodibenzazepine and R2 is a donor which is substituted or unsubstituted and is selected from dibenzazepinyl, thienyldibenzazepine, bithienyldibenzazepine, thienodibenzazepine, dihydrodibenzazepinyl, thienyldihydrodibenzazepine, bithienyldihydrodibenzazepine, thienodihydrodibenzazepine, 1-(thiophen-2-yl)piperidine, 1-(thieno[3,2-b]thiophen-2-yl)piperidine, 1-([2,2′-bithiophen]-5-yl)piperidine, C2-C12 dialkyl amino, indolizine-3-yl, diphenylamino, and julolidinyl. In Group C, the dyes are unsymmetrical. For instance, the donor at R1 could be thienyldibenzazepine and at R2 could be thienodibenzazepine; or both R1 and R2 can be thienyldibenzazepine but one is substituted and the other is not.
Preferably, the compound formed in step (a) has the following structure:
In one aspect, the solvent in step (a) is selected from N-methyl-2-pyrrolidone (NMP), N,N-dimethylacetamide (DMA), dimethylformamide (DMF), toluene, tetrahydrofuran (THF), dioxane, and any combination thereof.
In one aspect, in step (a), the reaction mixture is heated at a temperature from about 80° C. to about 150° C., or at about 80° C., 85° C., 90° C., 95° C., 100° C., 105° C., 110° C., 115° C., 120° C., 125° C., 130° C., 135° C., 140° C., 145° C., or about 150° C., or a combination of any of the foregoing values, or a temperature within a range encompassing any of the foregoing values. In another aspect, the reaction mixture can be heated for from about 6 hours to about 30 hours, or for about 6 hours, 6.5 hours, 7 hours, 7.5 hours, 8 hours, 8.5 hours, 9 hours, 9.5 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, 18 hours, 19 hours, 20 hours, 21 hours, 22 hours, 23 hours, or about 24 hours, or a combination of any of the foregoing values, or for a time within a range encompassing any of the foregoing values. In one aspect, step (a) can be conducted in an inert atmosphere such as, for example, nitrogen.
In one aspect, step (a) further includes admixing a catalyst with the compound of Formula II and the Donor. The catalyst can be bis(triphenylphosphine)palladium(II) dichloride (PdCl2(PPh3)2), Palladium(II)acetate (Pd(OAc)2), tris(dibenzylideneacetone)dipalladium(0) (Pd(dba)3)CHCl3, or any combination thereof. In a further aspect, from about 0.01 to about 0.1 moles of catalyst can be used per mole of compound of Formula II. Further in this aspect, about 0.01, 0.02, 0.03, 0.04 or about 0.05 moles of catalyst can be used, or a combination of any of the foregoing values, or an amount within a range encompassing any of the foregoing values.
In another aspect, step (a) further includes admixing a base with the compound of Formula II and the Donor. In still another aspect, the base can be potassium acetate (KOAc), sodium acetate (NaOAc), Cs2CO3, KOtBu, NaOtBu, K2CO3, Na2CO3, or any combination thereof. In one aspect, from about 2.0 moles to about 6.0 moles of base can be used per mole of compound of Formula II. Further in this aspect, about 2.0, 2.7, 3.0, 3.2, or about 3.3 moles to about 6.0, 5.5, 5.0, or about 4.5 moles of base can be used, or a combination of any of the foregoing values, or a number of moles within a range encompassing any of the foregoing values.
In still another aspect, step (a) further includes admixing a ligand with the compound of Formula II and the Donor compound. In a further aspect, the ligand can be triphenylphosphine (PPh3), Dicyclohexyl[2′,4′,6′-tris(propan-2-yl)[1,1′-biphenyl]-2-yl]phosphane (Xphos), 2,2′-bis(diphenylphosphino)-1-1′-binaphthyl (BINAP), (tBu)2PMeHBF4, or any combination thereof. In an aspect, from about 1 to about 4 moles of ligand can be used per mole of catalyst, or about 1.5 to about 2.5, or about 2 moles of ligand per mole of catalyst can be used, or a combination of any of the foregoing values, or a number of moles within a range encompassing any of the foregoing values.
In an aspect, in step (b), the alcohol used in the transesterification reaction is also a solvent or cosolvent. The alcohol may be selected from methanol, ethanol, propanol, isopropanol, and butyl alcohol. In an aspect, the reaction mixture is heated to reflux. In another aspect, the reaction mixture can be heated for from about 6 hours to about 24 hours, or for about 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or about 24 hours, or a combination of any of the foregoing values, or for a time within a range encompassing any of the foregoing values. In one aspect, step (b) can be conducted in an inert atmosphere such as, for example, nitrogen.
As noted above, the donors can be symmetrical or unsymmetrical in a D-A-D designed molecule. For symmetrical dyes, such as XanthCR-880, the donor is combined with the acceptor using twice the moles of the donor when compared to the moles of acceptor. On the other hand, an SWIR dye having an unsymmetrical D1-A-D2 design could be prepared by essentially the same synthesis except that some of the DI reactants are replaced with D2 reactants, such that: the total moles (D1+D2) is twice the moles of A. In addition, SWIR dyes having R1═H could be made by starting with an acceptor having only one leaving group and using equimolar amounts of acceptor and donor.
Also disclosed are SWIR dyes produced by the disclosed methods.
In one aspect, disclosed herein is a composition including an SWIR dye disclosed herein and which optionally includes a carrier. In a further aspect, the carrier can be a pharmaceutically acceptable carrier. In still another aspect, the compositions can be biocompatible and/or nontoxic.
Also disclosed herein are methods for imaging a biological sample. In one aspect, the method includes the steps of (a) contacting the biological sample with a disclosed composition; (b) exposing the biological sample and the composition to SWIR radiation; and (c) observing PA emission in the biological sample. In a further aspect, the biological sample includes an organelle, a cell, a tissue, an organ, or any combination thereof.
Other potential applications include composites comprising the dyes in a polymer matrix for commodity items such as eyeglasses, night vision glasses or smart glasses, sensors, laser, and optoelectronic materials for electrical devices.
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.
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 20° C.-22° C. or is at ambient temperature, and pressure is at or near atmospheric.
All chemicals and solvents were purchased from commercial suppliers and used without further purification unless other-wise specified. Tetrahydrofuran (THF), isopropyl alcohol (IPA), and hydrogen peroxide (H2O2, 30% w/v) were purchased from Macron Fine Chemicals. IR-1061, acetaminophen (APAP), potassium superoxide, tert-butyl hydroperoxide, and tetrakis(acetonitrile)copper(I) hexafluorophosphate were purchased from Sigma-Aldrich. mDSPE-PEG MW 5,000 was purchased from Laysan Bio. Sodium chloride, 50K centrifugal filters, and Mohr's salt were purchased from MilliporeSigma. DEA-NONOate was purchased from Cayman Chemical. 1× phosphate saline buffer (Corning) was purchased from Thermo-Fisher Scientific. 25 mM HEPES and L-cysteine were purchased from Oakwood Chemical. Glutathione (reduced) was purchased from Acros Organics Chemicals. Isopentyl nitrite was purchased from Alfa Aesar Chemicals. Thieno[3,2-b]thiophene, 2-bromothiophene, 5-bromo-2,2′-bithiophene, and 5H-dibenzo[b,f]azepine were purchased from Sigma Aldrich or Fisher Scientific and used directly without further purification.
Thiophene (SCR-1), thienothiophene (SCR-2), and bithiophene (SCR-3) were incorporated into xanthene dye structures. The synthesis of the dyes began with the C—N cross-coupling of the dibenzazepine core with the corresponding bromothiophene derivatives (see Schemes 1-3 below), followed by a direct diarylation reaction with 3′,6′-dibromofluoran. From here, the dyes were trapped into the opened form by converting them to the corresponding ethyl ester, giving rise to SCR-1, SCR-2, and SCR-3. It is noteworthy that the synthesis of the dyes occurs in three steps from the bromothiophene derivatives.
An assessment of the photophysical properties of SCR-1, SCR-2, and SCR-3 in dichloromethane (DCM) revealed the absorption and emission bands of all three dyes extended well into the NIR to SWIR region, thus fulfilling the primary design objective (
An aspect of the invention is NO-responsive molecules, an example of which is SCR-NO that used the SCR scaffold.
The NO reactive species, SCR-NO (see the first PA-Active species in
To ensure the relative ratio of the probe and reference remained constant, a nanoparticle (NP) system that demonstrates minimal leaching of the encapsulated dye components was used. Moreover, the NP would ideally display excellent biocompatibility, display sufficient permeability to allow NO to diffuse into the core to access the encapsulated probe, and exhibit intrinsic liver targeting. The SCR-NO and IR-1061 were encapsulated using DSPE-PEG (a common phospholipids-polymer conjugate) to give NP-SCR-NO and NP-IR-1061, respectively. Both sets of NPs showed the predicted absorbance properties, where the spectra in water were similar to the un-encapsulated parent molecules in organic solvent (
Next, the baseline stability of the NPs was tested. Each solution was found to be stable, as no change in the absorbance intensity was observed compared to the initial time point over several days. However, when rNP-NO was treated with NO, the signal corresponding to the probe decreased while the signal from the reference remained unchanged (
Beyond NO treatment, the rNP-NO were subjected to a panel of biologically relevant analytes found in the liver (
One determinant of a strong PA signal is the magnitude of the extinction coefficient (EC) of a molecule. This often supersedes the quantum yield (QY) since the former of these two terms dominates the PA brightness value (EC×(1−QY)). With a calculated value of 8.28×104 M−1cm−1 for SCR-1, it was anticipated that strong ultrasound waves will be produced upon irradiation of rNP-NO. To test this, dense tissue-mimicking phantoms comprised of milk and agar were formulated. Individual samples containing each of the NPs were inserted to perform PA imaging. In the case of NP-SCR-NO, a PA signal was observed only when excited at the probe wavelength. Similarly, a PA signal originating from IR-1061 irradiation was the only ultrasound source from NP-IR-1061. However, PA signals were recorded for both the probe and reference when rNP-NO was subjected to the same in vitro test. As can be seen in
For in vivo testing of rNP-NO in live mice, an advanced MSOT (multispectral optoacoustic (photoacoustic) tomography) system was employed to track rNP-NO. PA imaging was performed using the MSOT because: 1) it can be used to visualize the entire animal (including the liver) and present the processed PA images in an easy-to-interpret cross-sectional view; 2) signals from the probe (SCR-NO) and reference dye (IR-1061) can be readily isolated from each other, as well as from interfering endogenous PA-active pigments via spectral unmixing; and 3) it can rapidly switch between the two SWIR wavelengths to facilitate ratiometric calibration. It is noteworthy that the ratiometric feature of rNP-NO is superior to systems where monitoring occurs at a single wavelength. First, a cohort of BALB/c mice (female 6-8-weeks old) were treated with a solution of rNP-NO (3-4 mg/mL) via systemic administration (intraperitoneal (i.p.) injection). The biodistribution profile was determined by tracking rNP-NO circulation at two wavelengths. Within 15 minutes, rNP-NO had predominately localized to the liver.
Next, the utility of the nanosensor in a drug-induced liver injury (DILI) model was established. Drug-induced liver injury (DILI) is responsible for 60% of acute liver failure cases in the United States and is notoriously difficult to diagnose. Visualizing molecular level changes in the liver during DILI allows for early intervention and is an important step toward deciphering the mechanistic underpinnings of the disease. For example, NO is believed to contribute to the progression of DILI due to the upregulation of inducible nitric oxide synthases in hepatocytes. Moreover, it is one of the earliest indicators of liver failure as the immune response is rapidly and aggressively activated. By injecting BALB/c mice with acetaminophen (APAP), a drug known to cause DILI when given at high doses, it was reasoned that reliable monitoring of the overproduction of NO could be obtained using rNP-NO and SWIR PA imaging. Mice selected to generate the DILI model were intraperitoneal injection (i.p.) injected with a solution (200 μL) of APAP at a dose of 300 mg/kg. In contrast, mice belonging to the control group were injected with the same volume of 0.9% saline. Then 16 hours were allowed to lapse thereby giving the mice time to succumb to DILI before rNP-NO was administered for SWIR PA imaging. Interestingly, the SWIR PA signal from the reference dye was localized to the liver for both groups and the overall signal intensity (color coded in green) was nearly identical (
C—H activation chemistry was employed to develop a panel of synthetically accessible SWIR dyes and to highlight their utility for in vivo activity-base sensing by developing a SWIR PA nanosensor. By incorporating a dibenzazepine donor conjugated to thiophene (SCR-1), thienothiophene (SCR-2), and bithiophene (SCR-3) moiety, the absorbance band of the resulting xanthene dyes bathochromically shifted into the SWIR region. Capping of the pendant carboxylate group of SCR-1 with an NO-responsive unit resulted in SCR-NO, which remained open prior to encountering NO. This probe was able to stay “on” or turn “off” to design rNP-NO. When encapsulated, the SCR-NO dye resulted in a bathochromic shift to the SWIR region. Co-encapsulation of SCR-NO with IR-1061 as the internal reference dye allowed for a large dynamic range and reliable ratiometric confirmation of NO sensing. This is highlighted in the series of in vitro studies described above as well as in an in vivo DILI model, where rNP-NO was able to differentiate between mice primed for liver damage using APAP versus a saline control. The use of SWIR to monitor the probe and internal reference assures that the PA signals are originating from a deep region of the liver. Beyond activity-based sensing, the SCR panel of SWIR dyes can be readily transformed into contrast agents by capping carboxylate with an alcohol to yield stable ester products. These SWIR molecules are useful in cancer treatment (e.g., SWIR PA surgical guidance).
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.
Although specific terms are employed herein, they are used in a generic and descriptive sense only and not for purposes of limitation.
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.
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.
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. It is noted that the dates of publication provided herein can be different from the actual publication dates, which can require independent confirmation.
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.
All references cited herein are hereby incorporated by reference in their entirety as if fully set forth herein.
This application claims priority to U.S. provisional application No. 63/385,577, filed on Nov. 30, 2022, the disclosure of which is hereby incorporated by reference in its entirety as if fully set forth herein.
This invention was made with government support under grant number OIA-1757220, awarded by the National Science Foundation. The government has certain rights in this invention.
| Number | Date | Country | |
|---|---|---|---|
| 63385577 | Nov 2022 | US |
