FLUORESCENT POLYMERIC COATING FILM FOR MEDICAL DEVICES

Abstract

A group of fluorescent polymeric coating films visible in near-infrared light for coating a medical device, and a method for preparing such NIR visible polymeric coating films.

Claims

1. A fluorescent polymeric coating film visible in near-infrared light for coating a medical device, said coating film being a single layer or multiple layers of hydrophobic polymer, and at least one polymeric layer comprising a fluorescent dye and a counterion, said fluorescent dye being represented by Formula I ##STR00040## wherein: R.sub.8 and R.sub.9 are the same or different and independently selected from a group consisting of: an hydrogen, or a group chosen from a (C1-C20)alkyl, a cyclo(C3-C20)alkyl, a (C2-C20)alkenyl, a (C2-C20)alkynyl, a heterocyclic group, a cyclo(C3-C20)alkenyl, a heterocyclo(C2-C20)alkenyl, an aryl, a heteroaryl, a hetero(C1-C20)alkyl, a (C1-C20)alkylaryl, or a (C1 C20)alkylheteroaryl, said group being unsubstituted or substituted by one or two substituents chosen from a (C1-C5) alkyl, an aryl, or COOR.sub.11, R.sub.11 being a (C1-C20) alkyl, or a group of formula E-R.sub.10, wherein E is chosen from O, S, Se, NH, CH.sub.2; R.sub.10 is chosen from a (C1-C20)alkyl, a cyclo(C3-C20)alkyl, a (C2-C20)alkenyl, a (C2-C20)alkynyl, a heterocyclic group, a cyclo(C3-C20)alkenyl, a heterocyclo(C2-C20)alkenyl, an aryl, a heteroaryl, a hetero(C1-C20)alkyl, a (C1-C20)alkylaryl, a (C1-C20)alkylheteroaryl, R.sub.10 being unsubstituted or substituted by one to three substituents chosen from a (C1-C5) alkyl, an aryl, or COOR.sub.11, R.sub.11 being a (C1-C20) alkyl A.sub.1 is ##STR00041## and A.sub.2 is ##STR00042## or A.sub.1 is ##STR00043## and A.sub.2 is ##STR00044## wherein: R.sub.2 and R.sub.4 are independently selected from the group consisting of hydrogen, halogen, (C1-C10)alkyl, OR.sub.5, NR.sub.5R.sub.6, NO.sub.2, CF.sub.3, CN, SR.sub.5, N.sub.3, C(O)R.sub.5, OC()OR.sub.5, C(O)NR.sub.5R.sub.6, NR.sub.5C(O)R.sub.6, wherein R.sub.5 and R.sub.6 are independently selected from hydrogen, unsubstituted (C1-C10)alkyl, unsubstituted (C2-C10)alkenyl, unsubstituted (C2-C10)alkynyl, cyclo(C3-C10)alkyl, heterocyclic group, cyclo(C3-C10)alkenyl, heterocyclo(C2-C10)alkenyl, aryl, heteroaryl, aryl(C1-C10)alkyl, hetero(C1-C10)alkyl, (C1-C10)alkylaryl, (C1-C10)alkylheteroaryl; R.sub.1 and R.sub.3 are independently selected from the group consisting of a (C1-C20)alkyl eventually substituted by a hydrophobic group, a cyclo(C3-C20)alkyl eventually substituted by a hydrophobic group, a (C2-C20)alkenyl eventually substituted by a hydrophobic group, a (C2-C20)alkynyl, a heterocyclic group eventually substituted by a hydrophobic group, a cyclo(C3-C20)alkenyl eventually substituted by a hydrophobic group, a heterocyclo(C2-C20)alkenyl eventually substituted by a hydrophobic group, an aryl eventually substituted by a hydrophobic group, a heteroaryl eventually substituted by a hydrophobic group, a hetero(C1-C20)alkyl eventually substituted by a hydrophobic group, a (C1-C20)alkylaryl eventually substituted by a hydrophobic group, a (C1-C20)alkylheteroaryl eventually substituted by a hydrophobic group, said hydrophobic group being selected from methyl, ethyl, methoxy, ethyloxy; said hydrophobic polymer is chosen from poly(methyl methacrylate), poly(ethyl methacrylate), poly(propyl methacrylate), poly(butyl methacrylate), poly(methyl methacrylate-co-methacrylic acid), poly(lactide-co-glycolide), polylactic acid, polyglycolic acid, polycaprolacton, cellulose triacetate, nitrocellulose, polydimethylsiloxane, poly(ethylene terephthalate), polycarbonate, polyethylene, ethylene vinyl acetate copolymer, polyurethane, polystyrene, and copolymers thereof with poly(ethylene glycol).

2. The fluorescent polymeric coating film according to claim 1, the outermost polymeric layer is based on a biocompatible hydrophobic polymer.

3. The fluorescent polymeric coating film according claim 1, wherein the counterion is chosen from: an inorganic counterion, an organic counterion, or a bulky organic counterion chosen from tetraphenylborate, tetrakis(pentafluorophenyl)borate, tetrakis(4-fluorophenyl)borate, tetraphenylborate, tetrakis[3,5-bis-(trifluoromethyl)phenyl]borate, tetrakis[3,5-bis-(1,1,1,3,3,3-hexafluoro-2-methoxy-2-propyl)phenyl]borate and tetrakis[perfluoro-tert-butoxy]aluminate.

4. The fluorescent polymeric coating film according to claim 1, wherein the fluorescent dye is represented by the formula II: ##STR00045## wherein: A.sub.1, A.sub.2, R.sub.9, and E are as defined in claim 1; R.sub.12 and R.sub.13 are the same or different and independently selected from an hydrogen, an aryl, a (C1-C5) alkyl, or COOR.sub.11, R.sub.11 being a (C1-C20) alkyl.

5. The fluorescent polymeric coating film according to claim 4, wherein the fluorescent dye is represented by the formula II(a): ##STR00046## wherein: A.sub.1, A.sub.2, R.sub.9, and E are as defined in claim 1; R.sub.12 and R.sub.13 are the same or different and independently selected from an aryl; the counterion for said fluorescent dye is an inorganic counterion.

6. The fluorescent polymeric coating film according to claim 1, formed by a hydrophobic polymer, in particular a biocompatible hydrophobic polymer, an ionic fluorescent dye and a counterion, said fluorescent dye being represented by Formula I ##STR00047## wherein: R.sub.8 and R.sub.9 are the same or different and independently selected from the group consisting of an hydrogen, an unsubstituted (C1-C20)alkyl, an unsubstituted cyclo(C3-C20)alkyl, an unsubstituted (C2-C20)alkenyl, an unsubstituted (C2-C20)alkynyl, an unsubstituted heterocyclic group, an unsubstituted cyclo(C3-C20)alkenyl, an unsubstituted heterocyclo(C2-C20)alkenyl, an unsubstituted aryl, an unsubstituted heteroaryl, an unsubstituted hetero(C1-C20)alkyl, an unsubstituted (C1-C20)alkylaryl, an unsubstituted (C1-C20)alkylheteroaryl; A.sub.1 is ##STR00048## and A.sub.2 is ##STR00049## or A.sub.1 is ##STR00050## and A.sub.2 is ##STR00051## wherein: R.sub.2 and R.sub.4 are independently selected from the group consisting of hydrogen, halogen, (C1-C10)alkyl, OR.sub.5, NR.sub.5R.sub.6, NO.sub.2, CF.sub.3, CN, SR.sub.5, N.sub.3, C(O)R.sub.5, OC()OR.sub.5, C(O)NR.sub.5R.sub.6, NR.sub.5C(O)R.sub.6, wherein R.sub.5 and R.sub.6 are independently selected from hydrogen, unsubstituted (C1-C10)alkyl, unsubstituted (C2-C10)alkenyl, unsubstituted (C2-C10)alkynyl, cyclo(C3-C10)alkyl, heterocyclic group, cyclo(C3-C10)alkenyl, heterocyclo(C2-C10)alkenyl, aryl, heteroaryl, aryl(C1-C10)alkyl, hetero(C1-C10)alkyl, (C1-C10)alkylaryl, (C1-C10)alkylheteroaryl; R.sub.1 and R.sub.3 are independently selected from the group consisting of a (C1-C20)alkyl eventually substituted by a hydrophobic group, a cyclo(C3-C20)alkyl eventually substituted by a hydrophobic group, a (C2-C20)alkenyl eventually substituted by a hydrophobic group, a (C2-C20)alkynyl, a heterocyclic group eventually substituted by a hydrophobic group, a cyclo(C3-C20)alkenyl eventually substituted by a hydrophobic group, a heterocyclo(C2-C20)alkenyl eventually substituted by a hydrophobic group, an aryl eventually substituted by a hydrophobic group, a heteroaryl eventually substituted by a hydrophobic group, a hetero(C1-C20)alkyl eventually substituted by a hydrophobic group, a (C1-C20)alkylaryl eventually substituted by a hydrophobic group, a (C1-C20)alkylheteroaryl eventually substituted by a hydrophobic group, said hydrophobic group being selected from methyl, ethyl, methoxy, ethyloxy; said counterion being an inorganic counterion, or an organic counterion or a bulky organic counterion.

7. The fluorescent polymeric coating film according to claim 1, wherein A.sub.1 is ##STR00052## and A.sub.2 is ##STR00053## R.sub.1 and R.sub.3 being defined as claim 1.

8. The fluorescent polymeric coating film according claim 1, wherein R.sub.1 and R.sub.3 are independently selected from the group consisting of unsubstituted (C1-C20)alkyl, an unsubstituted cyclo(C3-C20)alkyl, an unsubstituted (C2-C20)alkenyl, an unsubstituted (C2-C20)alkynyl, an unsubstituted heterocyclic group, an unsubstituted cyclo(C3-C20)alkenyl, an unsubstituted heterocyclo(C2-C20)alkenyl, an unsubstituted aryl, an unsubstituted heteroaryl, an unsubstituted hetero(C1-C20)alkyl, an unsubstituted (C1-C20)alkylaryl, an unsubstituted (C1-C20)alkylheteroaryl.

9. The fluorescent polymeric coating film according to claim 1, wherein R.sub.1 and R.sub.3 are independently selected from the group of unsubstituted (C8-C20)alkyl, particularly the group of unsubstituted (C12-C18)alkyl.

10. The fluorescent polymeric coating film according to claim 1, wherein the fluorescent dye is of Formula Ia ##STR00054## or of Formula Ib ##STR00055## R.sub.1 and R.sub.3 being defined as claim 8.

11. The fluorescent polymeric coating film according to claim 1, wherein the fluorescent dye is ##STR00056##

12. The fluorescent polymeric coating film according claim 1, wherein the weight of fluorescent dye in a polymeric layer is from 0.1 to 50, particularly from 0.5 to 10%, still more particularly 1%, by weight of the hydrophobic polymer in said polymeric layer.

13. A method for coating a medical device by a fluorescent polymeric coating film visible in near-infrared light as defined according to claim 1, comprising: (i) a step of contacting said medical device with a solution comprising one or several solvents, a hydrophobic polymer, an ionic fluorescent dye and a counterion, said hydrophobic polymer being chosen from poly(methyl methacrylate), poly(ethyl methacrylate), poly(propyl methacrylate), poly(butyl methacrylate), poly(methyl methacrylate-co-methacrylic acid), poly(lactide-co-glycolide), polylactic acid, polyglycolic acid, polycaprolacton, cellulose triacetate, nitrocellulose, polydimethylsiloxane, poly(ethylene terephthalate), polycarbonate, polyethylene, ethylene vinyl acetate copolymer, polyurethane, polystyrene, and copolymers thereof with poly(ethylene glycol); said fluorescent dye being represented by Formula I ##STR00057## wherein: R.sub.8 and R.sub.9 are the same or different and independently selected from the group consisting of: an hydrogen, a group chosen from a (C1-C20)alkyl, a cyclo(C3-C20)alkyl, a (C2-C20)alkenyl, a (C2-C20)alkynyl, a heterocyclic group, a cyclo(C3-C20)alkenyl, a heterocyclo(C2-C20)alkenyl, an aryl, a heteroaryl, a hetero(C1-C20)alkyl, a (C1-C20)alkylaryl, or a (C1 C20)alkylheteroaryl, said group being unsubstituted or substituted by one or two substituents chosen from a (C1-C5) alkyl, an aryl, or COOR.sub.11, R.sub.11 being a (C1-C20) alkyl, or a group of formula E-R.sub.11, wherein E is chosen from O, S, Se, NH, CH.sub.2; R.sub.10 is chosen from a (C1-C20)alkyl, a cyclo(C3-C20)alkyl, a (C2-C20)alkenyl, a (C2-C20)alkynyl, a heterocyclic group, a cyclo(C3-C20)alkenyl, a heterocyclo(C2-C20)alkenyl, an aryl, a heteroaryl, a hetero(C1-C20)alkyl, a (C1-C20)alkylaryl, a (C1 C20)alkylheteroaryl, R.sub.10 being unsubstituted or substituted by one or two substituents chosen from a (C1-C5) alkyl, an aryl, or COOR.sub.11, R.sub.11 being a (C1-C20) alkyl A.sub.1 is ##STR00058## and A.sub.2 is ##STR00059## A.sub.1 is ##STR00060## and A.sub.2 is ##STR00061## wherein: R.sub.2 and R.sub.4 are independently selected from the group consisting of hydrogen, halogen, (C1-C10)alkyl, OR.sub.5, NR.sub.5R.sub.6, NO.sub.2, CF.sub.3, CN, SR.sub.5, N.sub.3, C(O)R.sub.5, OC()OR.sub.5, C(O)NR.sub.5R.sub.6, NR.sub.5C(O)R.sub.6, wherein R.sub.5 and R.sub.6 are independently selected from hydrogen, unsubstituted (C1-C10)alkyl, unsubstituted (C2-C10)alkenyl, unsubstituted (C2-C10)alkynyl, cyclo(C3-C10)alkyl, heterocyclic group, cyclo(C3-C10)alkenyl, heterocyclo(C2-C10)alkenyl, aryl, heteroaryl, aryl(C1-C10)alkyl, hetero(C1-C10)alkyl, (C1-C10)alkylaryl, (C1-C10)alkylheteroaryl; R.sub.1 and R.sub.3 are independently selected from the group consisting of a (C1-C20)alkyl eventually substituted by a hydrophobic group, a cyclo(C3-C20)alkyl eventually substituted by an hydrophobic group, a (C2-C20)alkenyl eventually substituted by a hydrophobic group, a (C2-C20)alkynyl, a heterocyclic group eventually substituted by a hydrophobic group, a cyclo(C3-C20)alkenyl eventually substituted by a hydrophobic group, a heterocyclo(C2-C20)alkenyl eventually substituted by a hydrophobic group, an aryl eventually substituted by a hydrophobic group, a heteroaryl eventually substituted by a hydrophobic group, a hetero(C1-C20)alkyl eventually substituted by a hydrophobic group, a (C1-C20)alkylaryl eventually substituted by a hydrophobic group, a (C1-C20)alkylheteroaryl eventually substituted by a hydrophobic group, said hydrophobic group being selected from methyl, ethyl, methoxy, ethyloxy; said counterion being an inorganic counterion, or an organic counterion or a bulky organic counterion; (ii) Evaporating the solvent in said solution to form one polymeric layer comprising a fluorescent dye and a counterion on said medical device.

14. The method according to claim 13, wherein said coating film is multi-layered, said method comprises step (iii) and step (iv) after the step (ii): (iii) Contacting coated medical device obtained in step (ii) with a solution comprising one or several solvents and a hydrophobic polymer and eventually devoid of any fluorescent dye; (iv) Evaporating the solvent in said solution to produce a polymeric layer on said medical device.

15. The method according to claim 13, wherein said solution contains an organic solvent, in particular chosen from acetonitrile, acetone, tetrahydrofuran, dioxane, ethyl acetate, propyl acetate, butyl acetate, amyl acetate, diethyl ether, 1,2-dimethoxyethane, 1,2-dioxane, dichloromethane, chloroform, petroleum ether, toluene, o-xylene, m-xylene, p-xylene.

16. The method according to claim 13, wherein the hydrophobic polymer is from 1 to 100 mg/ml, preferably 30 mg/ml of solution; the weight of fluorescent dye in a polymeric layer is from 0.1 to 50%, particularly from 0.5 to 10%, still more particularly 1%, by weight of the hydrophobic polymer in said polymeric layer.

17. The method according to claim 13, wherein the medical device is chosen from a surgical packing, fiducial marker, a surgical tap, a needle, a surgical retractor, a clip, a surgical thread, a staple, a knife, a safety pin, a scissor, a clamp, a scalpel, a hemostat, a tweezer, a forcep, a suction tip, a gauze, a cottonoid, a sponge, a catheter, a stainless steel wire, a surgical site marker, a stent, a pacemaker, a nerve stimulator, a drug delivery system, a port-a-cath, a magnetic anastomosis system, an intravascular embolization device, a mechanical linear stapler, a mechanical circular stapler, a drug-delivery implant, a piezoelectric implant, a diagnostic video-capsule, a magnetic tracking capsule.

18. The method according to claim 13, wherein the steps (i) and (ii) are repeated from 1 to 50 times, preferably 1 to 20.

19. The method according to claim 13, wherein the steps (iii) and (iv) are repeated from 1 to 20 times.

Description

FIGURES

[0162] FIG. 1 concerns photos of coated stainless steel wires taken by a fluorescent image system. These wires are respectively coated 1, 2, 3 or 5 times by a coating film of the invention (New, Cy7.5-C18-TPB-based coating) or by three different ICG-based coating films (ICG, ICG salt with tetraphenyl phosphonium counterion (ICG-P), ICG salt with tetrabutyl ammonium counterion (ICG-N)).

[0163] FIG. 2 displays photos by a fluorescent image system of coated silicon pieces taken just after coating procedure (initial, before drying) or after 30 s, 60 s or 90 s of drying. These pieces are respectively coated 1 time by a coating film of the invention (New, Cy7.5-C18-TPB-based coating) or 5 times by three different ICG-based coating films (ICG, ICG-P, ICG-N).

[0164] FIG. 3 displays photos of coated surgical thread taken just after coating procedure (basal) or one week after coating by a fluorescent image system. These surgical thread are respectively coated by a coating film of the invention (New, Cy7.5-C18-TPB-based coating) or by ICG-based coating films (ICG, ICG-P, ICG-N).

[0165] FIG. 4 shows that the ureter (indicated by 2 arrows) during a image-guided surgery is identified by an inserted catheter, which is coated by Cy7.5-C18-TPB-based coating film and visible via fluorescent imaging system.

[0166] FIG. 5 shows that a tip of naso-gastric tube coated by Cy7.5-C18-TPB-based coating film can be visualized during surgery (indicated by 2 arrows).

[0167] FIG. 6 shows that four-corners of the pathologic lesion of the stomach were marked by insertion of the marker coated by Cy7.5-C18-TPB-based coating film (indicated by 4 arrows).

[0168] FIG. 7 shows that gauze pieces coated by Cy7.5-C18-TPB-based coating film and plunged by blood during a surgery is identified in NIR via a fluorescent imaging system.

[0169] FIG. 8 shows synthetic protocols for CY7.5 dye and its derivatives of formula I.

[0170] FIG. 9 shows fluorescence images of the coating films containing different dyes of the present invention deposited on the glass surface.

[0171] FIG. 10 (A) shows fluorescence images of coating films of the invention based on fluorescent dye CY7.5-C18-TPB, HA-06-I, HA-60-TPB and different hydrophobic polymers: PMMA, PLGA or PLC. The coating films based on ICG are prepared as control. FIG. 10 (B) shows emission intensity values obtained from the images of the thin films containing different types of fluorescent dyes and polymers.

[0172] FIG. 11 shows tissue imaging with steel fiducials coated with different dyes (1% loading) in PMMA: (a) Chicken tissue under normal light; (b) fluorescence image of the tissue; (c) Fluorescence images of the coated fiducials inserted into the chicken tissues (1 for HA-06-I, 2 for HA-60-TPB, 3 for Cy7.5-C18-TPB).

EXAMPLES

1. Materials and Methods

[0173] 1.1 Synthesis of Cy7.5-C18-TPB

[0174] Fluorescent dye Cy7.5-C18-TPB of structure before:

##STR00031##

is synthesized according to protocol below and summarized in schema 1(a) of FIG. 8.

1.1.2-trimethyl-3-octadecyl-1H-benzo[e]indol-3-ium iodide

[0175] 250 mL round-bottom flask equipped with magnetic stirring bar was charged with 1,1,2-Trimethylbenz[e]indole (1 eq., 6.88 g, 32.9 mmol) and 1-iodooctadecane (2 eq., 25 g, 65.7 mmol), 100 mL of 2-butanone was added subsequently. Reaction mixture was refluxed for 24 h, then cooled down to r.t.

[0176] Reaction mixture was cooled down to r.t., diethyl ether was added and formed solid part was filtered off and washed with 100 mL of diethyl ether. Obtained crystals of crude product were redissolved in DCM and precipitated back while by adding diethyl ether, afterwards filtered and washed with diethyl ether. Product was obtained as slightly green crystals in 76% yield (14.73 g).

[0177] .sup.1H NMR (400 MHz, CDCl.sub.3) 8.10 (d, J=8.7 Hz, 1H), 8.08 (dd, J=8.0, 1.1 Hz, 1H), 8.04 (dd, J=8.2, 1.3 Hz, 1H), 7.76 (d, J=8.9 Hz, 1H), 7.72 (ddd, J=8.3, 6.9, 1.4 Hz, 1H), 7.65 (ddd, J=8.1, 6.9, 1.2 Hz, 1H), 4.78 (t, J=7.7 Hz, 2H), 3.19 (s, 3H), 1.97 (p, J=7.8 Hz, 2H), 1.87 (s, 6H), 1.52-1.41 (m, 2H), 1.40-1.30 (m, 2H), 1.28-1.19 (m, 26H), 0.85 (t, J=7.0 Hz, 3H).

[0178] .sup.13C NMR (100 MHz, CDCl.sub.3) 195.25, 138.34, 137.29, 133.82, 131.56, 130.17, 128.77, 127.97, 127.74, 122.96, 112.59, 56.04, 50.56, 32.00, 29.77, 29.76, 29.73, 29.70, 29.65, 29.55, 29.43, 29.41, 29.24, 28.26, 26.92, 22.85, 22.76, 17.03, 14.18.

[0179] HRMS (m/z): [M].sup.+ calculated for C.sub.33H.sub.52N 462.40943; found 462.40854.

Dioctadecylcyanine 7.5 Chloride:

[0180] 1,1,2-trimethyl-3-octadecyl-1H-benzo[e]indol-3-ium iodide (1) (2.2 eq., 1029 mg, 1.75 mmol) and glutaconaldehydedianil hydrochloride (1 eq., 226 mg, 0.794 mmol) were mixed in 10 ml of pyridine, afterwards Ac.sub.2O (13.4 eq., 1087 mg, 1 mL, 10.6 mmol) was added and the reaction mixture was heated to 60 C. while stirring and left for 3 h. After reaction was finished, solvents were evaporated at vacuum, and the crude product was dissolved in DCM, washed with 0.1 N HCl (3 times), brine and water. DCM layer was dried over Na.sub.2SO.sub.4, the solvent was evaporated and the product was purified by column chromatography on silica (gradient DCM/MeOH 99/1-95/5). Product was obtained as a green solid (926 mg, 0.906 mmol, 52%)

[0181] .sup.1H NMR (400 MHz, CDCl.sub.3): 8.16 (d, 1=8 Hz, 2H), 8.06 (t, J=12 Hz, 2H), 7.97 (bs, 1H), 7.92 (d, J=8 Hz, 4H), 7.61 (t, J=7 Hz, 2H), 7.46 (t, J=7 Hz, 2H), 7.35 (d, J=9 Hz, 2H), 6.67 (t, J=12 Hz, 2H), 6.25 (d, J=12 Hz, 2H), 4.14 (bs, 4H), 2.04 (s, 12H), 1.87 (m, J=7 Hz, 4H), 1.49 (m, J=7 Hz, 4H), 1.39 (m, J=7 Hz, 4H), 1.26 (bs, 52H), 0.88 (t, J=7 Hz, 6H)

[0182] .sup.13C NMR (100 MHz, CDCl.sub.3): 173.07, 157.04, 151.02, 139.74, 133.98, 131.83, 130.59, 130.09, 128.41, 127.87, 126.27, 125.06, 122.47, 110.56, 103.39, 51.155, 44.76, 32.06, 29.842, 29.807, 29.76, 29.72, 29.60, 29.53, 29.50, 27.86, 27.14, 22.82, 14.26

[0183] HRMS (m/z): [M].sup.+ calculated. for C.sub.71H.sub.105N.sub.2.sup.+ 985.8272; found 985.8290.

Dioctadecylcyanine 7.5 tetraphenylborate (Cv7.5-C18-TPB):

[0184] Dioctadecylcyanine 7.5 chloride (1 eq., 200 mg, 0.18 mmol) was dissolved in 5 mL of DCM, sodium tetraphenylborate (3 eq., 184 mg, 0.539 mmol) was added and the dispersion was sonicated for 5 min. TLC control has shown full conversion. Afterwards, the mixture was purified on a silica column, eluent DCM/MeOH 95/5 (product goes almost with front). Dioctadecylcyanine 7.5 tetraphenylborate (218 mg, 0.167 mmol, 93%) was obtained as green viscous oil and used without further characterisation.

Synthesis of CY 7.5-C8-I:

[0185] ##STR00032##

[0186] CY 7.5 C8 was synthesized by following our previous method. 1,1,2-trimethyl-3-octyl-1H-benzo[e]indol-3-ium iodide (2 equiv.), glutaconaldehydedianil hydrochloride (1 equiv.) were mixed in pyridine. Acetic anhydride (13.4 equiv.) was added and stirred at 60 C. for 3 hours. Reaction was monitored by TLC. Upon completion, solvent was evaporated under vacuum, dissolved in DCM and washed with 0.1 N HCl for 3 times. Organic mass was further washed with water and brine, dried on magnesium sulphate and purified by silica gel column chromatography using DCM and Methanol. Yield 60% .sup.1H NMR (CD.sub.3OD, 500 MHz) 0.85-0.89 (t, 6H), 1.26-1.27 (m, 12H), 1.37*1.49 (m, 6H), 1.70-1.74 (m, 1H), 1.79-1.85 (m, 2H), 1.96 (s, 7H), 1.99 (s, 2H), 2.11 (s, 6H), 2.16 (s, 2H), 3.81-3.85 (m, 1H), 4.15-4.19 (m, 2H), 6.29-6.32 (d, 1H, J=13.68 Hz), 6.56-6.63 (t, 1H, J=12.55 Hz), 7.05-7.08 (t, 2H, J=7.40 Hz), 7.26-7.30 (t, 5H, J=8.53), 7.43-7.47 (t, 1H, J=7.91 Hz), 7.52-7.54 (d, 5H, J=9.79 Hz), 7.59-7.63 (t, 1H, J=6.90 Hz), 7.81-7.86 (m, 1H), 7.95-8.04 (m, 2H), 8.19-8.21 (d, 1H, J=8.53 Hz). BRMS (ES+), m/z [M+H] 705.5210.

Synthetic Routes for Other Cyanine 7.5 Derivatives.

[0187] Substituted cydic analogues of cyanine 7.5 dye (scheme 1b of FIG. 8) were synthesized in five steps. Indolinium salts with C8 or C18 alkyl chains were synthesized by the condensation of trimethyl benzindole with alkyl iodide. Formylation of cyclohexanone followed by condensation with aniline in the presence of HCl resulted in the cyclic inner salt (b). Condensation of inner salt (b) and indolinium salt in the presence of sodium acetate resulted in a key intermediate cyclic chloro-cyanine. Substitution of -chloro with different phenol derivatives followed by the counter ion exchange with tetraphenyl borate resulted in a series of cyanine derivatives.

[0188] Cyclic cyanine dye having methyl group on the central cyclic ring (HA-04-I) was prepared by following the different route (scheme 1c of FIG. 8). Inner salt was prepared from 1-methylcyclohexene in the presence of N-methyl-N-phenyl formamide and POCl.sub.3. Condensation of inner salt with indolinium salt in the presence of pyridine resulted in the final compound.

Synthesis of Inner Salt (b):

[0189] ##STR00033##

[0190] This compound was prepared by following the literature method (Bioconjugate Chem. 1997, 8, 751-756; Chem. Commun., 2014, 50, 15439-15442). Yield 57%

[0191] .sup.1H NMR (DMSO d.sub.6, 400 MHz) 1.84 (s, 2H), 2.74 (s, 4H), 7.20-7.26 (m, 2H), 7.41-7.46 (m, 6H), 7.55-7.57 (d, 4H, 3=7.91 Hz), 7.70-7.72 (d, 1H, J=7.71 Hz), 8.68 (s, 2H).

[0192] As what is shown in schema 1b of FIG. 8, chloro-cyanine (10 mg, 0.0084 mmol, 1 equiv.) was dissolved in dry DMF (2 ml) at 25 C. In a separate round bottom flask, 2,6-dipehenylphenol (10.34 mg, 0.0420 mmol, 5 equiv.) was dissolved in dry DMF (2 ml) and dry K.sub.2CO.sub.3 (2.32 mg, 0.0168 mmol 2 equiv.) was added to it and the solution was stirred for 10-15 minutes. It was carefully filtered to get rid of the unreacted K.sub.2CO.sub.3 and the filtrate was slowly added to the solution of chloro-cyanine. The reaction mass was stirred at 25 C. for 30 minutes. The reaction was monitored by UV-Vis spectroscopy characterized by the disappearance of the absorption band at around 820 nm (corresponds to chloro-cyanine) and the formation of a new absorption band at shorter wavelength region. Upon completion, the reaction was quenched with solid CO.sub.2 and concentrated under vacuum to remove the DMF. The reaction mass was redissolved in dichloromethane and purified by silica gel column chromatography using dichloromethane/methanol as eluents.

[0193] Characterisation of Substituted Cyanine 7.5 Dye

[0194] Six derivatives of cyanine 7.5 dye of formula (I) of the present invention: HA-06, HA-04, HA-05, HA-60, HA-63, HA-69, as iodide or tetraphenylborate (TB) salts, were synthesised according to the protocol described above.

HA-06-I:

[0195] ##STR00034##

[0196] The title compound (68%, green solid) was prepared from 2,6-diphenylphenol according to the general protocol S1. .sup.1H NMR (CD.sub.3OD, 400 MHz) 0.84-0.87 (t, 6H), 1.24 (s, 59H), 1.39-1.47 (m, 9H), 1.71 (s, 10H), 1.84-1.88 (t, 3H), 2.03 (s, 2H), 2.15-2.18 (t, 4H), 4.16-4.20 (t, 4H), 4.60 (s, 1H), 5.96-5.99 (d, 2H J=14.05 Hz), 7.30-7.34 (t, 1H, J=7.40 Hz), 7.37-7.47 (m, 9H), 7.49-7.64 (m, 8H), 7.95-7.99 (t, 4H, J=8.53 Hz), 8.08 (s, 1H), 8.12-8.15 (t, 3H, J=3H) BRMS (ES+), m/z [M+H] 1270.9522.

HA-04-I:

[0197] ##STR00035##

[0198] .sup.1H NMR (CD.sub.3OD, 400 MHz) 0.85-0.88 (t, 6H), 1.26 (s, 51H), 1.44-1.47 (m, 4H), 1.50-1.53 (m, 3H), 1.89-1.93 (m, 4H), 1.96-1.98 (m, 2H), 2.04 (s, 12H), 2.56 (s, 3H), 2.64-2.67 (t, 4H), 4.25-4.28 (t, 4H), 4.61 (s, 1H), 6.28-6.30 (2H, d, J=13.73), 7.47-7.50 (t, 2H, J=7.17 Hz) 7.58-7.60 (2H, J=8.85 Hz), 7.63-7.66 (2H, J=7.17 Hz), 7.99-8.03 (q, 4H), 8.25-8.27 (d, 2H, J=8.54 Hz), 8.29-8.31 (d, 2H, J=13.29 Hz). BRMS (ES+), m/z [M+H] 1039.8734.

HA-05-I:

[0199] ##STR00036##

[0200] The title compound (65%) was prepared from 3,5-dimethylphenol according to the general protocol S1. .sup.1H NMR (CD.sub.3OD, 400 MHz) 0.84-0.88 (t, 6H), 1.25 (s, 55H), 1.44 (br, 12H), 1.84-1.87 (t, 6H), 2.03 (s, 3H), 2.09-2.12 (t, 2H), 2.17 (s, 3H), 2.67 (s, 2H), 2.79 (br, 4H), 4.20-4.24 (t, 4H), 4.60 (s, 1H), 6.18-6.21 (d, 2H, J=14.43 Hz), 6.55 (s, 1H), 6.80-6.82 (d, 1H, J=7.65 Hz), 7.43-7.47 (t, 2H, J=7.15 Hz), 7.53-7.55 (d, 2H, J=8.78 Hz), 7.57-7.66 (m, 3H), 7.94-7.96 (2H, d, J=17.57 Hz), 7.98-8.04 (m, 3H), 8.08-8.10 (2H, J=8.53 Hz). BRMS (ES+), m/z [M+H] 1145.9127.

HA-60-I:

[0201] ##STR00037##

[0202] Yield (68%) This compound was prepared from methyl-4-hydroxybenzoate and chloro-cyanine C8 according to the general protocol S1. BRMS (ES+), m/z [M+H] 895.5898.

HA-63-I:

[0203] ##STR00038##

[0204] Yield (50%). This compound was prepared from methyl-4-hydroxybenzoate and chloro-cyanine C18 according to the general protocol S1. BRMS (ES+), m/z [M+H] 1175.8981.

HA-69-I:

[0205] ##STR00039##

[0206] The title compound (63%, green solid) was prepared from 3,5-dimethylphenol and chloro-cyanine C8 according to the general protocol S1. .sup.1H NMR (CDCl.sub.3, 400 MHz) 0.87 (s, 6H), 1.16-1.42 (m, 26H), 1.68 (s, 7H), 1.86 (s, 4H), 2.00 (s, 4H), 2.16 (s, 3H), 2.76-2.86 (d, 2H), 4.17-4.29 (d, 4H), 5.49 (s, 3H), 5.94-5.98 (d, 1H, J=13.68 Hz), 6.30-6.33 (1H, d, J=14.18 Hz), 7.30-7.62 (m, 15H), 7.94-8.14 (m, 7H), 8.24-826 (1H, d, J=8.51 Hz), 8.51-8.55 (1H, d, J=14.10 Hz). BRMS (ES+), m/z [M+H] 989.6366.

[0207] Tetraphenylborate salts of Cv7.5 derivatives were prepared by the similar protocol as for Cy7.5-C18-TPB.

[0208] 1.2 Chemical Composition of the Coating Solution

[0209] Commercially available poly(methylmethacrylate), Cy7.5-C18-TPB synthesized before, tetraphenylborate and acetonitrile are used for preparing a tested coating solution.

[0210] Polymethylmethacrylate is dissolved at 30 mg/ml in acetonitrile. Cy7.5-C18-TPB dye with tetraphenylborate counterion is added at 1 wt % with respect to the polymer.

[0211] A comparative coating solution is prepared according to the same protocol. The comparative coating solution is the same as tested coating solution, except that a ICG derivative (ICG, ICG-N, or ICG-P) is used as fluorescent dye in place of Cy7.5-C18-TPB.

[0212] ICG-N is an ICG salt with tetrabutyl ammonium as counterion. ICG-P is an ICG salt with tetraphenyl phosphonium as counterion.

[0213] These two reference dyes are used to show that counterions in combination with ICG cannot produce the same performance as the specially designed hydrophobic dyes and counterions.

[0214] 1.3 Protocol of Coating

[0215] The coating solution prepared in section 1.2 is placed in the eppendorf tube. The medical device to be coated is immersed into this solution and then immediately taken out by tweezers. It is placed on the aluminium foil to dry for 15-20 min in the dark. Then, the procedure can be repeated 5-10 times in order to increase the thickness of the polymer layer and thus the brightness of the fiducial. After the coating, fiducial is left for 24 h in the dark for complete drying of remaining acetonitrile. At this step the fiducial is ready to use.

[0216] A stainless steel wire, silicon pieces, a surgical thread, a catheter, a nano-gastric tube, surgical markers and medical gauze are coated by the tested coating solution according to this protocol.

[0217] A stainless steel wire, silicon pieces, a surgical thread are also coated by the comparative coating solutions.

2. Results

[0218] Brightness

[0219] Compared to ICG-based polymeric coating films, the coating film of the present invention is adhesive to smooth surface, such as on stainless steel wires.

[0220] Indeed, the NIR fluorescence of costing film based on Cy7.5-C18-TPB on a stainless steel wire became visible after the first coating and increases by multiple coating, while ICG-based coating films could generate fluorescent labelling of the wires at all even after 5 time coating procedure (FIG. 1).

[0221] Stability

[0222] The coating film of the present invention is also more stable than ICG-based coating film.

[0223] The FIG. 2 shows that fluorescence observed for the coating film of the invention is more homogeneous and can remain stable 90 seconds after the coating procedure, while the fluorescence observed for ICG-based coating films is not homogeneous and is quickly lost.

[0224] The FIG. 3 shows that the fluorescence brightness observed for the coating film of the invention on surgical thread is preserved a week after the coating procedure, while the brightness of ICG-based coating films endures significant decrease during 1 week.

[0225] It is interesting to note that the presence of a counterion in ICG derivative dyes (ICG-P or ICG-N) does not have significant impact to the brightness and stability of ICG-based polymeric coating films.

[0226] In Vivo Application of Coated Surgical Devices

[0227] The FIG. 4 shows that the ureter, which is an anatomical structure vulnerable to injury during lower abdominal operation, can be easily identified by the insertion of a catheter coated by a Cy7.5-C18-TPB-based coating film via a fluorescent imaging system.

[0228] The FIG. 5 shows that a tip of naso-gastric tube, which is sometimes difficult to be identified during surgery, can be easily located after being coated by a Cy7.5TPB-based coating film.

[0229] The FIG. 6 shows that surgical markers coated by a Cy7.5-C18-TPB-based coating film can be visualized from outside of the intestinal organ, which can help the surgeon to find the location of the tumor and be useful for registration of digital augmented reality technology.

[0230] The FIG. 7 shows that the coating film of the present invention can also be used for coating a woven fabric.

[0231] Coating of Glass Surface by Films of the Invention

[0232] Coating films of the invention containing PMMA and respectively CY7.5-C18-TPB, CY7.5-C8-TPB, HA-60-TPB, or HA-06-I as fluorescent dye and counterion, and comparative coating films containing PMMA with ICG or PMMA only, were prepared by spin coating a mixture of PMMA (30 mg/ml) with different dyes (1% loading) on a 25 mm glass substrate. Images were taken by using a home built NIR imaging setup.

[0233] The results illustrated in FIG. 9 show that the coating films of the present invention are brighter than the coating film containing ICG.

[0234] Coating films made from different polymers were also tested. For all studied polymers, coating films based on fluorescent dyes Cy7.5-C18-TPB and the derivatives HA-06-I, HA-60-TPB systematically display better brightness than a coating film based on ICG (FIG. 10A, 10B).

[0235] Moreover, the NIR fluorescence detected from steel fiducials coated with PMMA films based on HA-06-I or HA-60-TPB inserted into muscle tissue is comparable with that from coating PMMA film based on CY7.5-C18-TPB (FIG. 11).