Near-infrared fluorescent probe specifically targeting tumors as well as synthesis method and use thereof

Abstract

The present disclosure provides a near-infrared fluorescent probe specifically targeting tumors as well as a synthesis method and use thereof. The near-infrared fluorescent probe specifically targeting tumors is a compound represented by formula I or a pharmaceutically available salt thereof: ##STR00001## wherein, X is a linker molecule selected from PEG.sub.n and glycine (G.sub.m), n=0-10, m=0-10, one end of the linker molecule is amino, and the other end of the linker molecule is carboxyl; Y is a dye molecule having a fluorescence excitation and emission spectrum within a near-infrared (NIR) range, and the compound represented by formula I or pharmaceutically available salt thereof can maintain or enhance the fluorescence of the dye molecule Y. The near-infrared fluorescent probe of the present disclosure can be rapidly cleared away in normal tissues, and remains for a long time in tumor sites, thereby taking the effect of living diagnosis. Hence, the near-infrared fluorescent probe of the present disclosure has a certain clinical application prospect, and can be applied to clinical surgery navigation.

Claims

1. A method for synthesizing a near-infrared fluorescent probe specifically targeting tumors, wherein the near-infrared fluorescent probe is a compound represented by formula I or a pharmaceutically available salt thereof: ##STR00012## wherein: X is a linker molecule selected from PEG.sub.4, PEG.sub.6, G.sub.3 and G.sub.6, have the following structures respectively: ##STR00013## one end of the linker molecule is amino, and the other end of the linker molecule is carboxyl; Y is a dye molecule having a fluorescence excitation and emission spectrum within a near-infrared (NIR) range, and the compound represented by formula I or pharmaceutically available salt thereof can maintain or enhance the fluorescence of the dye molecule Y; the method comprises the following steps: Step a, mixing lapatinib and X in the presence of 2-(7-azabenzotriazole)-N,N,N,N-tetramethylurea hexafluorophosphate, an alkaline and a polar solvent; Step b, dropwise adding the product obtained in Step a into water, extracting and then concentrating, subsequently adding trifluroracetic acid to remove a BOC protective group, and concentrating to obtain a lapatinib-X intermediate compound; Step c, mixing the lapatinib-X intermediate compound with a dye molecule Y in the presence of 2-(7-azabenzotriazole)-N,N,N,N-tetramethylurea hexafluorophosphate, an alkaline and a polar solvent; and Step d, purifying the product obtained in Step c with a preparative liquid phase to obtain a target compound lapatinib-X-Y, the near-infrared fluorescent probe.

2. The method for synthesizing the near-infrared fluorescent probe specifically targeting tumors according to claim 1, wherein in the Step a and Step c, the polar solvent is one or more of N,N-dimethylformamide, anhydrous dimethylsulfoxide and N-methylpyrrolidone.

3. The method for synthesizing the near-infrared fluorescent probe specifically targeting tumors according to claim 1, wherein in the Step a and Step c, the alkaline is one or more of triethylamine and N,N-diisopropylethylamine (DIEA).

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) FIG. 1 is an absorption spectrum of YQ-H-01 and others prepared in embodiments;

(2) FIG. 2 is an fluorescence spectrum of YQ-H-01 and others prepared in embodiments;

(3) FIG. 3 shows in-vivo imaging of YQ-H-01 and others prepared in embodiments in colorectal cancer HT29 tumor-bearing mice;

(4) FIG. 4 shows in-vivo imaging of a YQ-H-01 probe prepared in an embodiment in pancreatic cancer PANC1, liver cancer HepG2 and lung cancer H460 tumor-bearing mice.

DETAILED DESCRIPTION OF THE EMBODIMENTS

(5) Next, the present disclosure will be further explained in combination with embodiments.

(6) The present disclosure will be better understood according to embodiments below. However, those skilled in the art will easily understand that specific material proportions, process conditions and results described in embodiments are only for illustrating the present disclosure, but should not or do not limit the present disclosure described in claims.

Example 1: Synthesis of YQ-H-01

(7) ##STR00005##

(8) Lapatinib (5 mg, 1.0 eq), ICG-02 (12 mg, 1.5 eq), 2-(7-azabenzotriazole)-N,N,N,N-tetramethylurea hexafluorophosphate (HATU, 13 mg, 4.0 eq) and triethylamine (6 l, 5.0 eq) were reacted for 3 h at room temperature in the dark, the reaction was monitored by HPLC, the obtained product was purified by a preparative liquid phase after the reaction was completed, and a target fraction was lyophilized to obtain green solid YQ-H-01 (5 mg, Y=38.9%). By mass spectrum and nuclear magnetic hydrogen spectrum, its structure is verified as follows: m/z=, .sup.1H NMR (300 MHz, DMSO) 8.88 (dd, J=15.1, 10.5 Hz, 2H), 8.66 (t, J=12.4 Hz, 2H), 8.24 (dd, J=11.1, 6.4 Hz, 1H), 8.01 (t, J=2.8 Hz, 1H), 7.85-7.66 (m, 4H), 7.60-7.45 (m, 3H), 7.42-7.16 (m, 7H), 6.52 (dt, J=24.9, 7.5 Hz, 3H), 5.34 (s, 2H), 4.64 (s, 2H), 4.26 (d, J=7.2 Hz, 4H), 3.39 (t, J=8.0 Hz, 2H), 3.17 (s, 2H), 3.04 (d, J=9.4 Hz, 4H), 2.73-2.59 (m, 8H), 2.55 (s, 3H), 2.07-1.88 (m, 4H), 1.78-1.65 (m, 2H), 1.58 (d, J=7.6 Hz, 12H).

Example 2: Synthesis of YQ-H-03

(9) ##STR00006##

(10) Lapatinib (10 mg, 1.0 eq), 6-Boc-aminocaproic acid (8 mg, 2 eq), 2-(7-azabenzotriazole)-N,N,N,N-tetramethylurea hexafluorophosphate (HATU, 13 mg, 2.0 eq) and triethylamine (7.2 l, 3.0 eq) were reacted for 1 h at room temperature, the reaction was monitored by TLC, the reaction solution was dropwise added into water after the reaction was completed and then extracted with ethyl acetate, organic layers were combined and concentrated, then trifluoroacetic acid was added to remove Boc, the reaction was also monitored by TLC, trifluoroacetic acid was evaporated after the reaction was completed, the obtained product was purified to obtain lapatinib linking to 6-aminocaproic acid, and the rest steps referred to synthesis of compound YQ-H-01, so as to obtain a target compound YQ-H-03, a green solid. By mass spectrum and nuclear magnetic hydrogen spectrum, its structure is verified as follows: m/z=, .sup.1H NMR (300 MHz, DMSO) 8.88 (dd, J=15.1, 10.5 Hz, 2H), 8.66 (t, J=12.4 Hz, 2H), 8.24 (dd, J=11.1, 6.4 Hz, 1H), 8.01 (t, J=2.8 Hz, 1H), 7.85-7.66 (m, 4H), 7.60-7.45 (m, 3H), 7.42-7.16 (m, 7H), 6.52 (dt, J=24.9, 7.5 Hz, 3H), 5.34 (s, 2H), 4.64 (s, 2H), 4.26 (d, J=7.2 Hz, 4H), 3.39 (t, J=8.0 Hz, 2H), 3.17 (s, 2H), 3.04 (d, J=9.4 Hz, 4H), 2.73-2.59 (m, 8H), 2.55 (s, 3H), 2.07-1.88 (m, 4H), 1.78-1.65 (m, 2H), 1.58 (d, J=7.6 Hz, 12H).

Example 3: Synthesis of YQ-H-04

(11) ##STR00007##

(12) The synthesis method refers to synthesis of YQ-H-03. By mass spectrum and nuclear magnetic hydrogen spectrum, its structure is verified as follows: m/z=, .sup.1H NMR (300 MHz, DMSO) 8.96 (dd, J=8.0, 4.8 Hz, 2H), 8.66 (d, J=14.0 Hz, 2H), 8.36 (d, J=8.9 Hz, 1H), 8.01-7.88 (m, 3H), 7.75 (d, J=1.2 Hz, 2H), 7.65 (ddd, J=9.6, 8.6, 1.9 Hz, 3H), 7.53-7.46 (m, 1H), 7.45-7.30 (m, 5H), 7.25-7.14 (m, 2H), 6.64-6.43 (m, 3H), 5.32 (s, 2H), 4.72 (d, J=23.4 Hz, 2H), 4.40-4.23 (m, 4H), 3.61-3.53 (m, 2H), 3.51-3.41 (m, 12H), 3.41-3.30 (m, 4H), 3.19-3.12 (m, 2H), 3.03 (d, J=4.6 Hz, 2H), 2.97 (td, J=6.6, 3.9 Hz, 2H), 2.76 (dd, J=11.9, 5.0 Hz, 2H), 2.73-2.59 (m, 8H), 2.55 (s, 3H), 2.42-2.31 (m, 2H), 2.10-1.91 (m, 4H), 1.80-1.71 (m, 2H), 1.65 (d, J=2.0 Hz, 12H).

Example 4: Synthesis of YQ-H-05

(13) ##STR00008##

(14) The synthesis method refers to synthesis of YQ-H-03. By mass spectrum and nuclear magnetic hydrogen spectrum, its structure is verified as follows: m/z=, .sup.1H NMR (300 MHz, DMSO) 8.95 (t, J=2.6 Hz, 2H), 8.67 (d, J=14.1 Hz, 2H), 8.37 (dd, J=12.4, 3.7 Hz, 1H), 7.99 (dd, J=7.1, 3.6 Hz, 1H), 7.96-7.88 (m, 2H), 7.75 (d, J=1.0 Hz, 2H), 7.70-7.59 (m, 3H), 7.54-7.30 (m, 6H), 7.25-7.14 (m, 2H), 6.66-6.42 (m, 3H), 5.32 (s, 2H), 4.73 (d, J=23.7 Hz, 2H), 4.33 (s, 4H), 3.59-3.54 (m, 2H), 3.51-3.41 (m, 20H), 3.36 (dd, J=11.4, 5.8 Hz, 4H), 3.19-3.11 (m, 2H), 3.03 (s, 2H), 2.98 (d, J=6.8 Hz, 2H), 2.77 (dd, J=11.5, 5.5 Hz, 2H), 2.61 (dd, J=18.9, 12.8 Hz, 8H), 2.52 (d, J=1.6 Hz, 3H), 2.36 (t, J=6.9 Hz, 2H), 2.01 (dd, J=13.8, 7.4 Hz, 4H), 1.76 (d, J=3.2 Hz, 2H), 1.68 (d, J=10.9 Hz, 12H).

Example 5: Synthesis of YQ-H-06

(15) ##STR00009##

(16) The synthesis method refers to synthesis of YQ-H-03. By mass spectrum and nuclear magnetic hydrogen spectrum, its structure is verified as follows: m/z=, .sup.1H NMR (300 MHz, DMSO) 8.95 (s, 2H), 8.68 (dd, J=13.9, 6.0 Hz, 2H), 8.49-8.38 (m, 1H), 8.32-8.11 (m, 4H), 7.90 (dd, J=7.0, 2.9 Hz, 2H), 7.76 (s, 2H), 7.65 (dd, J=12.0, 5.6 Hz, 3H), 7.52-7.44 (m, 1H), 7.44-7.30 (m, 5H), 7.21 (dd, J=11.5, 5.7 Hz, 2H), 6.71-6.42 (m, 3H), 5.32 (s, 2H), 4.78-4.63 (m, 2H), 4.32 m, 4H), 4.16 (m, 2H), 3.48-3.25 (m, 6H), 3.17 (s, 2H), 3.09-2.99 (m, 4H), 2.76-2.56 (m, 8H), 2.52 (s, 3H), 2.45 (m, 2H), 2.02 (m, 4H), 1.76 (m, 2H), 1.72-1.55 (m, 12H).

Example 6: Synthesis of YQ-H-07

(17) ##STR00010##

(18) The synthesis method refers to synthesis of YQ-H-03. By mass spectrum and nuclear magnetic hydrogen spectrum, its structure is verified as follows: m/z=, .sup.1H NMR (300 MHz, DMSO) 8.94 (s, 2H), 8.67 (d, J=14.0 Hz, 2H), 8.51-7.99 (m, 8H), 7.90 (dd, J=5.4, 2.8 Hz, 2H), 7.77 (s, 2H), 7.63 (d, J=8.4 Hz, 3H), 7.53-7.29 (m, 6H), 7.25-7.15 (m, 2H), 6.72-6.41 (m, 3H), 5.32 (s, 2H), 4.80-4.65 (m, 2H), 4.38-4.20 (m, 6H), 3.71 (d, J=4.9 Hz, 12H), 3.35 (p, J=7.9 Hz, 2H), 3.03 (s, 2H), 3.00-2.92 (m, 2H), 2.65 (dd, J=13.6, 8.1 Hz, 8H), 2.55 (s, 3H), 2.43 (dd, J=8.7, 2.8 Hz, 2H), 2.11-1.94 (m, 4H), 1.84-1.73 (m, 2H), 1.66 (s, 12H).

Example 7: Synthesis of YQ-H-08

(19) ##STR00011##

(20) The synthesis method refers to synthesis of YQ-H-03. By mass spectrum and nuclear magnetic hydrogen spectrum, its structure is verified as follows: m/z=, .sup.1H NMR (300 MHz, DMSO) 8.94 (d, J=3.5 Hz, 2H), 8.42 (dd, J=28.0, 9.0 Hz, 1H), 8.22-8.02 (m, 6H), 7.94-7.86 (m, 2H), 7.76 (dd, J=13.8, 7.2 Hz, 2H), 7.68-7.56 (m, 5H), 7.49 (td, J=8.0, 6.1 Hz, 1H), 7.37 (dd, J=12.7, 9.3 Hz, 5H), 7.20 (dd, J=9.0, 6.1 Hz, 4H), 7.01 (t, J=7.3 Hz, 2H), 6.63 (dd, J=43.4, 3.1 Hz, 1H), 6.34 (d, J=14.4 Hz, 2H), 5.32 (s, 2H), 4.73 (d, J=15.3 Hz, 2H), 4.21 (d, J=33.3 Hz, 6H), 3.78-3.64 (m, 12H), 3.36 (dd, J=12.9, 5.9 Hz, 2H), 3.03 (s, 2H), 2.83-2.64 (m, 6H), 2.59 (t, J=6.5 Hz, 4H), 2.55 (s, 3H), 2.35 (dd, J=13.1, 5.9 Hz, 2H), 2.05-1.91 (m, 4H), 1.91-1.80 (m, 2H), 1.22 (d, J=4.2 Hz, 12H).

(21) The above descriptions are only preferred embodiments of the present disclosure. It should be noted that for persons of ordinary skill in the art, several improvements and modifications can also be made without departing from the principle of the present disclosure, and these improvements and modifications should be deemed as the protective scope of the present disclosure.

Example 8

(22) In a subcutaneous tumor HT29 tumor-bearing mouse model, YQ-H (30 nmol, dissolved with 100 L of normal saline for injection) series probes respectively underwent tail vein administration, fluorescence imaging was respectively photographed at different time points (0 h, 6 h, 12 h, 24 h and 48 h in sequence) by small animal living imaging CCD. Results show that in HT29 tumor-bearing mice, probes YQ-H-01/03 have a certain tumor targeting effect while having strong fluorescence signals in liver and the like, which is possibly disadvantageous for development of subsequent probes; probe YQ-H-08 introduces an aroma ring structure between target molecule lapatinib and fluorescent dye, which enhances the intake of tumors to a certain extent, and meanwhile leads to obvious whole body signal of mice probably caused by a fact that fat solubility is enhanced to bring about prolonged in-vivo metabolism time; probe YQ-H-04/06/07 exhibit better in-vivo metabolism features, fluorescence signals in liver are significantly reduced, and fluorescence signals in tumor sites are obvious, wherein the tumor effect of the probe YQ-H-06 is more excellent and has follow-up development potential.

(23) On the basis of this, through tumor targeting validation of pancreatic cancer (PANC1), liver cancer (HepG2) and lung cancer (H460) tumor-bearing mice on the probe YQ-H-06, it is found that the probe YQ-H-06 has a good tumor targeting effect on three subcutaneous tumor bearing mice, and needs further research and development to be applied to clinical surgery navigation.