WO2021204802A1 - Labelled substance and methods of detection of inflammation and infection using said substance - Google Patents

Abstract

The present invention relates inter alia to a labelled substance, wherein the substance comprises a carrier exhibiting at least one scavenger structure, said scavenger structure comprising a nucleophilic centre complying with the formula X¹(-R''-)(-R')_mH_n (formula I) wherein a) X¹ is a single-bonded heteroatom selected amongst N, O and S and exhibits a free electron pair, b) m is 0 or 1 and n is 1 or 2 with the sum of m plus n being 2 for X¹ = N and 1 for X¹ = S 10 and O, c)-R''-is a bivalent organic group providing attachment to the carrier via one of its free valences and direct attachment to the heteroatom X¹ at the other one of its free valences, and d) R'-is a monovalent organic group directly attached to the heteroatom X¹ via its free valence, and wherein the substance further comprises a chemical entity comprising a detectable label. It also relates to a composition comprising said radiolabelled substance, and the use thereof in methods of detection of regions of inflammation, ischemia, infections, and malignancies in a subject.

Description

LABELLED SUBSTANCE AND METHODS OF DETECTION OF INFLAMMATION AND INFECTION USING SAID SUBSTANCE

TECHNICAL FIELD The present invention relates to labelled substance or composition comprising said labelled substance, and to the use thereof in methods of detection of inflammation, ischemia, infections, and malignancies in a subject.

BACKGROUND TECHNOLOGY Advanced lipid peroxidation end products (ALEs), i.e. aldehydes including malondialdehyde (MDA), 4-hydroxy-2-nonenal (4-HNE) and acrolein, are highly reactive molecules derived from oxidative degradation of polyunsaturated fatty acids in the cell membrane. These aldehydes react with proteins and form adducts which ultimately disrupt the structure and function of the protein. Thus, the detection of aldehydes and aldehyde-adducted proteins may be valuable as marker of any inflammation or inflammation related condition, including sterile inflammatory reactions, such as ischemia-reperfusion, infections and malignancies in the body.

PVAC is a polymer where polyvinyl alcohol (PVA) has been functionalized with multiple pendant carbazate groups. The hydrazine moiety of the carbazate group is nucleophilic and reacts with electrophiles such as carbonyls present in lipid peroxidation products to form Schiff-base-like carbazones, thereby inactivating endogenous aldehydes, such as MDA, 4-HNE and acrolein. The function of the PVA backbone is to serve as a carbazate carrier under the assumption that PVA modifies the pharmacokinetic profile of the attached carbazate groups. In W02009108100, inter alia carbazate-functionalized polyvinyl alcohol

(PVAC) and related substances was used to treat inflammatory conditions caused by various antigens, reactive mediators of inflammation, by scavenging said mediators. Still W02009108100 provides no indication on how to use said substances for detecting inflammation and especially not local inflammation.

SUMMARY OF THE INVENTION

The present inventors have surprisingly found that using a detectable amount of a detectable labelled substance, inflammations, in particular local inflammations, in body regions of a subject may be detected. What the present inventors saw was that when the labelled substance was administered to a subject it resulted in local signals in specific regions with high perfusion (short-term signals), but also in long-term signals (long half- life) indicating low or high degree inflammation. The results also indicated specific signalling from ischemic tissue or organs.

The labelled substance according to the present disclosure may thus be used to detect inflammatory body regions, infectious body regions, ischemic tissue or body region, malignant body regions and/or metastasis.

Surprisingly the labelled substance showed in vivo specificity to ischemic body tissues.

By detecting said inflammatory regions, it is also possible to detect infections such as local infections as said inflammation is a result of the infection, and the infection will be localised in or in the vicinity of the inflamed region. In one instance, the method was so specific as to detect a one-sided tooth inflammation/infection in a control animal.

By detecting said inflammatory regions, it is also possible to detect malignant body regions as these lead to inflammation, and the malignancies will be localised in or in the vicinity of the inflamed region.

The present labelled substance has the advantage of being specific for inflammatory sites but also of having a treating or preventing effect for inflammatory related conditions.

The above-mentioned methods are particularly suitable for detecting local conditions. In one embodiment, this may be in an organ or in a part of an organ.

First main aspect - Labelled substance

According to a first aspect, there is provided a labelled substance, wherein the substance comprises a carrier exhibiting at least one scavenger structure, said scavenger structure comprising a nucleophilic centre complying with the formula

X^l(-R”-)(-R’)_mH_n (formula I) wherein a) X¹ is a single-bonded heteroatom selected amongst N, O and S and exhibits a free electron pair, b) m is 0 or 1 and n is 1 or 2 with the sum of m plus n being 2 for X¹ = N and 1 for X¹ = S and O, c)-R”-is a bivalent organic group providing attachment to the carrier via one of its free valences and direct attachment to the heteroatom X¹ at the other one of its free valences, and d) R’-is a monovalent organic group directly attached to the heteroatom X¹ via its free valence, and wherein the substance further comprises a chemical entity comprising a detectable label.

The labelled substance may be used for detection of inflammatory body regions in a subject, in particular local inflammatory body regions.

In the present application the term “detectable label” means a substance, atom or a group that is detectable using any suitable means preferably imaging systems such as PET scans, SPECT scans or technetium scans. The detectable label may be but is not limited to radioactive, fluorescent or luminescent.

Second main aspect - Composition comprising the labelled substance

According to a second aspect, there is provided a composition comprising a detectable amount of the labelled substance according to the present invention.

The composition may be used for detection of inflammatory body regions in a subject, in particular local inflammatory regions.

Third main aspect - Labelled substance for use in detection of inflammation.

According to a third aspect, there is provided a labelled substance according to the present invention for use in detection of inflammatory body regions, infectious body regions, ischemic tissue or body region, malignant body regions or metastasis in a subject.

Fourth main aspect - A method for detection of inflammatory body regions in a subject a. According to a fourth aspect, there is provided a method for detection of inflammatory body regions in a subject, comprising the steps of b. administering a detectable amount of the labelled substance according to the present invention or the composition according to the present invention to the subject, c. allowing the labelled substance to bind to an inflammatory site, and d. detecting the signal from the labelled substance bound to the inflammatory site. Fifth main aspect - Method for detection of infectious body regions in a subject

According to a fifth aspect, there is provided a method for detection of infectious body regions in a subject, comprising the steps of: a. administering a detectable amount of the labelled substance according to the present invention or the composition according to the present invention to the subject, b. allowing the labelled substance to bind to an inflammatory site, and c. detecting the signal from the labelled substance bound to the inflammatory site; and d. detecting the infectious body regions located in or in the vicinity of said binding site.

Sixth main aspect - Method for detection of malignant body regions in a subject

According to a sixth aspect, there is provided a method for detection of malignant body regions in a subject, comprising the steps of: a. administering a detectable amount of the labelled substance according to the present invention or the composition according to the present invention to the subject, b. allowing the labelled substance to bind to an inflammatory site, and c. detecting the signal from the labelled substance bound to the inflammatory site; and d. detecting the malignant body regions located in or in the vicinity of said binding site.

Seventh main aspect - Method for detection of ischemic body regions in a subject

According to a seventh aspect, there is provided a method for detection of ischemic body regions in a subject, comprising the steps of: a. administering a detectable amount of the labelled substance according to the present invention or the composition according to the present invention to the subject, b. allowing the labelled substance to bind to an inflammatory site, and c. detecting the signal from the labelled substance bound to the inflammatory; and d. detecting the ischemic body regions located in or in the vicinity of said binding site. DESCRIPTION OF THE DRAWINGS

Figure 1 shows a synthesis scheme of the non-labelled substance (n), in this case PVAC, comprising the polymer functionalized with multiple carbazate groups (m).

Figure 2 shows a synthesis scheme of the conjugation labelling of the substance, in this case, PVAC with [¹⁸F]FBA at pH 4.0. 1 n = 90%, m = 10% PVAC. 3 n = 90, m = > 9.5%, o = < 5%.

Figure 3 shows photos of an ischemic kidney model with injection of the radiolabelled substance (PVAC) or blocking with an infusion of cold PVAC before injection of the radiolabelled PVAC and dynamic PET scan for 90 minutes. A - F. Region of interest (ROI) from fusion images (PET/CT, PET/MR) using CT or MR as a guide used to outline different parts of the kidneys: A Axial cross section of kidney, B axial outlined cortex. C frontal cross section of cortex. D Bladder with a 1.5 mm sphere. E Frontal cross section of the outflow tract with a 1 mm sphere. F Medulla represented by a 3 mm sphere. G Graph of the dynamic PET scan showing standardized uptake value (SUV) for the kidneys. The difference was 10.85 (8.1 - 15.8, =<0.0001) between the ischemic kidney and non- ischemic kidney at 60 minutes. Similar results between the ischemic kidney and the blocked kidney 11.9 (7.4 - 14.3,/ =<0.0001). H Ex vivo measurement of unblocked and blocked kidneys, a difference was seen in the ischemic kidney compared to the non ischemic kidney 3.86 (1.5 - 6.2, p = 0.0095 ), no difference was seen between the ischemic and non-ischemic kidney in blocked animals (p = 0.9454). I Frontal PET scan of a blocked ischemic kidney (B.K) and a non-ischemic kidney (nl.K). J Frontal PET scan of an ischemic kidney (I.K) compared with a non-ischemic kidney (nl.K). K Video with uptake of radiolabelled PVAC over time in control animal with no previous surgery. L Rotating 3D reconstruction of an ischemic kidney model animal 60 minutes into the PET scan. Figure 4 shows graphs and photos of an ischemic limb model with injection of the radiolabelled substance (PVAC) followed by dynamic PET scan for 90 minutes. A, B. Region of interest (ROI) from fusion images (PET/CT, PET/MR) using CT or MR as a guide used to outline muscular tissues from the hind legs. A. Axial cross section of hind limbs B. Length of the limb (tibia) with a small cylinder (SC) and a large cylinder (LC). The following formula was used to calculate the standardized uptake value (SUV) in the limb;

C. Data from dynamic PET scans. D. Ex vivo measurement from ischemic limb, non ischemic limb and paired non-ischemic limb. A higher SUV was seen in ischemic limb compared with non-ischemic limb, 0.134 (0.075 - 0.193, ? = 0.0003, but no difference was seen in ischemic limb compared with paired non-ischemic limb,/? = 0.0699). E. Coronary section of ischemic limb (I.L) and non-ischemic limb (Ni.L).

Figure 5 shows graphs and photos of the pharmacokinetics of the radiolabelled substance (PVAC) using PET scans of rats injected (I.V or I.J) with either radiolabelled PVAC or fluorochrome labelled PVAC. A. The elimination Ti_/2for aorta was 0.17h = 10.2min (1.9 - 40.8). The accumulation n_/2for the bladder was 0.17h = 10.3min (5.6 - 19.2). B. Long term pharmacokinetic studies of fluorochrome labelled PVAC injected in rats either I.V or I.J. For the I.V. injection the elimination phase was split into: a fast phase and a slow phase. The fast phase (50% of the elimination) had a t_\h of 0.2h (0.11 - 0.33). The slow phase had a t_\h of 10.73h (7.1 - 15). I.J injection gave rapidly increased serum concentrations followed by a steady state at 2h between elimination and absorption and at 6h elimination took over, leading to a n/2 of 34.90h (24.92 - 51.45). The dotted line marked the transition from fast to slow phase. C. Video showing uptake of PVAC over 90 min PET scan. D. Post I.J injection with arrows marking the bladder and kneecap. E. PVAC uptake in the bladder. F. Total excreted PVAC in gathered urine (at 96 h) was 37.75% (17.58 - 57.91) after I.V injection and 20.30% (13.39 - 27.21) after I.J injection. Figure 6 shows a graph the pharmacokinetics of the Ex vivo biodistribution of radiolabelled substance (PVAC) for different organs on x-axis and standardized uptake value (SUV) on y-axis. The numbers above the bars are average SUV for each organ from nine different animals (3 male, 6 female).

Figure 7. Different reactive mediators formed during IRI tested for their affinity towards the non-radiolabelled substance (PVAC). PVAC reduced the signal for oxidized albumin ip = 0.0001), methylglyoxal (p = 0.0015), malondialdehyde ip = 0.0022) and, acrolein ip = 0.0073). ITEMIZED EMBODIMENTS

Item 1. A labelled substance, wherein the substance comprises a carrier exhibiting at least one scavenger structure, said scavenger structure comprising a nucleophilic centre complying with the formula

X¹(-R”-)(-R^,)mH_n (formula I) wherein a) X¹ is a single-bonded heteroatom selected amongst N, O and S and exhibits a free electron pair, b) m is 0 or 1 and n is 1 or 2 with the sum of m plus n being 2 for X¹ = N and 1 for X¹ = S and O, c) -R”- is a bivalent organic group providing attachment to the carrier via one of its free valences and direct attachment to the heteroatom X¹ at the other one of its free valences, and d) R’ - is a monovalent organic group directly attached to the heteroatom X¹ via its free valence, and wherein the substance further comprises a chemical entity comprising a detectable label.

Item 2. The substance according to item 1, wherein in that either one or both of the organic groups R’ - and -R”-, with preference for -R”-, comprise a structure of the formula:

-CH₂(X⁴)_o-(C=X'V(X²)_m- (formula II) wherein a) each of m’, n’ and o’ is 0 or 1, with preference for m’ being 1 with further preference for either one or both of n’ and o’ also being 1, b) each of X², X³, and X⁴, is selected amongst NH and a heteroatom S or O, with preference for either one or both of X² and X⁴ being selected amongst NH and O with further preference for X³ being selected amongst NH, O and S, c) the left free valence provides binding to a monovalent alkyl group R*- or to the carrier via at least a bivalent alkylene group -R**-, each of which two groups comprises the methylene group -CH2- shown in formula II, and d) the right free valence binds directly to the first heteroatom X¹.

Item 3. The substance according to item 1 or 2, wherein in that the nucleophilic center is part of a group selected amongst a) amino groups, preferably primary or secondary amino groups, b) hydrazide groups, preferably-NH-NH2, preferably as part of a -CONHNH2 group, a semicarbazide group, preferably -NHCONHNH2, a carbazate group, preferably - OCONHNH2, a thiosemicarbazide group, preferably -NHCSNHNH2, a thiocarbazate group, preferably -OCSNHNH2, c) aminooxy groups, preferably -ONH2, and d) thiol groups, preferably -SH.

Item 4. The substance according to any of the preceding items, wherein in that the carrier is a macromolecular carrier and/or is water-soluble or water-insoluble and preferably exhibits polymer structure.

Item 5. The substance according to any of the preceding items, wherein in that a) the carrier is water-insoluble and defines a support, and/or b) the substance is attached to a water-insoluble support, and preferably the carrier is polyvinyl alcohol.

Item 6. The substance according to any of the preceding items, wherein in that the scavenger structure is capable of undergoing an addition reaction with a carbonyl group of an aldehyde group and/or with a carbon-carbon multiple bond to which is directly attached a carbonyl group, such as an aldehyde group.

Item 7. The substance according to any of the preceding items, wherein the scavenger is a carbazate group.

Item 8. The substance according to item 1, wherein in that the substance is carbazate- functionalized polyvinyl alcohol.

Item 9. The substance according to anyone of the preceding items, wherein the detectable label is selected from radioactive labels and fluorochrome or fluorophores. Item 10. The substance according to item 9 wherein the detectable label is radioactive and comprises a radioactive isotope selected from ^UC, ¹³N, ¹⁵0, ¹⁹F, ⁶⁸Ga, ⁸⁹Zr, and ⁸²Rb. Item 11. The substance according to anyone of items 1 to 10, wherein 0.1-7.5%, preferably 0.1-5% of the scavenger structures are labelled.

Item 12. The substance according to item 9, wherein the detectable label is fluorochrome and wherein said fluorochrome is fluorescein isothiocyanate.

Item 13. A composition comprising a detectable amount of the labeled substance defined in anyone of items 1-10 and diagnostically suitable excipients.

Item 14. A labelled substance according to anyone of items 1 to 12 for use in detection of inflammation body regions, infectious body regions, ischemic tissue or body region, malignant body regions or metastasis.

Item 15. The labelled substance according to item 14 wherein the inflammation is a local inflammation.

Item 16. A method for detection of inflammatory body regions in a subject, comprising the steps of a. administering a detectable amount of the labelled substance defined in anyone of items 1-11 or the composition defined in item 12 to the subject, b. allowing the labelled substance to bind to an inflammatory site, and c. detecting the signal from the labelled substance bound to the inflammatory site.

Item 17. The method according to item 16, where the substance is administered intravenously.

Item 18. The method according to item 16 or 17, wherein the inflammatory region is inflammatory intestines, liver, pancreas, spleen, lung, muscles, tendons, joints, blood vessels, oral cavities, subcutaneous fat, and tissue surrounding implanted implants. Item 19. The method according to item 16 or 17, wherein the inflammatory region is atherosclerotic plaques in blood vessels. Item 20. A method for detection of infectious body regions in a subject, comprising the steps of: a. administering a detectable amount of the labelled substance defined in anyone of items 1-11 or the composition defined in item 12 to the subject, b. allowing the labelled substance to bind to an inflammatory site, and c. detecting the signal from the labelled substance bound to the inflammatory site; and d. detecting the infectious body regions located in or in the vicinity of said binding site Item 21. The method according to item 20, wherein said infectious region is selected from intrabdominal muscle, muscle fascia, joint, and lung.

Item 22. A method for detection of malignant body regions in a subject, comprising the steps of: a. administering a detectable amount of the labelled substance defined in anyone of items 1-11 or the composition defined in item 12 to the subject, b. allowing the labelled substance to bind to an inflammatory site, and c. detecting the signal from the labelled substance bound to the inflammatory site; and d. detecting the malignant body regions located in or in the vicinity of said binding site.

Item 23. The method according to item 22, wherein said malignant body regions is a cancer tumour or metastasis.

Item 24. A method for detection of ischemic body regions in a subject, comprising the steps of: a. administering a detectable amount of the labelled substance defined in anyone of items 1-11 or the composition defined in item 12 to the subject, b. allowing the labelled substance to bind to an inflammatory site, and c. detecting the signal from the labelled substance bound to the inflammatory site; and d. detecting the ischemic body regions located in or in the vicinity of said binding site.

Item 25. The method according to item 24, wherein the ischemic region is the result of ischemic reperfusion injury in kidney, intestines, extremities, and cardiovascular system.

Item 26. The method according to item 16 or 25, wherein the subject is a mammal.

Item 27. The method according to anyone of items 16-26, wherein the detection of the signal from the labelled substance is achieved by positron emission tomography.

DETAILED DESCRIPTION OF THE INVENTION

The present disclosure is generally directed to a labelled substance or composition comprising said labelled substance, and to the use thereof in methods of detection of inflammation, infections, and malignancies in a subject.

Definitions:

"Administration" or "administering", as used herein, refers to providing, contacting, and/or delivering a compound or compounds by any appropriate route to achieve the desired effect. Administration is preferably intravenous. "Subject", as used herein, is intended to include human and non-human animals. In examples, the subject is a mammal and/or a human. Exemplary human subjects include a human patient having a disorder, e.g. an inflammatory or infectious disease or disorder. The term "non-human animals" includes all vertebrates, e.g., non mammals (such as chickens, amphibians, reptiles) and mammals, such as non-human primates, domesticated and/or agriculturally useful animals (such as sheep, dogs, cats, cows, pigs, etc.), and rodents (such as mice, rats, hamsters, guinea pigs, etc.).

The subject or the individual is typical an animal, such as a vertebrate, with emphasis on a mammal such as a human being.

First main aspect - Labelled substance

The substance according to the present invention comprises a carrier exhibiting at least one scavenger structure, said scavenger structure comprising a nucleophilic centre complying with the formula

X¹(-R”-)(-R^,)mH_n (formula I).

According to an embodiment of the labelled substance, either one or both of the organic groups R’-and-R”-, with preference for -R’ comprise a structure of the formula:

-CH₂(X⁴)_o-(C=X'V(X²)_m- (formula II) wherein a) each of m’, n’ and o’ is 0 or 1, with preference for m’ being 1 with further preference for either one or both of n’ and o’ also being 1, b) each of X² , X³, and X⁴, is selected amongst NH and a heteroatom S or O, with preference for either one or both of X² and X⁴ being selected amongst NH and O with further preference for X³ being selected amongst NH, O and S, c) the left free valence provides binding to a monovalent alkyl group R*-or to the carrier via at least a bivalent alkylene group -R**-, each of which two groups comprises the methylene group-CFF-shown in formula II, and d) the right free valence binds directly to the first heteroatom X¹.

According to another embodiment of the labelled substance, the nucleophilic center is part of a group selected amongst a) amino groups, preferably primary or secondary amino groups, b) hydrazide groups, preferably-NH-NH2, preferably as part of a -CONHNH2 group, a semicarbazide group, preferably as -NHCONHNH2, a carbazate group, preferably as - OCONHNH2, a thiosemicarbazide group, preferably as -NHCSNHNH2, a thiocarbazate group, preferably as -OCSNHNH2, c) aminooxy groups, preferably as -ONH2, etc, and d) thiol groups, preferably-SH.

In a preferred embodiment the nucleophilic center is a carbazate group. According to a still further preferred embodiment of the labelled substance, the carrier is a macromolecular carrier and/or is water-soluble or water-insoluble and preferably exhibits polymer structure. The carrier may be water-insoluble and define a support and/or the labelled substance is attached to a water-insoluble support. A water soluble carrier is more preferred since it can be dissolved in buffers and body fluids and is also easier administered. In a preferred embodiment the carrier is a water-soluble macromolecular carrier preferably a polymer, preferably selected from polyvinyl alcohol or polysaccharide wherein said polysaccharide is preferably selected from dextran, starch, agarose, agaropektin, cellulose or glucose amino glucan (GAG) preferably hyaluronic acid.

According to another example, the labelled substance has a scavenger structure capable of undergoing an addition reaction with a carbonyl group of an aldehyde group and/or with a carbon-carbon multiple bond to which is directly attached a carbonyl group, such as an aldehyde group. Without being bound by theory, it is believed that the scavenger structure binds to said aldehyde found at inflammatory sites.

In a preferred embodiment the labelled substance is carbazate-functionalized polyvinyl alcohol.

The scavenger structure

The scavenger structure comprises a first nucleophilic centre which preferably is capable of participating in an addition reaction with the carbonyl group (C=0) of an aldehyde group, -and/or with a C,C-multiple bond to which one or more electron-withdrawing substituents preferably are directly attached.

The nucleophilic centre (first centre) of the scavenger structure preferably comprises a single-bonded first heteroatom N, O or S (= X¹) which exhibits a) a free electron pair, b) one or two hydrogens, and c) one or two organic groups R’ -(monovalent) and-R” -(divalent) directly bound to the heteroatom.

The preferred heteroatoms are N and S. S is preferably combined with the presence of a second nucleophilic centre, such as a primary or secondary amino, in the same scavenger structure as discussed below. The bivalent organic group-R” -provides binding to the carrier via one of its free valencies. The other free valency of -R”- as well as the free valency of the other organic group R’-are directly attached to the heteroatom X¹.

Generically a nucleophilic centre has the formula:

X¹(-R”-)(-R^,)mH_n (formula I) where X¹ , R’-and-R”-are as defined in the preceding paragraph and m is 0 or 1 and n is 1 or 2 with the sum of m plus n being 2 for X¹ = N and 1 for X¹ = S and O.

A single-bonded atom means that the atom is directly bound to other atoms only by single bonds. A multiple-bonded atom means that the atom is directly bound to another atom by a triple or a double bond. The atoms referred to are primarily N, O, S and carbon. The preferred nucleophilic centres are typically uncharged when interacting with an aldehyde. For a nucleophilic centre which is an uncharged base or acid form of an acid-base pair > 5%, such as > 25% or > 50 or > 75%, of the total concentration of the acid-base pair should be in uncharged form.

When the heteroatom X¹ is N, the ability to react with an aldehyde group will include that the adduct formed is capable of undergoing spontaneous elimination of water (H2O) to the formation of an imine structure (-CH=NR”-, m = 0 and n = 2) and/or an enamine structure (-CH=CHNHR”-, m = 0 and n = 2) or-CH=CHNR’R”-, m = 1 and n = 1) with both alternatives requiring a hydrogen (a-hydrogen) on a sp³-hydridised a-carbon of the aldehyde group-CHO). When the heteroatom X¹ is S or O, m = 0 and n = 1 which means that the structure obtained upon elimination of H2O is thioenolate or enolate (- CH=CHX'R’) (provided there is an a-hydrogen of the aldehyde group-CHO). These elimination reactions typically mean formation of a more stable product and/or a product that may react further to a further stabilized “end”-product. The selection of scavenger structures containing groups permitting subsequent reactions which end up in stabilized end products will support irreversibility of the initial addition reaction and are as a rule preferred.

The reaction of the first nucleophilic centre and a C,C-multiple bond on a acetaldehyde will result in a primary adduct which comprises the structure >CH-CHX¹- (for C,C-double bonds) and if the multiple bond is a,b to an aldehyde group there can be formed different tautomeric adducts. e.g.-CHX¹-CH=CHOH (enol) and-CHX¹-CH2- CH=0 (keto) which will enable another nucleophilic centre of the same or another scavenger structure to react with the aldehyde.

Either one or both of the organic groups R’-and-R” -comprise a structure of the formula

(formula II) where a) each of m’, if and o’ is 0 or 1, with preference for m’ being 1 with further preference for either one or both of n’ and o’ also being 1, b) each of X² , X³, and X⁴, is selected amongst NH and a heteroatom S or O, with preference for either one or both of X² and X⁴ being selected amongst NH and O with further preference for X³ being selected amongst NH, O and S, c) the left free valence provides binding to a monovalent alkyl group R*-or to the carrier via at least a bivalent alkylene group -R**-, each of which comprises the methylene group-CH2-shown of formula II, d) the right free valency binds directly to the first heteroatom X¹.

The substructure C=X³ (= B) includes also other ester-and amide-forming substructures which derive from acid functions and form an ester function when X² and/or X⁴ are oxygen and/or an amide function when X² and/or X⁴ are NH, e.g. sulphonamide (B is S(=0)₂) or phosphone amide (B is P=0(NH₂) or P=0(0H), n’ = 1).

Either one or both of the monovalent alkyl group R*-and the bivalent alkylene -R**-may be straight, branched or cyclic and possibly contain one or more structures selected amongst ethers (-0-,-S-), hydroxy (-OH), mercapto (-SH) and amino (-NH-,-ME). Each free valences represent binding to sp³-hybridised carbon (= alkyl carbon). Either one or both of these alkyl groups are preferably a lower alkyl which in this context means that they comprise one, two, three, four, five up to ten sp³-hybridised carbons typically with at most one heteroatom O, N and S bound to one and the same carbon. The groups are typically inert in the sense that they are not participating in the reaction which interferes with the aldehyde. The hydrogens given in formula (I) and/or its substructures may be replaced with an alkyl group selected amongst the same alkyl groups as discussed for R*-.

It is preferred that the bivalent group -R” -which attaches the first nucleophilic centre to the carrier comprises a substructure complying with formula I and/or II. The structural elements (substructures) discussed in the preceding paragraphs will support delocalisation of electrons and therefore further support irreversibility of the initial addition reaction.

Preferred scavenger structures thus have a nucleophilic centre which contain the first heteroatom X¹ together with a structure complying with formula II and are selected amongst: a) amino groups preferably primary or secondary amino groups b) hydrazide groups such as -NH-NH₂, e.g. as part of a-CONHNTk group, a semicarbazide group such as -NHCONHNH₂, a carbazate group such as -OCONHNH₂, a thiosemicarbazide group such as -NHCSNHNH₂, a thiocarbazate group such as - OCSNHNH₂ (formation of hydrazone, semicarbazone, thiocarbazone linkages/groups, etc when undergoing addition/elimination reactions with an aldehyde group) c) aminooxy groups, such as -ONH₂, etc (formation oxime linkages/groups, etc when undergoing addition/elimination reactions with an aldehyde group), d) a thiol group e.g.-SH (Michael addition products are formed when the thiol group reacts with a C,C-double bond. The product may undergo keto-enol tautomerisation when the double bond is a,b to a keto-or aldehyde-carbonyl, see above).

The free valence indicated in each of the groups given in the preceding paragraph preferably attaches the nucleophilic centre to the carrier via a linker structure comprising the above-mentioned bivalent alkylene group -R**-. A hydrogen bound directly to nitrogen may be replaced with a monovalent alkyl group selected amongst the same alkyl groups as R*-as long as they are not substantially counteracting the desired reactivity of the unsubstituted form of the nucleophilic centre. Thus the hydrogen in a thiol group and in a hydroxyl group cannot be replaced, for instance. Two replacing alkyl groups may form a cyclic structure together with atom to which they are attached, i.e. form a bivalent alkylene group e.g. selected amongst the alternatives for the -R**-group.

The bivalent structures -R**- and -R”- discussed above comprises next to the carrier a linker structure which does not negatively affect the desired effect of the nucleophilic centre of the scavenger structure. Such structures are not part of the invention and suitable such structures can be designed by the average-skilled person in the field.

In certain preferred scavenger structures there may be a second nucleophilic centre which a) may be part of one of the organic groups, e.g. the R*-or the -R**-group, and b) contain a first heteroatom N, O or S (= Y¹) in the same manner as for the first nucleophilic centre. In substance this means that this second nucleophilic centre complies with the formula:

Y¹(-R”-)(-R^,)mH_n (formula III) and the formula

-CH₂(Y⁴)_0”(C=Y³)n”(Y²)m”- (formula IV) where m, n, m”, n”, o”, Y¹, Y², Y³, Y⁴,-R”-and-R’ are selected in the same groups of variables as m, n, m’, n’, o’, X¹, X², X³, X⁴,-R”-and-R’of formula I and II. This includes that hydrogens (H) may be replaced as suggested for formulae I and II.

The heteroatom Y¹ preferably is part of a) an -NIT group where the free valence preferably may bind to a sp³-hybridised carbon, or b) a thiol group -SH where the free valence preferably may bind to a sp³-hybridised carbon. Each of m”, n” and o” in formula IV is 0 in both (a) and (b).

The distance between the first heteroatom Y¹ and the first heteroatom X¹ is typically larger than two or three atoms with upper limits being e.g. 20 atoms with preference for 4, 5 or 6 atoms between these two heteroatoms. The distance should support intra-molecular cyclisation, typically via one or more addition reactions. This cyclisation typically comprises an addition reaction between the second nucleophilic centre and a) a carbon-carbon or a carbon-heteroatom double bond formed as described above by reaction of the first nucleophilic centre with the starting aldehyde group, and/or b) a multiple C,C-bond present already in the starting aldehyde, such as a double C,C- bond, e.g. a,b to the aldehyde group, and/or c) a second keto or aldehyde carbonyl group provided such a group is present in the acetaldehyde molecule.

The result of the cyclisation is an n-membered ring-structure containing the first heteroatom Y¹ and the first heteroatom X¹ with n in n-membered being an integer > 3 with preference for 5 or 6. Larger rings may also be formed, such as 7-, 8-, 9-, 10-, 11-, 12-, 13-, 14-membered rings, as long as steric considerations and relative positions of functional groups so admit. The cyclisation may be followed by rearrangement reactions, e.g. intramolecularly, and/or elimination reactions creating carbon-heteroatom double bond(s), ring-openings, etc.

The carrier

The selection of suitable carriers depends on the requirements of a particular use. The typical carrier is selected amongst macromolecular compounds, i.e. is a compound which has a molecular weight of > 2000 dalton, preferably > 10000 dalton or > 50000 dalton, and preferably exhibits a polymeric structure, i.e. is a polymer which may be a homopolymer, copolymer or a chemical adduct between two or more polymers of different polymeric structure. Other suitable carriers may have molecular weights < 2000 dalton and exhibit polymeric structure as indicated by the possibility of the low numbers of monomeric units discussed below, e.g. > 20 and < 100. The term “adduct polymer” in this context means a product formed by reacting two polymers exhibiting mutually groups capable of forming covalent bonds that link the two polymers together upon reaction of the two mutually groups with each other. See for instance WO 2009108100 (IPR-Systems AB) and references cited therein. Suitable macromolecular carriers may thus be selected amongst synthetic polymers (= man-made polymers), biopolymers (nature-made polymers such as polysaccharides, polypeptides, proteins, etc) and biosynthetic polymers where “biosynthetic polymer” refers to a macromolecular carrier or compound exhibiting both a synthetic polymeric structure and a biopolymeric structure. A carrier polymer may be cross-linked or not cross-linked. With respect to branching the polymer may be unbranched, i.e. linear, or branched including either hyperbranched or dendritic. The degree of branching may thus vary between 0 and 1, such as be > 0.10 or > 0.25 > 0.5 > 0.75 or > 0.90 and/or < 0.90 or < 0.75 or < 0.50 or < 0.25 or < 0.10. Cross-linked polymers are as a rule insoluble in aqueous liquids while the solubility of non-cross- linked polymers depend on the overall structure of the polymer, e.g. presence and amount of polar and/or hydrophilic groups. Carrier polymers may also be derivatized to contain non-polymeric or polymeric groups, for instance cross-links, substituents, charged or uncharged groups, scavenger structures (as discussed above), etc. Macromolecular carriers which are insoluble in aqueous liquids may have different physical and geometric shapes as discussed for support materials elsewhere in this specification.

The term polymer above includes organic as well as inorganic polymers.

The macromolecule or polymer used in the carrier may be water-insoluble and suspensible in aqueous liquid media (when in particle form).

Polymers and other macromolecules suitable as carrier material may be hydrophilic or hydrophobic with preference for hydrophilic. Pronounced hydrophobic macromolecular carriers are as a rule insoluble in aqueous liquids meaning that there may be a risk for host defence reactions with them and also that the availability of nucleophilic centres for reaction with aldehyde may not be optimal. In order to overcome this kind of problems, it is often preferred to introduce hydrophilic groups on their surfaces (hydrophilization). The introduction of hydrophilic groups may among others be accomplished by a) coating with a hydrophilic material, b) selecting building blocks/monomers which exhibit hydrophilic groups and appropriate conditions during synthesis of the macromolecular compound, and c) chemical derivatisation with hydrophilic groups subsequent to the synthesis of the basic hydrophobic polymer, etc.

The hydrophilicity of a group, structure or carrier molecule increases as a rule with an increase in the ratio r = the sum of the number of heteroatoms O, N and S divided by the sum of the number of carbon atoms. Hydrophilic groups/compounds typically have an r > 0.5, preferably > 1.0, and for hydrophobic groups r < 1.0, preferably < 0.5. Typical hydrophilic groups are hydroxy, amino, amido, carboxy (including free acid carboxyl as well as carboxylate (ester and salt), etc. Typical hydrophobic groups are alkyls (CnH^n+i)- , C_nH(2n-i)-, C_nH(2n-3)-, etc), phenyls including alkyl phenyls, benzyl including other phenylalkyls, etc.

A carrier macromolecule typically comprises a polymer backbone which comprises > 5, or more preferably > 10 such as > 25 different and/or identical monomeric units linked together. The polymer may carry projecting or pending polymeric and/or non-polymeric groups of various lengths and kinds. A carrier polymer is preferably hydrophilic with hydrophilic groups selected amongst those given elsewhere in this specification. The most preferred hydrophilic group is hydroxy with the preferred carrier polymers and/or other macromolecular carrier being selected by poly hydroxy polymers (PHP or PH-polymers) exhibiting > 5, with preference for > 10, such as > 25 or > 50 hydroxyl groups and/or > 5 monomeric subunits each of which exhibits one, two, three, four or more hydroxyl groups per unit.

Typical polymers that may be present in polymeric carriers are a) polyester polymers, b) polyamide polymers, c) polyether polymers, d) polyvinyl polymers, e) polysaccharides, etc. A carrier may comprise one or more of these polymers/polymeric structures.

Polyester polymers are in particular obtained by polymerisation of a) monomers exhibiting at least one hydroxy group and at least one carboxy group, or b) a mixture containing monomers exhibiting two or more hydroxy groups and monomers exhibiting two or more carboxy group.

Polyamide polymers are in particular obtained by polymerisation of a) monomers exhibiting at least one amino group and at least one carboxy group, or b) a mixture containing monomers exhibiting two or more amino groups and monomers exhibiting two or more carboxy group. An important group of polyamides are those that exhibit polypeptide structure together with a plurality of hydroxy groups (PH-polymers). Suitable polyamide polymers of this kind are typically based on hydroxy-, amino- carboxylic acids as monomers, in particular with the amino group positioned a to the carboxylic group, e.g. serine, threonine, tyrosine, proline, etc. Poly ether polymers are typically used in combination with other polymeric structures, e.g. polymers of (a), (b), (d) and/or (e) above, which are polyfunctional with respect to the presence of groups such as hydroxy, amino, etc. Typical polyether polymers are polyethylene oxide and various copolymeri sates between ethylene oxide and other lower alkylene oxides, lower epihalohydrins, etc. Polyvinyl polymers which may be suitable as polymeric carriers in the invention are typically found amongst polymers containing one, two or more different monomeric units selected amongst hydroxyalkyl acrylates and methacrylates, N- hydroxyalkyl acryl-and N-hydroxyalkyl methacrylamides, hydroxyalkyl vinyl ethers, vinyl esters, etc. Polyvinyl alcohols are typically obtained by partial hydrolysis of polyvinyl esters meaning that polyvinyl alcohols that are preferred in the invention typically exhibit residual amounts of ester groups (< 10% or < 5%).

Typical polysaccharides that may be present in carriers used in the invention include dextran, starch, agarose, agaropektin, cellulose, glucosamino glucanes (GAG), and derivates of these polysaccharides, etc. The most interesting polysaccharides are dextran, certain glucosamino glucanes (GAG) such as hyaluronic acid, etc.

A polymer to be used in the carrier may have been derivatized, e.g. cross- linked and/or functionalized after its synthesis.

The scavenger structure including the first, the optional second nucleophilic centre and the various heteroatoms discussed for the scavenger structures are typically part of one and the same organic group/substituent attached to the macromolecular carrier. In certain variants different parts of a scavenger structure may be part of different group s/substituents attached to the carrier and/or part of the carrier.

Sizes/molecular weights of suitable carrier polymers will among others depend on the actual application/use of the composition/method of the invention. Thus, suitable polymeric carriers with respect to a particular polymeric structure and/or size may vary within a wide interval. Thus as a rule the number of monomeric subunits (mean value) of a polymer present in the carrier may be > 20 or > 100 or > 200 or > 300 or > 500 or >

1000 or > 2000 or > 20 000 or > 50 000 and/or < 50000 or < 20000 or < 2000 or < 1000 or < 500 or < 300 or < 200, or < 100 (with the proviso that >-limit always is lower than a <-limit when these values are combined to define intervals). Preferred numbers of monomeric units may in some cases be found in the interval of 200 - 600 which in particular applies to the polyvinyl alcohol used in the experimental part.

Suitable numbers of scavenger structures or nucleophilic centres per monomeric unit of a polymer of the carrier will also depend on the use, the scavenger structure, etc, and may thus be found within a wide interval, such as < 80%, preferably < 50%, <40% or < 30% with lower limits being preferably 0.01% or 0.1% or 1% or 5% or 10% or 15% where 100% corresponds to one scavenger structure or nucleophilic centre per monomeric unit. For scavenger structures containing two or more nucleophilic centres the number of nucleophilic centres per monomeric unit may exceed 100%, such as >

100% or > 125% or > 150%. In a preferred embodiment the number of scavenger structures per carrier is 15-50%, more preferably 20-40%.

Labelling

The labelling of the substance may be obtained by coupling or binding it to a chemical entity comprising the detectable label.

The chemical entity comprising the detectable label may be coupled or bound to the carrier (backbone) or to one or more of the scavenger structures.

The label is preferably detectable using immunological test, projection radiography, fluoroscopy, PET (Positron Emission Tomography) scans, PET -MR or PET- MRI (Magnetic Resonance), PET-CT, X-ray, CT (Computer Tomography) or CAT (Computer Axial Tomography) scans or combinations thereof.

The labelling may also be achieved by using radiolabelled reagents when preparing the carrier or the scavenger.

The skilled person selects according to need a suitable chemical entity comprising the detectable label with the proviso the chemical entity when bound to the substance does not sterically hinder the function of the substance and in particular the scavenger structures.

If the entity is coupled to the substance via the scavenger structures such as the nucleophilic centre, there must be enough remaining scavenger structures that are available for binding to the inflammatory sites. The labelled substance, when testing PVAC, retains 100% of the scavenging potential compared to non-labelled substance when labelling of less than 1% of scavenging structures. When 5% of the scavenging structures groups were labelled around 80% of its scavenging properties remained.

In one embodiment, 0.1-25% of the scavenger structures are radiolabelled. The degree of radiolabelling depends on the tissue to be analysed. In a preferred embodiment 0.1-10% preferably 0.1-7.5% of the scavengers are radiolabelled, more preferably 0.1-5% or 1-5%. In other embodiments, 5-25% or 7.5-25% or 10-25% or 15-25% or 10-20% of the scavengers are labelled.

As only a part of the scavenger functions is blocked the labelled substance will also have a scavenging effect. Thus, the labelled substance will both indicate a site of inflammation and treat said inflammation as disclosed in W02009108100.

The labelled substance as well as the carrier as such is preferably soluble in aqueous liquids such as water, buffer, body fluids, such as blood, serum, plasma, urine, lymph, lachrymal fluid, intestinal juice, gastric juice, saliva, synovial fluid, etc, depending on the desired use.

Radiolabelling

The label of the chemical entity defined above may be radioactive (i.e. a radiolabel). In one embodiment, the radiolabel is selected from isotopes with short half- lives or long half-lives. In one preferred embodiment the isotope is selected from ^UC, ¹³N, ¹⁵0, ¹⁸F, ⁶⁸Ga, ⁸⁹Zr, and ⁸²Rb.

Fluorochrome labelling

A fluorochrome (or fluorophore ) is a fluorescent chemical compound that can re-emit light upon light excitation. Fluorochrome molecules can generally be classified into four categories: proteins and peptides, small organic compounds, synthetic oligomers and polymers, and multi-component systems. In one embodiment, fluorescein isothiocyanate (FITC) was used. Non-radioactive contrast agent

Contrast agents or contrast materials are used to improve pictures of the inside of the body produce by for example x-ray computed tomography (CT), magnetic resonance (MT) or ultrasound. According to the present invention the contrast agent may be iodine, barium or gadolinium based. Iodine and barium based contrast agents are preferably used in combination with x-ray, CT or x-ray CT and gadolinium is preferably used in combination with x-ray, x-ray MR or MR.

Synthesis of the labelled substance The substance may be synthesized according to well-known protocols, for instance of the kinds given in WO 2009108100 and references cited therein. The labelling of the substance may be done using well-known protocols including radiolabeling including PET radionucleotides and fluorescent reagents (Mol. Imaging boil., 2008 Jul-Aug, 10(4), 177-181, Pharmaceuticals (Basel), 2014 April, 7(4), 392-418, Semin nucl. Med., 2017, 47(5), 454-460, Polym. Int., 2015, 64, 174-182).

All variants and examples of the first main aspect can be combined with the second, third, fourth, fifth, sixth, or seventh main aspects unless expressly stated otherwise. Second main aspect - Composition comprising a labelled substance

According to the second aspect, there is provided a composition comprising a detectable amount of the labelled substance.

The labelled substance may take the form of a composition containing one or more formulations where at least one of them comprises the labelled substance. The labelled substance may be present in the composition: a) in dry form, for instance as free particles, b) in dissolved form, typically in an aqueous liquid medium, and c) in suspended/dispersed form, i.e. as water-insoluble particles suspended in an aqueous liquid medium. The term “dissolved” in this context means that the labelled substance is present as a solute. The labelled substance particles comprise substance in a pure form or diluted with some solid material. Useful concentrations of labelled substance in formulations according to (b) can be found within a broad interval. The composition may in addition to the labelled substance contains buffers, salts, etc required for enabling acceptable conditions in vivo for the patient. These constituents may be co-formulated with the substance in the composition.

In the below-mentioned methods for detection, the labelled substance or the composition mentioned above would be administered to a patient in need thereof. The patient may be an animal or a human. The administration ways or routes may vary according to the specific medical situation and are part of the knowledge of a medical practitioner.

The administration may be done locally or systemically or in combination. Some are given below as illustrative examples of such administration ways or routes.

Formulations may be in the form of liquids, solutions, suspensions, or emulsions.

Formulations suitable for intravenous injection, include aqueous and nonaqueous isotonic, pyrogen-free, sterile injection solutions which may contain antioxidants, buffers, preservatives, stabilizers, bacteriostats, and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non aqueous sterile suspensions which may include suspending agents and thickening agents, and liposomes or other microparticulate systems which are designed to target the compound to blood components or one or more organs. Examples of suitable isotonic vehicles for use in such formulations include Sodium Chloride Injection, Ringer's

Solution, or Lactated Ringer's Injection. The formulations may be presented in unit-dose or multi-dose sealed containers, for example, ampoules and vials, and may be stored in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules, and tablets. Formulations may be in the form of liposomes or other microparticulate systems which are designed to target the active compound to blood components or one or more organs.

All variants and examples of the second main aspect can be combined with the first, third, fourth, fifth, sixth, or seventh main aspects unless expressly stated otherwise.

Third main aspect - Labelled substance for use in detection of inflammation. According to a third aspect, there is provided a labelled substance or a composition according to the present invention for use in detection of inflammatory body regions in a subject.

All variants and examples of the third main aspect can be combined with the first, second, fourth, fifth, sixth, or seventh main aspects unless expressly stated otherwise.

Fourth main aspect - A method for detection of inflammatory body regions in a subject According to the fourth aspect, there is provided a method for detection of inflammatory body regions in a subject using the labelled substance or the composition according to the present invention.

The inflammatory region can be any anatomical location, but specific regions may e.g. be parts of the intestines, liver, pancreas, spleen, lung, muscles, tendons, joints, blood vessels, oral cavities, subcutaneous fat, and tissue surrounding implanted implants.

One example is of such an inflammatory region is atherosclerosis, arthritis, ischemic, atherosclerotic plaques on blood vessels, e.g. aorta arteries, carotid arteries.

Preferably the detection is done by using an immunological test, projection radiography, fluoroscopy, PET (Positron Emission Tomography) scans, PET -MR or PET- MRI (Magnetic Resonance), PET-CT, X-ray, CT (Computer Tomography) or CAT (Computer Axial Tomography) scans or combinations thereof.

All variants and examples of the fourth main aspect can be combined with the first, second, third, fifth, sixth, or seventh main aspects unless expressly stated otherwise.

Fifth main aspect - Method for detection of infectious body regions in a subject According to the fifth aspect, there is provided a method for detection of infectious body regions in a subject using the labelled substance or the composition according to the present invention.

The step of detecting the infectious body regions may be done using any suitable technique depending on the tissue. Preferably the detection is done by using an immunological test, projection radiography, fluoroscopy, PET (Positron Emission Tomography) scans, PET-MR or PET-MRI (Magnetic Resonance), PET-CT, X-ray, CT (Computer Tomography) or CAT (Computer Axial Tomography) scans or combinations thereof. The infectious region can be any anatomical location, but specific regions may e.g. be parts of the intrabdominal muscle, muscle fascia, joint, and lung.

All variants and examples of the fifth main aspect can be combined with the first, second, third, fourth, sixth, or seventh main aspects unless expressly stated otherwise.

Sixth main aspect - Method for detection of malignant body regions in a subject According to the sixth aspect, there is provided a method for detection of malignant body regions in a subject using the labelled substance or the composition according to the present invention. The malignant body regions may be a cancer tumour or metastasis.

All variants and examples of the fifth main aspect can be combined with the first, second, third, fourth, fifth, or seventh main aspects unless expressly stated otherwise. Preferably the detection is done by using an immunological test, projection radiography, fluoroscopy, PET (Positron Emission Tomography) scans, PET -MR or PET- MRI (Magnetic Resonance), PET-CT, X-ray, CT (Computer Tomography) or CAT (Computer Axial Tomography) scans or combinations thereof. Seventh main aspect - Method for detection of ischemic body regions in a subject According to the seventh aspect, there is provided a method for detection of ischemic body regions in a subject using the labelled substance or the composition according to the present invention.

The ischemic body regions may for example be the result of ischemic reperfusion injury in kidney, intestines, extremities, and cardiovascular system.

Preferably the detection is done by using an immunological test, projection radiography, fluoroscopy, PET (Positron Emission Tomography) scans, PET -MR or PET- MRI (Magnetic Resonance), PET-CT, X-ray, CT (Computer Tomography) or CAT (Computer Axial Tomography) scans or combinations thereof. In ischemic models of both the kidney and hind limb, PVAC accumulated selectively in ischemic tissues. The ischemic kidney model had a high uptake of radiolabelled PVAC readily detectable by both PET scans and ex vivo biodistribution, while the ischemic limb model only showed a statistically significant increase ex vivo. Still, PET images of the hind limb displayed SUVs about twice the level of the control signifying that qualitative visual assessment could also be useful in tissues with low blood flow.

The elimination of P VAC through the kidneys followed a two-phase elimination curve with a rapid phase lasting approximately 2 hours followed by a slower phase with an accumulation in the bladder. The two-phase elimination could possibly be explained by the size range (15-35 kDa) of the PVAC molecules where the elimination through the kidneys would be more rapid in the smaller size range. Alternatively, PVAC may bind to circulating molecules, e.g. carrier proteins, which could slow down elimination of the bound fraction. Still, after 48 hours circulating levels were very low, suggesting a near complete elimination from the circulation.

Ex vivo biodistribution demonstrated that the majority of the labelled substance, PVAC, is excreted in urine with the remaining fraction confined to the vascular system. Among solid organs the uptake was highest in well-perfused organs such as kidneys, liver and lungs. In contrast, uptake in low-perfused organs such as skeletal muscle was substantially lower indicating that the signal more directly correlates to blood content ex vivo rather then specific uptake. In fact, the only specific uptake that was not apparently correlated to the level of blood content was observed in the ovaries, which displayed a notable uptake that increased over time. In all other non-ischemic organs, excluding the bladder, the uptake was diminished over time. A similar pattern was not seen in the male sex organs. Generally, administration of PVAC have been safe and our unpublished toxicity data have not shown any adverse events even after very high doses (50 mg/kg bw). As expected local administration in to the joint resulted in slower kinetics with less PVAC excreted in urine, signifying that a macro-molecule such as PVAC is retained in tissues.

Interestingly, the increased uptake in ischemic kidneys was effectively prevented by a previous administration of non-labelled PVAC. It is likely that the uptake of PVAC was saturated by injecting non-radiolabelled PVAC prior to administering radiolabelled PVAC, which indicates that PVACs binding to ischemic tissues was both specific and saturable. This is further supported by PVAC’s ability to effectively and almost completely scavenge reactive mediators formed during IRI.

All variants and examples of the seventh main aspect can be combined with the first, second, third, fourth, fifth, or sixth main aspects unless expressly stated otherwise. EXAMPLES

Materials and methods

Chemistry Carbazate substitution of PVAC was measured via by 2,4,6-trinitrobenzene sulfonic acid assay, the degree of substitution ( m ) (Fig. 1) was calculated to be 7.5%. The aldehyde scavenging capabilities are affected linearly when labelling is performed, this is due to both labelling and scavenging reactions utilize the carbazate group.

Monitoring of the reaction with the radiolabelled was achieved indirectly by tracking consumption of [¹⁸F]FBA at a wavelength of 254 nm. Reaction conditions were optimized and the maximum reaction speed was seen at pH 4.00. Radioactivity was tracked during synthesis of precursor and in the final product and losses were small with 80% of radioactivity reaching the final reaction. Isolation of PVAC-[¹⁸F]FBA from [¹⁸F]FBA was achieved by size exclusion chromatography, an exclusion limit of 5 kDa was used. PBS (pH 7.4) was used to elute from the column and equilibrate the solution prior to administration. Mean radioactivity in the final solution was 7.48 MBq for animals undergoing dynamic scan after I.V administration. Roughly 1 MBq was left in the syringe and the mean administered radioactivity was 6.08 MBq (5.04 - 7.13) or 29.33 (23.98 - 34.69) MBq/gbw. Animals

The studies were approved by the regional animal ethics committee in Uppsala (C56/16) and by regional ethics committee in Stockholm north (N81/14). All animals were treated in accordance with the recommendations from the National Guide for the Care and Use of Laboratory Animals. (17) The animals were housed for one week before surgery and maintained under standard laboratory conditions (+ 22°C, with a 12-h light/dark cycle and fed pellet food and water ad libitum).

Experimental design

In order to determine the kinetics and biodistribution of radioactive PVAC eleven more Sprague-Dawley rats, 8 female and 3 male, were after sedation injected with either I.V (tail vein, n= 9) or I. J (hind limb knee, n= 2) radiolabelled PVAC. Both I. J animals and five of the I.V animals were then placed in a PET camera. For 90 minutes a dynamic scan took place, followed by a CT scan and finally the animals were euthanized. The remaining four animals, 2 female 2 male, were sedated for 90 minutes. They were then euthanized, and organs recovered for the study of biodistribution.

Nine Sprague-Dawley rats, all female, were used in the ischemic models for kidney (n= 6) and hind limb (n= 3). There was no need for sham-operations since each animal could serve as their own internal control by having both a naive kidney and hind limb. Instead, in the ischemic kidney model three animals were blocked by injecting cold (unlabelled) PVAC prior to the injection of radiolabelled PVAC. All animals were then placed in a PET camera and underwent a 60 (kidney) or 90 (hind limb) minute dynamic scan, followed by a CT or MRI scan. They were then euthanized, and organs recovered for the study of biodistribution.

Radioactivity in the organs was measured using Nal well counter to determine the biodistribution of PVAC. These studies were performed at the preclinical PET -MRI platform, Uppsala.

The kinetic study of fluorochrome labelled PVAC was performed at Adlego, Stockholm where six male Sprague-Dawley rats were injected either I.V. (n=3, 2.5 mg/kg bw) or I.J. (n=3, 2.5 mg/animal) with fluorochrome labelled PVAC. Blood was drawn via tail vein before injection and at multiple time-points (5, 15, 30, 60 minutes, 2, 4, 6, 24, 48, 72, 96,168 hours) post injection, to later be stored frozen as serum. Urine was collected during 0-6, 6-24, 24-48, 48-72 and, 72-96 hours to later be stored frozen. The animals were euthanized at the end of the experiment.

Surgical procedure

Anesthesia was induced in a sealed chamber with inhaled 4% isoflurane and maintained using a facemask delivering 2.5% isoflurane. All procedures were performed under clean but nonsterile conditions. Both a heating lamp and a blanket were used to prevent heat loss during surgery and later ischemia. For the renal ischemia model an incision was placed under the left rib arc, through the skin, muscle and peritoneum, approximately 3 cm in length and a cotton swab was used to gently visualize the left kidney and the left renal artery. Ischemia was induced by placing a micro-clip on the left renal artery for 30 - 45 minutes. The clip was then removed and re-perfusion of the kidney was observed before closure of the wound using continuous resorbable 5-0 Vicryl® sutures (Johnson & Johnson AB, Sollentuna, Sweden) for the fascia, and continuous non-resorbable 4-0 Ethilon® sutures (Johnson & Johnson AB) for the skin. The surgical procedure took ~ 60- 75 min. For the hind limb ischemic model, a 2cm long incision was made through the skin in the femoral triangle to visualizing the femoral artery. Ischemia was achieved by placing a micro-clip on the femoral artery resulting in a whitening of the rats left foot. Ischemia was manifest for 45 - 60 minutes before the clip was removed and reperfusion occurred. The wound was closed using continuous resorbable 5-0 Vicryl® sutures (Johnson & Johnson AB, Sollentuna, Sweden) for the muscles, and continuous nonresorbable 4-0 Ethilon® sutures (Johnson & Johnson AB) for the skin. The surgical procedure took -75-90 min.

Imaging The ischemia and re-perfusion injury was followed by a single bolus injection (max volume 500 mΐ) of ¹⁸F-PVAC via a tail vein catheter, and the animal underwent small- animal PET examination of the organ area of interest for 90 minutes in list mode, followed by a CT examination for 3 minutes. The rats were kept sedated during the whole procedure by 3.0% isoflurane, blended with 450 ml/min air/0₂ (controlled by an anesthesia vaporizer) which were delivered through a face mask and placed on a heated bed of the PET-SPECT-CTsystem (TriumphTMTrimodality System, TriFoil Imaging, Inc., Northridge, CA, USA) to prevent hypothermia. Breathing rate and body temperature were monitored by an integrated physiologic monitoring system. A whole-body scan was performed by multiple bed positioning that lasted for 15 minutes. The dynamic datasets were reconstructed into 26 timeframes (12 frames of 10 sec, 3 frames of 1 min, 5 frames of 5 min, 6 frames of 10 min) using a maximum- likelihood expectation maximization 3- dimensional algorithm (10 iterations). Small-animal PET data were analysed using PMOD (version 3.510; PMOD Technologies Ltd. Switzerland) and Image J (Fiji, 2.0.0).

PET measurement was followed by MR examinations (in n= 2 rats from the renal ischemia model) with a 3 Tesla scanner (nanoScan, Mediso, Medical Imaging

Systems, Budapest, Hungary). Images were acquired in the coronal and axial planes over the region of interest using a T1 -weighted spin-echo sequence and whole body transmit/receiver coil. FOV 64x64 mm, acquisition matrix 256x192, slice thickness 1.3 mm, intersection gap 0.2 mm, resolution in plane 0.25 x 0.33 mm², bandwidth per pixel 156.25 Hz, number of signal averages 2 or 3, and echo time (TE) 11 ms. Time repetition

(TR) was between 540 and 880 ms depending on the number of slices.

Mechanistic studies in vitro

To study the kinetic of fluorochrome labelled PVAC a 7-point standard curve was made by dissolving FITC-PVAC in water. Frozen serum and urine samples were thawn and diluted 1:1 in water. Standards and samples were placed in a 96-well plate in duplicates. The samples were then exited at 485 nm and emission was registered at 528/20 nm using a Synergy HTX plate reader (BioTek). The data was exported and analysed in GraphPad prism, the standard curve was calculated using a 4-parameter logistic curve fit. Blank was subtracted for the standard curve and urine samples wells containing water was used for serum samples the value obtained prior to injection was used, the values were then interpolated and reported as concentration (pg/ml). For urine samples the total amount was also calculated by taking the concentration * the urine volume. To understand more about PVACs ability to scavenge known mediators of substances formed during IRI a Megazyme Acetaldehyde Assay Kit (Bray, Ireland) was used. The mediators assayed were oxidized proteins, methylglyoxal, malondialdehyde and acrolein (all obtained from Sigma). (21,22) The kit is designed to measure levels of acetaldehyde by using the physiological reaction with the enzyme aldehyde dehydrogenase (ALDH). In the reaction where aldehydes are reduced to carboxylic acids a side product NADH is formed which correlates to the present amount of IRI mediators. NADH can be quantified by measuring absorbance at 340 nm. The kit was used according to the manufacturer’s instructions with the addition of several targets for the enzyme and a preincubation step with or without PVAC. To produce an oxidized protein albumin (Sigma) was dissolved in PBS (1 ml,

100 mM) acrolein was added to the mixture to final concentration of 200 mM. The mixture was incubated for 30 minutes under stirring and then purified via dialysis (ThermoFisher, SnakeSkin dialysis tubing, 3500 MWCO). PVAC was dissolved in PBS (5 ml, 2.5 mg/ml) the mediators were dissolved in PBS (1 ml per mediator, 2.2 mM). Mediators and were mixed with PVAC or vehicle (PBS) in a 96-well microplate (100 mΐ mediator, 100 mΐ of PVAC/PBS) the plate was placed on a shaker (450 rpm) for 30 minutes. The mixture was then added to wells containing reaction buffers and NAD⁺ and baseline absorbance was measured. ALDH was added and the plate was incubated at 37°C until absorbance stabilized. The plate was read on a Synergy HTX plate reader (BioTek).

Statistics

Graphpad prism version 5.0c. (GraphPad Software, Inc, La Jolla, CA, USA) was used for all data handling and statistic analysis. Shapiro-Wilk normality test was performed to determine the nature of the data before any further analysis was made. In experiments with two groups: t-test (and none-parametric counterparts) was used, in experiments with >two groups: ANOVA was used. Experiments with two parameters, e.g. SUV and time: two-way ANOVA was calculated. All data in the results are described as mean difference, 95% Cl, p value and graphs are presented with error bars (SEM). Statistically significant p values are displayed and defined as <0.05, denoted by *.

EXAMPLE 1 - SYNTHESIS OF CARBAZATE- FUNCTIONALIZED POLYVINYL ALCOHOL (P AC)

To 0.50 g of polyvinyl alcohol (PVA, average molecular weight 16 kDa; Sigma Chemical Co., St Louis, MO) was added 20 ml dimethyl sulfoxide (DMSO, > 99.9%, Sigma). The mixture was heated at 80 °C for 15 min to completely dissolve PVA. After cooling to room temperature, 1.00 g of 1, 1 '-carbonyldiimidazole (CDI, > 97.0%, Sigma) was added and the mixture was stirred for 24 h. Hydrazine hydrate, 3.2 ml (80%, Sigma), was added to the mixture, which was left to stir for another 24 h. The reaction mixture was diluted with water and dialyzed (Spectra/Por ® 6 Dialysis Membrane, molecular weight cut-off, MWCO, 3.5 kDa) for three days and the product was isolated as a white powder after lyophilization.

Characterization: The degree of substitution of carbazate groups was investigated by 2,4,6-trinitrobenzene sulfonic acid, 5% (w/v) in H2O (Sigma) assay. One ml samples were dissolved in 20 ml sodium tetraborate decahydrate (> 99.5%, Sigma) buffer (pH 9.3, 0.1 M). From the prepared solutions, 1 ml was mixed with 25 pi 2,4,6- trinitrobenzene sulfonic acid solution. After three hours of reaction the mixture was analyzed by ultraviolet-visible spectroscopy at 505 nm and compared to a standard curve based on /c/V-butyl carbazate (> 98.0%, Sigma).

EXAMPLE 2 - LABELLING OF SUBSTANCE

All chemicals used in radiolabelling were bought from Sigma-Aldrich and used as received unless otherwise stated. Water used was MillieQ water 18.2 MW. For buffers pH was measured with a calibrated Mettler Toledo SevenEasy pH meter. Acetate buffers, 50 mM, were prepared by dissolving 0.205 g sodium acetate in 50 mL of water. The solution was fractionated and acetic acid was added until desired pH was achieved. 40 mM K2CO3 was prepared by dissolving 27.6 mg K2CO3 in 5000 pi water. 88 mM Kryptofix® 222 solution was prepared by dissolving 16.6 mg 4,7,13,16,21,24-hexaoxa- l,10-diazabicyclo[8.8.8]hexacosane (Kryptofix® 222) in 500 mΐ acetonitrile. 40 mM FBA-precursor solution was prepared from 6.3 mg (4-formylphenyl)trimethylammonium trifluoromethansulfonate in 0.50 mL anhydrous DMSO. Ethyl acetate was dried over 3 A molecular sieves (activated over night at 200°C) (19)

Radioactivity was measured in an ion chamber (Veenstra Instruments). Reactor vials were heated in a Reacti-Therm™ I aluminium heating block (Thermo Fisher Scientific) equipped with an external temperature probe immersed in a capped reference vial filled with DMSO placed next to the reaction vial. Sep-Pak® Light Alumina N cartridge (Waters) was preconditioned with ethanol (5 ml) and water (10 ml). Sep-Pak® Plus Short tC18 cartridge (Waters) was preconditioned with ethanol (5 ml) and water (10 ml). Illustra NAP™-5 column (GE Healthcare Life Sciences) was preconditioned with PBS (2 ml). High performance liquid chromatography, HPLC-UV-Radiosystem was a Knauer UV-detector with remote flow cell. Radioactivity was detected using a Bioscan flow count equipped with an extended range module FC 6106 (Eckert and Ziegler) and a PMT/Nal well detector FC-3300 (Eckert and Ziegler) which was attached in series after the UV-detector. A manual injector valve (Rheodyne) with a 100 mΐ loop was used. The HPLC apparatus was software controlled using OpenLAB (Agilent). [¹⁸F]Fluoride anion was produced at Uppsala PET Centre via the ¹⁸0(p,«)¹⁸F nuclear reaction using a cyclotron (Scanditronix MC-17, Uppsala, Sweden) equipped with a niobium or silver body target filled with ¹⁸0-enriched water (98%, Rotem GmbH, Leipzig, Germany). For the niobium target the beam current was 45 mA and a total bombardment of 1 pAh was typically used. For the silver target the beam current was 25 mA and a total bombardment of 5 pAh was typically used. The produced radioactivity was transferred from the cyclotron target using acetonitrile/water (35/65 v/v) and concentrated online on a Chromafix PS HCO₃ cartridge (Macherey-Nagel) before being transferred to a hotcell where the column was rinsed with water (5 ml).

For radionucleotide labelling of PVAC, 4-[¹⁸F] fluorobenzaldehyde (¹⁸FBA) was used. Synthesis of the precursor was carried out as described in the literature

(Speranza et al. , Appl. Radiat Isot, 2009, 67(9), 1664)) with some modifications. To 4- (dimethylamino) benzaldehyde (3.73 g, 35 mmol) in anhydrous ethyl acetate was added dropwise methyl trifluoro-methanesulfonate (1.81 g, 11.0 mmol). The reaction mixture was stirred under N2 at room temperature. The white precipitate was filtered and rinsed with anhydrous ethyl acetate and the residual solvent was removed under reduced pressure. The product was kept in the dark at +4°C until used for further experiments.

The radioactivity on the PS CO3 cartridge was eluted in one fraction directly into a 5 ml Thermo Reacti reaction vial with KiCCb-solution (0.4 ml, 40.0 mM) + 0.60 ml air over 60 seconds (<1% of the radioactivity remained on the cartridge). The solution was added to a vial containing a solution of Kryptofix® 222 (0.40 ml, 88 mM). The water was azeotropically removed at 110 °C by acetonitrile (3x0.5 ml) under a stream of nitrogen (100 ml-min ¹). Total drying time was between 20-25 min. The precursor solution (0.40 ml DMSO, 40 mM) in DMSO was added and the reaction was heated to 65 °C under nitrogen atmosphere (balloon) for 15 min. The reaction was quenched with water (2.5 ml) and the reaction vial was rinsed once (2.5 ml). The combined aqueous solution was loaded onto a SepPak AluN cartridge in series with a tC18 Plus cartridge over 90 seconds (<2% of the radioactivity remained in the reaction vial). The tC18 cartridge was detached and washed with 0.1 M HC1 (10 ml) and water (5 ml) over 60 seconds and dried with air. Finally, another AluN cartridge was attached in series after the tC18 cartridge and both were eluted with ethanol (1.0 ml) over 60 seconds into 0.20 ml fractions (<5% of the radioactivity remained on the SPE cartridge). Approx. 80% of radioactivity on the tC18Plus cartridge was collected in ethanol (0.60 mL). For HPLC- analysis 10 pi was withdrawn and diluted with 90 mΐ FEOMeCN (70/30 v/v). From the resulting solution 15 mΐ was injected for HPLC-UV-radio analysis. Freeze-dried PVAC 1 (2.8 mg) was dissolved in sodium acetate buffer (pH 4.0, 200 mΐ). To the solution was added 4-[¹⁸F]fluorobenzaldehyde 2 as a solution in ethanol (100 mΐ) and the reaction was allowed to stand in room temperature for 10 min before it was quenched with PBS (200 mΐ). The solution was added to a NAP™-5 column and eluted in 250 mΐ fractions with PBS. Radioactivity and pH of the resulting carbazate hydrazine conjugate 3 product solution was measured (Figure 2).

Fluorochrome labelled PVAC was produced via coupling of PVAC to fluorescein isothiocyanate (FITC) using the following protocol: 1 mg commercially available FITC (Sigma Aldrich) was incubated together with 100 mg PVAC for 2h with stirring to obtain o low level substitution PVAC -FITC (< 1%). The reaction mix was dialyzed against deionized water with stirring over night at room temperature. The final PVAC -FITC was finally freeze-dried and stored at -20°C until use. EXAMPLE 2 - ISCHEMIC PET MODELS

The ischemic kidney model had a higher uptake of the radiolabelled PVAC in the ischemic kidney compared to the non-ischemic kidney. Instead of an early peak during initial high circulating concentrations, the levels increased over time. The mean difference in standardized uptake value (SUV) at time point 60 minutes post-injection was 10.85 (8.1 - 15.8, =<0.0001). The three ischemic kidney animals that were blocked by injecting cold PVAC prior to radiolabelled PVAC had no difference in the SUV (p = 0.6432 ) when comparing the ischemic kidney to the non-ischemic kidney, tested at the same time point (60 minutes post-injection of radiolabelled PVAC). The graph showed no differences (p = 0.6432 ) between the ischemic kidney and the non-ischemic kidney over 60 minutes or later during the dynamic PET scan, figure 3g. Also the ex vivo measurements confirmed a difference between ischemic and non-ischemic kidneys (3.86, 1.5 - 6.2, p = 0.0095 ) and no difference (p = 0.9454 ) when the animals were blocked, figure 3h. Representative examples of PET scan shows an increased uptake in the ischemic kidney compared to the non-ischemic kidney but not when the animal was blocked, figure 3.

The ischemic limb model revealed no difference in uptake between the ischemic and non-ischemic limb during the dynamic scan, figure 4c. The ex vivo measurements showed an increased SUV in the ischemic limb muscle compared to a non ischemic limb, 0.134 (0.075 - 0.193 ,p = 0.0003). There was no difference (p = 0.0699 ) when comparing an ischemic limb muscle with a paired non-ischemic limb in the same animal, figure 4d.

In control animals, PVAC was mainly confined to the bloodstream followed by elimination via kidneys and accumulation in the bladder. Ex vivo biodistribution of PVAC confirmed highest uptake in urine followed by blood, kidneys and other well- perfused organs. The elimination of I.V administered PVAC was split into a fast phase (xl/2 = 0.2 h) followed by a slow phase (xl/2 = 10.73 h), with near complete elimination from blood after 48 hours. Both the ischemic kidney (fourfold increase, p = <0.001) and limb models (threefold increase, p = <0.001) demonstrated a higher uptake of PVAC in ischemic tissues, ex vivo radioactivity detection.

EXAMPLE 3 - PHARMACOKINETICS The pharmacokinetics of PVAC was studied in PET scans of rats injected (I.V or I.J) with either radiolabelled PVAC or fluorochrome labelled PVAC. Radiolabelled PVAC showed an uptake in the blood, the half-life (n_/2) in aorta was 10.2 minutes (1.9 - 40.8) and an increased uptake was seen in the bladder, n/2 for the bladder was 10.3 minutes (5.6 - 19.2), figure 5a. For fluorochrome labelled PVAC the elimination phase was split into: a fast phase and a slow phase. The fast phase (50% of the elimination) had a n/2 of 0.2 hours (0.11 - 0.33). The slow phase had a n/2 of 10.73 hours (7.1 - 15). I.J injection gave rapidly increased serum concentrations followed by a steady state at 2 hours between elimination and absorption and at six hours elimination was dominant leading to a n_/2of 34.90 hours (24.92 - 51.45), figure 5b. Video shows the initial PVAC uptake after I.V. injection, which is limited to the aorta before the kidneys start to light up. The kidneys slowly excrete the substance and finally the bladder starts to increase in intensity, figure 5c. I.J injection instead showed localized uptake in the joint and low levels in the bladder and kidneys, figure 5d,e. Total excreted PVAC in gathered urine (at 96 h) was 37.75% (17.58 - 57.91) after I.V injection and 20.30% (13.39 - 27.21) after I.J injection, figure

5f.

EXAMPLE 4 - BIODISTRIBUTION

The ex vivo biodistribution after injection of radiolabelled PVAC had the highest uptake in urine (60.5 mean SUV ± 15.5) followed by blood (6.7 mean SUV ± 1.1) and kidneys (2.8 mean SUV ± 0.3). Well perfused organs such as the lungs (2.1 mean SUV ± 0.3), liver (1.7 mean SUV ± 0.3) and spleen (0.7 mean SUV ± 0.1) had a higher uptake than low perfusion organs such as muscle (0.1 mean SUV ± 0) and brain (0.2 mean SUV ± 0), figure 6. The differences between sexes were also studied, there was no difference between any organ system apart from the reproduction organs (p = 0.0446 ), with a higher uptake in ovaries compared to testis.

EXAMPLE 5 - INVESTIGATION OF AFFINITY TOWARDS IRI MEDIATORS Different reactive mediators formed during IRI (oxidized albumin, methylglyoxal, malondialdehyde, acrolein) were oxidised using ALDH. The IRI mediators that were pre incubated with PVAC showed a reduced signal: 79.23% (p = 0.0001 ) for free oxidized albumin, 100% (p = 0.0015 ) for methylglyoxal, 88.68% (p = 0.0022 ) for malondialdehyde and 99.33% (p = 0.0073 ) for acrolein, fig 7. A reduced signal in the absorbance at 340 nm equals a lower concentration of free IRI mediators, since the rest were bound to PVAC.

While the invention has been described and pointed out with reference to operative examples thereof, it will be understood by those skilled in the art that various changes, modifications, substitutions and omissions can be made without departing from the spirit of the invention.

Claims

C L A I M S

1. A labelled substance, wherein the substance comprises a carrier exhibiting at least one scavenger structure, said scavenger structure comprising a nucleophilic centre complying with the formula

X¹(-R”-)(-R^,)mH_n (formula I) wherein a) X¹ is a single-bonded heteroatom selected amongst N, O and S and exhibits a free electron pair, b) m is 0 or 1 and n is 1 or 2 with the sum of m plus n being 2 for X¹ = N and 1 for X¹ = S and O, c) -R”- is a bivalent organic group providing attachment to the carrier via one of its free valences and direct attachment to the heteroatom X¹ at the other one of its free valences, and d) R’ - is a monovalent organic group directly attached to the heteroatom X¹ via its free valence, and wherein in that the nucleophilic center is part of a group selected amongst hydrazide groups, preferably-NH-ML, preferably as part of a -CONHNH2 group, a semicarbazide group, preferably -NHCONHNH2, a carbazate group, preferably

OCONHNH2, a thiosemicarbazide group, preferably -NHCSNHNH2, a thiocarbazate group, preferably -OCSNHNH2; and wherein the carrier is a water-soluble macromolecular carrier preferably selected from polyvinyl alcohol or polysaccharide wherein said polysaccharide is preferably selected from dextran, starch, agarose, agaropektin, cellulose or glucose amino glucan (GAG) preferably hyaluronic acid; wherein the substance further comprises a chemical entity comprising a detectable label.

2 The substance according to any of the preceding claims, wherein in that the carrier exhibits polymer structure.

3. The substance according to any of the preceding claims, wherein the carrier is polyvinyl alcohol.

4. The substance according to any of the preceding claims, wherein the scavenger is a carbazate group.

5. The substance according to claim 1, wherein in that the substance is carbazate- functionalized polyvinyl alcohol.

6. The substance according to anyone of the preceding claims, wherein the detectable label is selected from radioactive labels, non-radioactive contrast agent fluorochrome or fluorophores.

7. The substance according to claim 6 wherein the detectable label is radioactive and comprises a radioactive isotope selected from ^UC, ¹³N, ¹⁵0, ¹⁸F, ⁶⁸Ga, ⁸⁹Zr, and ⁸²Rb, preferably [ 18F]fluorobenzaldehyde.

8. The substance according to anyone of claims 1 to 7, wherein 0.1-7.5%, preferably

0.1-5% of the scavenger structures such as the nucleophilic centres are labelled.

9. The substance according to claim 6, wherein the detectable label is fluorochrome and wherein said fluorochrome is fluorescein isothiocyanate.

10. The substance according to claim 6 wherein the detectable label is a non-radioactive contrast agent wherein said agent is iodine, barium or gadolinium based.

11. A composition comprising a detectable amount of the labeled substance defined in anyone of claims 1-10 and diagnostically suitable excipients.

12. A labelled substance according to anyone of claims 1 to 11 for use in detection of inflammation body regions, infectious body regions, ischemic tissue or body region, malignant body regions or metastasis.

13. The labelled substance according to claim 12 wherein the inflammation is a local inflammation.

14. A method for detection of inflammatory body regions in a subject, comprising the steps of a. administering a detectable amount of the labelled substance defined in anyone of claims 1-10 or the composition defined in claim 11 to the subject, b. allowing the labelled substance to bind to an inflammatory site, and c. detecting the signal from the labelled substance bound to the inflammatory site.

15. The method according to claim 14, where the substance is administered intravenously.

16. The method according to claim 14 or 15, wherein the inflammatory region is inflammatory intestines, liver, pancreas, spleen, lung, muscles, tendons, joints, blood vessels, oral cavities, subcutaneous fat, and tissue surrounding implanted implants.

17. The method according to claim 14 or 16, wherein the inflammatory region is atherosclerotic plaques in blood vessels.

18. A method for detection of infectious body regions in a subject, comprising the steps of: a. administering a detectable amount of the labelled substance defined in anyone of claims 1-10 or the composition defined in claim 11 to the subject, b. allowing the labelled substance to bind to an inflammatory site, and c. detecting the signal from the labelled substance bound to the inflammatory site; and d. detecting the infectious body regions located in or in the vicinity of said binding site.

19. The method according to claim 18, wherein said infectious region is selected from intrabdominal muscle, muscle fascia, joint, and lung.

20. A method for detection of malignant body regions in a subject, comprising the steps of: a. administering a detectable amount of the labelled substance defined in anyone of claims 1-10 or the composition defined in claim 11 to the subject, b. allowing the labelled substance to bind to an inflammatory site, and c. detecting the signal from the labelled substance bound to the inflammatory site; and d. detecting the malignant body regions located in or in the vicinity of said binding site.

21. The method according to claim 20, wherein said malignant body regions is a cancer tumour or metastasis.

22. A method for detection of ischemic body regions in a subject, comprising the steps of: a. administering a detectable amount of the labelled substance defined in anyone of claims 1-10 or the composition defined in claim 11 to the subject, b. allowing the labelled substance to bind to an inflammatory site, and c. detecting the signal from the labelled substance bound to the inflammatory site; and d. detecting the ischemic body regions located in or in the vicinity of said binding site.

23. The method according to claim 22, wherein the ischemic region is the result of ischemic reperfusion injury in kidney, intestines, extremities, and cardiovascular system.

24. The method according to claim 14 or 22, wherein the subject is a mammal.

25. The method according to anyone of claims 14-24, wherein the detection of the signal from the labelled substance is achieved by immunological test, projection radiography, fluoroscopy, PET (Positron Emission Tomography) scans, PET -MR or PET-MRI (Magnetic Resonance), PET-CT, X-ray, CT (Computer Tomography) or CAT (Computer Axial Tomography) scans or combinations thereof.