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Synthesis 2015; 47(23): 3717-3726
DOI: 10.1055/s-0034-1378876
paper
© Georg Thieme Verlag Stuttgart · New York

Synthesis of Chlorin–(Arylamino)quinazoline Hybrids as Models for Multifunctional Drug Development

Alexander V. Nyuchev
a   Department of Chemistry, N.I. Lobachevsky State University of Nizhny Novgorod, 23 Gagarin Avenue, 603950 Nizhny Novgorod, Russian Federation   Email: afnn@rambler.ru
,
Vasiliy F. Otvagin
a   Department of Chemistry, N.I. Lobachevsky State University of Nizhny Novgorod, 23 Gagarin Avenue, 603950 Nizhny Novgorod, Russian Federation   Email: afnn@rambler.ru
,
Andrei E. Gavryushin
b   Nanoscape AG, Am Klopferspitz 19, 82152 Planegg, Germany
,
Yuliya I. Romanenko
c   Research Institute of Macroheterocycles, Ivanovo State University of Chemical Technology, 153000 Ivanovo, Russian Federation
,
Oscar I. Koifman
c   Research Institute of Macroheterocycles, Ivanovo State University of Chemical Technology, 153000 Ivanovo, Russian Federation
,
Dmitrii V. Belykh
d   Institute of Chemistry, Komi Scientific Center, Urals Branch of the Russian Academy of Sciences, 167982 Syktyvkar, Russian Federation
,
Hans-Günther Schmalz
e   Department of Chemistry, University of Cologne, Greinstrasse 4, 50939 Cologne, Germany
,
Alexey Yu. Fedorov*
a   Department of Chemistry, N.I. Lobachevsky State University of Nizhny Novgorod, 23 Gagarin Avenue, 603950 Nizhny Novgorod, Russian Federation   Email: afnn@rambler.ru
› Author Affiliations
Further Information

Publication History

Received: 14 May 2015

Accepted after revision: 07 July 2015

Publication Date:
20 August 2015 (online)

 
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Abstract

A series of multifunctional conjugates each consisting of a fluorescent chlorin photosensitizer and an (arylamino)quinazoline-based epidermal growth factor receptor/vascular endothelial growth factor receptor ligand, potentially useful in site-selective photodynamic antitumor therapy, were prepared and their photochemical properties were investigated.


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Key words

porphyrins - cycloaddition - alkynes - azides - click chemistry - medicinal chemistry

Photodynamic therapy (PDT) is an effective method for the treatment of a number of oncological[1] and dermatological diseases.[1f] [2] It can also be used in antifungal[3] and antimicrobial therapies,[4] and it has other important biomedical applications.[1f] [5] A major problem in anticancer photodynamic therapy is a low selectivity of accumulation of the PDT agent in tumor tissues.[6] The accumulation of a photosensitizer, even in trace amounts, in skin or in mucous membranes can lead to serious phototoxicity.[7] One way to improve the situation is to construct hybrid therapeutic molecules bearing a photosensitizer conjugated to a molecular fragment possessing a high affinity to receptors expressed preferentially in tumor cells.[8] Such hybrid conjugates can be prepared in accordance with the concept of multivalent interactions.[9] [10] The aim of this work was to develop model hybrid conjugates that (a) would act both as PDT-photosensitizers and as cytotoxic agents; (b) would selectively accumulate in tumor cells; and (c) would permit fluorescence imaging of their tissue distribution. Such multifunctional agents for the simultaneous imaging and targeted (noninvasive) treatment of cancers have attracted serious interest in recent decades.[1f] [11]

Chlorin e 6 and related compounds, which are natural metabolites of chlorophyll a,[7] [12] are widely used as efficient photosensitizers in photodynamic therapy (Figure [1]).[7,12a,13] They display absorption maxima and significant fluorescence in the red part of the UV–visible spectrum[7] [14] (650–700 nm), and they can be used in fluorescent imaging techniques or as agents for therapy guidance, diagnostics, treatment assessment, or mechanistic studies.[11] Several chlorin-based antitumor drugs have been approved for clinical use in the last two decades, the most successful of which is the fully synthetic temoporfin[9] [12a] [15] and the semisynthetic NPe 6.[16]

For the PDT-active part of our new series of conjugates, we selected a chlorin e 6 fragment and, more specifically, its zinc complex. The zinc complex was selected as a means of increasing the pharmaceutical potential of the final conjugates, because the insertion of a diamagnetic metal into the core of a dihydroporphyrin often improves its photochemical properties, particularly its singlet-oxygen (Φδ) and triplet­-state (Φ t ) yields.[7] [17]

For the part of the conjugate responsible for selective delivery to tumor tissues, we chose 4-(arylamino)quinazolines (Figure [1]). These agents are known to be selective epidermal growth factor receptor (EFGR)/vascular endothelial growth factor receptor (VEGFR) ligands.[18] One of the most promising drugs of this family, Vandetanib[19] (Figure [1]), was approved by the US Federal Drug Administration in 2011 for the treatment of late-stage (metastatic) medullary thyroid cancer.

Zoom Image
Figure 1 Chlorin-type photosensitizers and epidermal growth factor receptor (EFGR)/vascular endothelial growth factor receptor (VEGFR) ligands

Thus, a 4-(arylamino)quinazoline fragment of the conjugates was intended to act as a selective anchor for the growth factor receptors of the tumor cells, as well as behaving as a tyrosine kinase inhibitor[20] that might significantly improve the antitumor effect of the PDT-conjugate (Scheme [1]).

Zoom Image
Scheme 1 Functionality-based design, synthesis strategy, and expected properties of hybrid molecules for photodynamic therapy

The chlorin e 6 derivatives and the 4-(arylamino)quinazoline EGFR/VEGFR ligands were assembled into conjugates by means of copper-mediated dipolar cycloaddition (click chemistry) reactions.[21] (Scheme [1]). Depending on the type of linker used to connect the PDT part and the targeting part, the resulting conjugate could be designed to contain one or several chlorin and/or quinazoline fragments.

The 4-(arylamino)quinazoline 1 was synthesized in seven steps and 45% overall yield from vanillic acid according to the reported procedure.[22] It was further transformed into the azide-containing derivative 3 by using 6-azidohexanoic acid (2)[23] in a carbodiimide-mediated acylation[24] (Scheme [2]).

Zoom Image
Scheme 2 Synthesis of the EGFR/VEGFR ligand core (1) and its azide-functionalized derivative 3

The PDT-active chlorin e 6 moieties (Scheme [3,] Table [1]) were prepared from naturally occurring methylpheophorbide a (4), isolated from spinach or spirulina.[25]

The five-membered exocycle in methylpheophorbide a (4) underwent ring opening on treatment with an excess of a polyamine 5a–c. The resulting products were subjected to insertion of a zinc cation in situ to give the corresponding chlorins 6a–c in excellent yields.

Zoom Image
Scheme 3 Synthesis of alkyne-functionalized chlorin e 6 derivatives

In the next step, the free amino groups in chlorins 6a–c were acylated with 3,5-bis(propargyloxy)benzoic acid[26] (7a) or pent-4-ynoic acid (7b) under Steglich conditions (Table [1]) to give the terminal alkyne group-bearing chlorins 8a–d.

Table 1 Synthesis of Alkyne-Bearing Chlorin e 6 Derivatives

Entry

Amine

Product

Yield (%)

Acylation conditions

Acylation product

Yield (%)

1

5a

6a

92

7a (1.9 equiv), EDC·HCl (2 equiv), HOBt (2 equiv), DMAP (0.5 equiv), CH2Cl2, argon, r.t.

8a

93

2

5b

6b

99

7a (3 equiv), EDC·HCl (3 equiv); HOBt (3 equiv); DMAP (1 equiv), CH2Cl2, argon, r.t.

8b

88

3

5c

6c

99

7b (2.1 equiv); EDC·HCl (2.1 equiv); DMAP (1.5 equiv), CH2Cl2, argon, r.t.

8c

90

4

7a (2.1 equiv); EDC·HCl (2.1 equiv); HOBt (2.1 equiv); DMAP (1.5 equiv), CH2Cl2, argon, r.t.

8d

90

The azide-functionalized (arylamino)quinazoline EGFR/ VEGFR ligand 3 was linked to the alkyne-functionalized chlorins 8a–d and 10 [27] by means of a click [3+2] Huisgen cycloaddition[21] with copper iodide as a catalyst (Scheme [4] and Scheme [5]) to afford the 1,4-regioisomeric triazoles 9a–d and 11, respectively, as sole products in 40–99% yield. This method was used to prepare a number of conjugates, containing one chlorin fragment and one to four 4-(arylamino)quinazoline groups.

Zoom Image
Scheme 4 Synthesis of conjugates 9a–c
Zoom Image
Scheme 5 Synthesis of conjugates 9d and 11

The bis(chlorin)–quinazoline conjugate 15 was prepared from the amine-bearing chlorin 6a and quinazolinol 1 in three steps (Scheme [6]). Compound 6a was acylated with 6-azidohexanoic acid (2) under Steglich conditions to afford the azide-functionalized chlorin 12 in quantitative yield. The reaction of quinazoline derivative 14, obtained by alkylation of the phenol function of quinazolinol 1 with 1-(bromomethyl)-3,5-bis(prop-2-yn-1-yloxy)benzene[28] (13), with chlorin 12 in the presence of copper(I) iodide catalyst in aqueous N,N-dimethylformamide gave the desired conjugate 15 in 57% isolated yield.

Zoom Image
Scheme 6 Synthesis of conjugate 15

Conjugates 9a-d, 11, and 15 were shown to display red fluorescence regardless of the number of the chlorin and quinazoline moieties present or the nature of the linker (Table [2] and Figures S24–S29 in the Supporting Information).

Table 2 Photophysical Properties of Conjugates 9a, 11, and 15

Conjugate

λex

λem

9a

420

650

11

330

649

15

335

647

In conclusion, we have synthesized a range of new conjugates consisting of one or more PDT-active chlorin-based photosensitizers and one or more EGFR/VEGFR ligands (targeted drug and tyrosine kinase inhibitor). These hybrid molecules are of serious interest as models for a new class of therapeutic agents that would permit multitargeted therapy in combination with imaging options. However, the aqueous solubility of these hybrid compounds was insufficient to permit a full assessment of their biological activities. Further work in this direction is now underway, and will be reported in due course.

Commercially available reagents were used without additional purification. TLC analyses were carried out on Merck TLC silica gel 60 F254. Column chromatography was performed by using Macherey–Nagel Kieselgel 60 (70–230 mesh). IR spectra were recorded on a Shimadzu IR Prestige 21 spectrophotometer. NMR spectra were obtained with a Bruker AV 600, Bruker DRX 500, Bruker AV 400, Bruker ARX 400, or Bruker DPX 300 spectrometer, with the residual solvent peak as an internal reference. Assignments were made by comparison of chemical shifts, peak multiplicities, and J values. UV/Vis spectra (200–700 nm) were recorded on a PerkinElmer UV/VIS Lambda 25 spectrophotometer; emission spectra were obtained with a PerkinElmer FL 55 spectrophotometer. High-resolution mass-spectra were recorded on a Finnigan­ MAT 900 [EI (70 еV) and ESI] spectrometer.


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4-[(4-Bromo-2-fluorophenyl)amino]-6-methoxyquinazolin-7-yl 6-Azidohexanoate (3)

A Schlenk flask was degassed, filled with argon, and charged with EDC­·HCl (0.465 g, 2.42 mmol, 2.2 equiv) and 6-azidohexanoic acid (2; 0.346 g, 2.2 mmol, 2 equiv). Anhydrous DMF (7 mL) was added, and the mixture was stirred for 30 min at 0 °С. A second Schlenk flask was charged with quinazolinol 1 (0.4 g, 1.1 mmol) and DMAP (0.062 g, 0.55 mmol, 0.5 equiv). The flask was then filled with argon and anhydrous DMF (3 mL) was added. The mixture from the first flask was transferred into the second flask by using a syringe. The resulting mixture was stirred at 0 °С for 1 h and then at 60 °С for 1 h; it was then cooled and concentrated under reduced pressure. The residue was dissolved in EtOAc (80 mL), washed with H2O (3 × 50 mL), dried (Na2SO4), and concentrated. The product was purified by column chromatography (50% EtOAc, 50% cyclohexane) to give a white solid; yield: 0.501 g (91%); mp 157 °C.

IR (neat): 2091 cm–1 (azide group).

1H NMR (300 MHz, CDCl3): δ = 1.55 (tt, J = 9.8, 5.1 Hz, 2 H, СН2), 1.68 (dd, J = 15.0, 8.1 Hz, 2 H, СН2), 1.78–1.88 (m, 2 H, СН2), 2.68 (t, J = 7.3 Hz, 2 H, СН2), 3.33 (t, J = 6.7 Hz, 2 H, СН2), 3.96 (s, 3 H, CH3O), 7.09 (s, 1 H), 7.35 (m, 3 H), 7.56 (s, 1 H), 8.43 (t, J = 8.8 Hz, 1 H), 8.68 (s, 1 H).

13C NMR (75 MHz, CDCl3): δ = 24.55, 26.30, 28.72, 33.88, 51.40, 56.39, 100.21, 113.79, 116.03, 118.94, 122.22, 124.74, 126.34, 127.85, 146.07, 146.20, 151.17, 151.88, 153.39, 155.91, 171.27.

HRMS (ESI): m/z [M + Na]+ calcd for C21H20BrFN6O3 + Na: 525.0657; found: 525.0655 (error 0.2 ppm).


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Alkyne-Functionalized Chlorins 8a–d; General Procedure

The appropriate amine 5a–d was added to a solution of methylpheophorbide a (4) in CHCl3 (35 mL per gram of 4), and the mixture was stirred at r.t. until 4 was completely consumed (TLC). MeOH (15 mL per gram of 4) and Zn(OAc)2 were added, and the mixture was stirred for 3 h, then diluted with CH2Cl2 (100 mL) and washed with H2O (3 × 50 mL). The organic phase was dried (Na2SO4) and concentrated under reduced pressure. Because of their low stability, products 6a–с were used immediately in subsequent steps without further purifications.

To the residue (product 6a–c), the corresponding acid 7a or 7b, EDC­·HCl, HOBt, and DMAP were added under argon. Anhydrous CH2Cl2­ (5 mL) was added, and the mixture was stirred at r.t. for 15 h. The mixture was then diluted with CH2Cl2 (100 mL), washed with H2O (3 × 50 mL), dried (Na2SO4), and concentrated under reduced pressure. The product was purified by column chromatography.


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Chlorin 8a

Chlorin 8a was prepared according to the general procedure starting from 4 (0.128 g, 0.21 mmol), H2N(CH2)2NH2 (0.4 mL), and Zn(OAc)2 (0.23 g) in the first step to obtain product 6a as a deep-green solid; yield: 0.144 g (92%). In the second step, 6a (0.128 g, 0.175 mmol, 1 equiv), HOBt (0.049 g, 0.36 mmol, 2 equiv), EDC·HCl (0.069 g, 0.36 mmol, 2 equiv), DMAP (0.001 g, 0.089 mmol, 0.5 equiv), and acid 7a (0.078 g, 0.34 mmol, 1.9 equiv) were used. Column chromatography (1.5% MeOH, 1.5% Et3N, 97% CH2Cl2) gave 8a as a deep-green solid; yield: 0.156 g (93%); mp 155 °C.

1H NMR (300 MHz, CDCl3): δ = 0.88–0.93 (m), 1.21–1.24 (m), 1.28 (s), 1.48–1.55 (m), 1.58–1.64 (m), 1.69–1.73 (m), 1.95–1.97 (m), 2.02–2.04 (m), 2.11–2.16 (m), 2.22–2.25 (m), 2.29–2.34 (m), 2.43 (s), 2.50–2.55 (m), 2.86 (s), 3.19 (s), 3.24 (s), 3.30–3.36 (m), 3.56 (s), 3.65 (s), 3.69–3.72 (m), 4.24–4.27 (m), 4.30–4.36 (m), 4.58 (s), 5.89–5.98 (m), 6.05–6.17 (m), 6.56–6.63 (m), 6.93–7.03 (m), 7.02 (s), 7.39–7.42 (m), 8.55 (s), 9.45 (s).

The 13C NMR could not be properly recorded due to the insufficient molar concentrations of the compound achievable in all tested solvents.

HRMS (ESI): m/z [M + Na]+ calcd for C51H52N6O8Zn + Na: 963.3030; found: 963.3038 (error 1.0 ppm).


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Chlorin 8b

Chlorin 8b was prepared according to the general procedure starting from 4 (0.073 g, 0.12 mmol), amine 5b (0.5 mL), and Zn(OAc)2 (0.12 g) in the first step to give product 6b as a deep-green solid; yield: 0.098 g (99%).

In the second step, 6b (0.056 g, 0.0675 mmol, 1 equiv), HOBt (0.027 g, 0.21 mmol, 3 equiv), EDC·HCl (0.041 g, 0.21 mmol, 3 equiv), DMAP (0.005 g, 0.045 mmol, 0.7 equiv), and acid 7a (0.047 g, 0.2 mmol, 3 equiv) were used to obtain 8b. This was purified by filtration through a pad of silica gel (0.5% MeOH, 0.5% Et3N, 99% CH2Cl2) to give a deep-green solid; yield: 0.062 g (88%). Because of its low stability, compound 8b was used directly, without further purification, in the synthesis of compound 9b.

HRMS (ESI): m/z [M + Na]+ calcd for C55H60N6O10Zn + Na: 1051.3555; found: 1051.3574 (error 1.8 ppm).


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Chlorin 8c

Chlorin 8c was prepared according to the general procedure from 4 (0.069 g, 0.114 mmol), amine 5c (0.6 mL), and Zn(OAc)2 (0.125 g) in the first step to give product 6c as a deep-green solid; yield: 0.095 g (99%).

HRMS (ESI): m/z [M + Na]+ calcd for C43H56N8O5Zn + Na: 851.3568; found: 851.2401.

In the second step, 6c (0.095 g, 0.112 mmol, 1 equiv), EDC·HCl (0.046 g, 0.24 mmol, 2.1 equiv), DMAP (0.019 g, 0.171 mmol, 1.5 equiv), and acid 7b (0.024 g, 0.024 mmol, 2.1 equiv) were used to obtain crude 8c. This was purified by column chromatography (0.5% MeOH, 0.5% Et3N, 99% CH2Cl2) to give a deep-green solid; yield: 0.102 g (90%); mp 144 °C.

1H NMR (300 MHz, CDCl3): δ = 0.86–0.92 (m, 4 H), 1.26 (s, 1 H), 1.38 (s, 3 H), 1.52–1.56 (m, 4 H), 1.72 (dd, J = 7.2, 5.3 Hz, 9 H), 2.13–2.23 (m, 2 H), 2.46–2.59 (m, 7 H), 3.15 (s, 3 H), 3.24–3.29 (m, 5 H), 3.48–3.50 (m, 6 H), 3.62 (s, 3 H), 3.74–3.81 (m, 4 H), 3.85 (s, 4 H), 4.33 (d, J = 9.1 Hz, 1 H), 4.47 (q, J = 7.0 Hz, 1 H), 5.25 (dd, J = 19.0 Hz, 1 H), 5.29 (s, 1 H), 5.66 (d, J = 19.1 Hz, 1 H), 6.14 (d, J = 11.5 Hz, 1 H), 6.35 (d, J = 17.9 Hz, 1 H), 6.72 (t, J = 5.1 Hz, 2 H), 8.07 (dd, J = 17.8, 11.5 Hz, 1 H), 8.80 (s, 1 H), 9.62 (s, 1 H), 9.68 (s, 1 H).

13C NMR (75 MHz, CDCl3): δ = 9.94, 11.47, 12.19, 12.30, 14.21, 17.90, 19.82, 23.21, 29.74, 31.29, 34.13, 37.18, 37.87, 45.73, 49.39, 51.78, 52.38, 53.35, 93.83, 99.03, 101.70, 121.98, 129.48, 129.84, 134.85, 169.60, 171.79.

HRMS (ESI): m/z [M + Na]+ calcd for C53H64N8O7Zn + Na: 1012.2479; found: 1012.2463 (error 1.5 ppm).


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Chlorin 8d

Chlorin 8d was prepared according to the general procedure from 6c (0.083 g, 0.1 mmol), HOBt (0.028 g, 0.21 mmol, 2.1 equiv), EDC·HCl (0.04 g, 0.21 mmol, 2.1 equiv), DMAP (0.017 g, 0.017 mmol, 1.5 equiv), and 7a (0.048 g, 0.21 mmol, 2.1 equiv). It was purified by column chromatography (1% MeOH, 1% Et3N, 98% CH2Cl2) to give a deep-green solid; yield: 0.112 g (90%); mp >150 °C (dec).

1H NMR (500 MHz, CDCl3): δ = 0.82 (dd, J = 36.1, 5.7 Hz, 5 H), 1.56–1.58 (m, 2 H), 1.66 (t, J = 7.6 Hz, 4 H), 2.23 (s, 6 H), 3.21 (s, 3 H), 3.30 (s, 4 H), 3.35 (s, 4 H), 3.52 (d, J = 6.1 Hz, 5 H), 3.58 (d, J = 7.3 Hz, 4 H), 3.69 (s, 3 H), 4.16 (d, J = 8.7 Hz, 1 H), 4.26 (dd, J = 14.3, 7.0 Hz, 1 H), 4.39 (d, J = 1.8 Hz, 7 H), 4.85 (s, 2 H), 5.02 (d, J = 16.7 Hz, 1 H), 5.49 (d, J = 18.9 Hz, 1 H), 5.98 (dd, J = 11.5, 1.4 Hz, 1 H), 6.18 (dd, J = 17.8, 1.3 Hz, 1 H), 6.39 (s, 2 H), 7.06 (d, J = 1.9 Hz, 4 H), 7.85 (s, 2 H), 8.09 (dd, J = 17.8, 11.5 Hz, 1 H), 8.45 (s, 1 H), 9.34 (s, 1 H), 9.49 (s, 1 H).

13C NMR (126 MHz, CDCl3): δ = 5.61, 10.18, 11.34, 12.13, 12.58, 17.91, 19.59, 23.05, 29.62, 29.67, 30.81, 38.38, 38.69, 38.74, 45.91, 47.05, 50.53, 51.65, 52.13, 52.67, 53.24, 53.55, 54.24, 54.91, 55.98, 75.70, 92.94, 100.35, 101.73, 102.67, 104.67, 105.75, 106.73, 106.98, 119.28, 130.83, 130.83, 132.63, 132.94, 133.51, 136.73, 138.07, 139.63, 141.58, 141.82, 143.76, 145.17, 147.30, 148.14, 152.65, 153.16, 158.41, 164.89, 167.30, 171.92, 173.72, 174.75.

HRMS (ESI): m/z [M]+ calcd for C68H70N8O11Zn: 1239.4528; found: 1239.4531 (error 1.2 ppm).


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Chlorin–Quinazoline Conjugates 9a–d; General Procedure

Chlorin 8a–d (1 equiv) and an appropriate quantity of compound 3 were placed in a flask and treated with CuI, CH2Cl2, and H2O, and the mixture was stirred for 15 h. The precipitate that formed was collected, washed with CH2Cl2 and 0.01 M HCl, and dried under vacuum at 50 °С.


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Conjugate 9a

Prepared according to the general procedure from 8a (0.043 g, 0.046 mmol, 1 equiv), 3 (0.058 g, 0.115 mmol, 2.5 equiv), CuI (0.9 mg, 0.1 equiv), CH2Cl2 (0.5 mL), and H2O (0.04 mL) as a deep-green solid; yield: 0.89 g (99%); mp 176 °C.

1H NMR (500 MHz, DMF-d 7): δ = 1.42 (dt, J = 18.0, 7.5 Hz, 6 H), 1.60–1.83 (m, 12 H), 1.89–1.98 (m, 4 H), 2.14–2.33 (m, 2 H), 2.67 (t, J = 7.3 Hz, 4 H), 3.37 (d, J = 3.8 Hz, 6 H), 3.40 (s, 3 H), 3.57–3.64 (m, 5 H), 3.76 (s, 3 H), 3.95 (s, 6 H), 4.44 (dd, J = 15.3, 8.2 Hz, 4 H), 5.28 (s, 4 H), 5.52 (s, 1 H), 5.82 (s, 2 H), 6.01 (dd, J = 11.5, 1.6 Hz, 1 H), 6.25 (dd, J = 17.8, 1.7 Hz, 1 H), 7.00 (t, J = 2.0 Hz, 1 H), 7.00 (t, J = 2.0 Hz, 1 H), 7.34 (m, 3 H), 7.48 (dd, J = 8.5, 1.2 Hz, 2 H), 7.62–7.69 (m, 4 H), 8.05 (s, 2 H), 8.21–8.30 (m, 2 H), 8.33 (s, 2 H), 8.73 (d, J = 7.4 Hz, 2 H), 8.87 (s, 1 H), 9.62 (s, 1 H), 9.65 (s, 1 H), 9.83 (s, 2 H).

13C NMR (126 MHz, DMF-d 7): δ = 7.02, 10.52, 11.36, 11.91, 17.51, 22.55, 24.19, 25.62, 33.23, 37.70, 39.87, 40.12, 46.66, 49.63, 51.06, 51.50, 52.51, 52.59, 54.83, 56.32, 61.89, 62.86, 93.25, 100.32, 102.46, 103.24, 104.44, 106.67, 118.02 (2 С), 118.89, 119.36, 119.55, 120.98, 124.45, 126.59 (2 С), 127.67 (2 С), 129.62, 130.90, 132.74, 133.37, 133.91, 137.30, 137.94, 138.88, 141.32, 141.79, 143.04, 143.78, 144.99, 145.82 (2 С), 147.11, 148.50, 150.84, 152.58, 153.39, 156.03, 157.72, 158.03, 159.77, 161.87, 162.10, 162.34, 165.68, 166.43, 170.95, 171.05, 171.20, 173.48, 174.29.

HRMS (ESI): m/z [M + Na]+ calcd for C93H92Br2F2N18O14Zn + Na: 1967.4559; found: 1967.4570 (error 0.6 ppm).


#

Conjugate 9b

Prepared according to the general procedure from 8b (0.036 g, 0.035 mmol, 1 equiv), 3 (0.037 g, 0.074 mmol, 2.1 equiv), CuI (0.3 mg, 0.05 equiv), and CH2Cl2 (0.5 mL) as a deep-green solid; yield: 0.029 g (40%); mp 135 °C.

1H NMR (300 MHz, DMF-d 7): δ = 1.42–1.51 (m, 4 H), 1.64 (d, J = 7.1 Hz, 3 H), 1.71 (t, J = 7.6 Hz, 4 H), 1.79 (p, J = 7.5 Hz, 6 H), 1.97 (p, J = 7.2 Hz, 4 H), 2.70 (t, J = 7.1 Hz, 5 H), 3.35–3.43 (m, 11 H), 3.60 (d, J = 14.0 Hz, 7 H), 3.68–3.78 (m, 8 H), 3.80 (dd, J = 6.0, 3.7 Hz, 3 H), 3.86 (d, J = 7.7 Hz, 4 H), 3.90–4.04 (m, 9 H), 4.40–4.55 (m, 6 H), 5.25 (s, 4 H), 5.46 (dd, J = 103.0, 18.6 Hz, 2 H), 6.02 (dd, J = 11.4, 1.7 Hz, 1 H), 6.26 (dd, J = 17.8, 1.8 Hz, 1 H), 7.27 (d, J = 2.3 Hz, 2 H), 7.38 (s, 1 H), 7.47–7.52 (m, 2 H), 7.62–7.69 (m, 4 H), 8.06 (s, 2 H), 8.25–8.35 (m, 4 H), 8.54 (t, J = 5.7 Hz, 1 H), 8.75 (s, 1 H), 9.65 (d, J = 11.6 Hz, 2 H), 9.88 (s, 2 H).

13C NMR (151 MHz, DMF-d 7): δ = 11.57, 12.37, 12.96, 18.56, 20.13, 23.63, 25.28, 26.63, 30.19, 30.25, 30.34, 30.39, 30.49, 30.53, 30.63, 30.67, 30.78, 30.92, 31.07, 31.56, 34.31, 35.30, 35.45, 35.50, 35.60, 35.64, 35.74, 35.90, 35.92, 36.05, 36.19, 38.70, 40.81, 40.98, 47.68, 50.76, 52.11, 52.53, 53.63, 57.38, 62.93, 70.52, 71.26, 94.28, 101.33, 103.48, 103.57, 104.34, 105.36, 107.70, 119.12, 119.87, 120.44, 120.59, 122.06, 125.59, 127.52, 128.74, 130.69, 131.97, 133.71, 134.39, 135.21, 138.18, 138.93, 140.09, 142.27, 142.96, 144.82, 145.95, 146.82, 148.03, 149.69, 151.95, 153.54, 157.24, 158.77, 160.70, 166.62, 167.16, 171.77, 171.99, 174.53, 175.30.

HRMS (ESI): m/z [M + H]+ calcd for C97H101Br2F2N18O16Zn: 2033.5264; found: 2033.5241 (error 1.1 ppm).


#

Conjugate 9c

Prepared according to the general procedure from 8с (0.044 g, 0.045 mmol, 1 equiv), 3 (0.056 g, 0.11 mmol, 2.5 equiv), CuI (1 mg, 0.1 equiv), CH2Cl2 (0.5 mL), and H2O (0.04 mL) as a deep-green solid; yield: 0.0639 g (71%); mp 174 °C.

1H NMR (600 MHz, DMF-d 7): δ = 1.67–1.81 (m, 12 H), 2.20–2.29 (m, 2 H), 2.64 (t, J = 6.9 Hz, 4 H), 2.73–2.80 (m, 14 H), 2.94 (tt, J = 3.7, 1.9 Hz, 10 H), 3.40 (s, 8 H), 3.43 (s, 6 H), 3.45–3.55 (m, 7 H), 3.60–3.65 (m, 4 H), 3.79 (s, 3 H), 3.82–3.91 (m, 3 H), 3.94 (d, J = 22.8 Hz, 5 H), 4.19 (s, 4 H), 4.49 (dd, J = 53.7, 8.2 Hz, 2 H), 5.36 (d, J = 16.3 Hz, 1 H), 5.51–5.63 (m, 1 H), 5.84 (s, 1 H), 6.04 (d, J = 11.7 Hz, 1 H), 6.29 (d, J = 18.0 Hz, 1 H), 7.51 (d, J = 8.4 Hz, 2 H), 7.67 (t, J = 8.4 Hz, 4 H), 7.87 (s, 2 H), 8.32 (dd, J = 17.8, 11.5 Hz, 1 H), 8.62 (s, 1 H), 8.78 (s, 1 H), 9.67 (s, 1 H), 9.70 (s, 1 H), 9.98 (s, 1 H).

13C NMR (151 MHz, DMF-d 7): δ = 8.05, 11.56, 12.56, 12.97, 18.54, 20.12, 22.56, 23.57, 25.31, 26.37, 26.99, 29.43, 31.31, 34.30, 38.41, 38.72, 39.20, 47.69, 50.61, 52.10, 52.57, 53.55, 53.62, 54.93, 55.16, 55.87, 57.39, 94.35, 101.41, 103.48, 104.47, 119.32, 119.92, 120.46, 120.62, 127.33, 127.41, 128.75, 128.78, 130.75, 131.95, 133.78, 134.47, 135.25, 138.99, 139.97, 142.33, 142.95, 144.82, 146.02, 146.71, 148.08, 149.66, 152.08, 153.61, 157.24, 158.91, 166.68, 171.88, 174.52, 175.23.

HRMS (ESI): m/z [M + 2Na]2+ calcd for C95H104Br2F2N20O13Zn + 2Na: 1021.5667; found: 1021.5672 (error 0.5 ppm).


#

Conjugate 9d

Prepared according the general procedure from 8d (0.042 g, 0.034 mmol, 1 equiv), 3 (0.085 g, 0.17 mmol, 5 equiv), CuI (1.3 mg, 1 equiv), CH2Cl2 (0.5 mL), and H2O (0.04 mL) as a deep-green solid; yield: 0.065 g (60%); mp 166 °C.

1H NMR (600 MHz, DMF-d 7): δ = 1.38–1.44 (m), 1.63–1.64 (m), 1.70–1.72 (m), 1.75–1.77 (m), 1.81–1.83 (m), 1.86–1.89 (m), 2.68–2.70 (m), 3.04 (s), 3.26 (s), 3.37 (s), 3.44–3.47 (m), 3.61–3.64 (m), 3.68–3.71 (m), 3.77 (s), 3.82–3.86 (m), 3.98 (s), 4.00–4.04 (m), 4.35–4.37 (m), 4.42–4.45 (m), 4.48–4.52 (m), 5.04 (br s), 6.02–6.04 (m), 6.27–6.30 (m), 6.84 (s), 7.28 (s), 7.50–7.54 (m), 7.65–7.70 (m), 8.07 (s), 8.14–8.20 (m), 8.29–8.34 (m), 8.56 (s), 8.60–8.62 (m), 8.68 (br s), 8.76 (s), 9.49 (s), 9.67 (s), 9.88–9.90 (m), 10.00 (s).

13C NMR (151 MHz, DMF-d 7): δ = 10.54, 11.38, 11.98, 17.54, 19.16, 24.18, 25.57, 33.25, 38.04, 38.37, 46.69, 49.61, 51.06, 51.52, 52.57, 53.78, 54.83, 56.35, 56.67, 61.57, 90.19, 93.26, 103.43, 104.38, 106.46, 118.78, 119.39, 119.55, 127.71, 129.62, 130.96, 137.18, 139.07, 143.73, 144.93, 145.69, 150.95, 154.13, 156.19, 159.64, 166.33, 170.98, 173.49.

HRMS (ESI): m/z [M + 2H]2+ calcd for C152H152Br4F4N32O23Zn: 1624.3829; found: 1624.3825 (error 0.2 ppm).


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Conjugate 11

Chlorin 10 (0.05 g, 0.067 mmol) and compound 3 (0.044 g, 0.087 mmol, 1.3 equiv) were dissolved in CH2Cl2 (2 mL). CuI (1.3 mg, 0.0067 mmol, 0.1 equiv) was added and the mixture was stirred at r.t. for 15 h. The solvent was removed under reduced pressure, and the residue was purified by column chromatography (1% MeOH, 1% Et3N, 98% CH2Cl2) to give a deep-green solid; 0.054 g (64%); mp 182 °C.

1H NMR (600 MHz, CDCl3): δ = 1.05–1.06 (m), 1.90–1.92 (m), 2.15–2.16 (m), 2.19–2.20 (m), 2.84 (br s), 3.28 (s), 3.39–3.50 (m), 3.61 (s), 3.65 (s), 3.74 (br s), 4.28–4.29 (m), 4.74–4.75 (m), 5.29 (s), 5.95–5.97 (m), 6.13–6.16 (m), 6.74 (br s), 6.95 (br s), 7.98–8.03 (m), 8.43 (s), 9.10 (br s), 9.41 (s).

13C NMR (126 MHz, CDCl3): δ = 5.61, 10.18, 11.34, 12.13, 12.58, 17.91, 19.59, 23.05, 29.67 (2 С), 30.81, 38.38, 38.74 (2 С), 45.91, 47.05, 50.53, 51.65, 52.13, 52.67, 53.24–53.55 (3 C), 54.24, 54.91, 55.98, 75.70, 92.94, 100.35, 101.73, 102.67, 104.67, 105.75, 106.73, 106.98, 119.28, 130.83, 130.83, 132.63, 132.94, 133.51, 136.73, 138.07, 139.63, 141.58, 141.82, 143.76, 145.17, 147.30, 148.14, 152.65, 153.16, 158.41, 164.89, 167.30, 171.92, 173.72, 174.75.

HRMS (ESI): m/z [M + H]+ calcd for C61H64BrFN11O8Zn: 1240.3393; found: 1240.3381 (error 1.0 ppm).


#

Chlorin 12

Synthesized according to the general procedure for compounds 8a–d from aminochlorin 6a (0.116 g, 0.156 mmol), 6-azidohexanoic acid (2; 0.073 g, 0.47 mmol, 3 equiv), EDC·HCl (0.096 g, 0.5 mmol, 3.5 equiv), and DMAP (0.012 g, 0.11 mmol, 0.7 equiv). The product was purified by column chromatography (1% MeOH, 1% Et3N, 98% CH2Cl2) to give a deep-green solid; yield: 0.136 g (99%) solid; mp >150 °С (dec).

IR (neat): 2090 cm–1 (azide group).

1H NMR (500 MHz, CDCl3): δ = 1.16 (t, J = 7.3 Hz, 2 H), 1.59 (d, J = 7.2 Hz, 4 H), 1.75 (d, J = 7.6 Hz, 4 H), 2.24 (s, 6 H), 2.39–2.46 (m, 1 H), 2.68 (s, 1 H), 3.20–3.29 (m, 5 H), 3.35 (d, J = 18.2 Hz, 7 H), 3.56 (s, 3 H), 3.68 (s, 3 H), 3.73–3.88 (m, 2 H), 4.17–4.31 (m, 2 H), 4.88 (s, 2 H), 5.11 (d, J = 19.1 Hz, 1 H), 5.38 (dd, J = 41.9, 11.1 Hz, 1 H), 5.99 (dd, J = 11.5, 1.6 Hz, 1 H), 6.20 (dd, J = 17.8, 1.6 Hz, 1 H), 8.11 (dd, J = 17.8, 11.5 Hz, 1 H), 8.46 (s, 1 H), 9.48 (s, 1 H), 9.52 (s, 1 H).

13C NMR (126 MHz, CDCl3): δ = 12.62, 14.89, 17.89, 19.70, 22.99, 24.61, 25.89, 26.44, 28.86, 29.50, 30.70, 35.46, 36.23, 38.40, 40.49, 43.02, 45.84, 47.23, 50.99, 51.28, 51.38, 51.60, 52.77, 92.95, 100.41, 101.62, 102.97, 104.68, 119.21, 130.90, 131.91, 133.53, 138.21, 139.35, 141.69, 143.95, 145.55, 147.64, 147.73, 152.93, 153.18, 165.10, 172.76, 173.73.

HRMS (ESI): m/z [M + Na]+ calcd for C44H53N9O6Zn + Na: 890.3302; found: 890.3308 (error 0.6 ppm).


#

7-{[3,5-Bis(prop-2-yn-1-yloxy)benzyl]oxy}-N-(4-bromo-2-fluorophenyl)-6-methoxyquinazolin-4-amine (14)

Quinazoline 1 (0.101 g, 0.28 mmol) and diyne 13 (0.078 g, 0.28 mmol) were dissolved in DMF (3 mL). K2CO3 (0.077 g, 2 equiv) was added, and the mixture stirred at 60 °С for 14 h. The solvent was removed under reduced pressure, and the residue was dissolved in CH2Cl2 (100 mL), washed with H2O (3 × 20 mL), dried (Na2SO4), and concentrated. The product was purified by column chromatography (50% EtOAc, 50% cyclohexane) to give a gray solid; yield: 0.091 g (58%); mp 168 °C.

1H NMR (600 MHz, DMSO-d6): δ = 3.57 (t, J = 2.4 Hz, 2 H), 3.97 (s, 3 H, CH3O), 4.80 (d, J = 2.1 Hz, 4 H, CH2CCH), 5.23 (s, 2 H), 6.63 (d, J = 1.9 Hz, 1 H), 6.76 (d, J = 1.7 Hz, 2 H), 7.28 (s, 1 H), 7.42–7.49 (m, 1 H), 7.53 (t, J = 8.3 Hz, 1 H), 7.66 (dd, J = 9.9, 2.2 Hz, 1 H), 7.83 (s, 1 H), 8.35 (s, 1 H), 9.56 (s, 1 H).

13C NMR (151 MHz, DMSO-d6): δ = 55.62, 56.19, 69.68, 78.39, 101.42, 102.11, 107.23, 108.42, 108.85, 117.54, 117.60, 119.26, 119.41, 126.34, 126.42, 127.51, 129.55, 138.75, 146.79, 149.13, 152.95, 153.18, 155.82, 156.91, 157.49, 158.42.

HRMS (ESI): m/z [M]+ calcd for C28H21BrFN3O4: 562.0772; found: 562.0771 (error 0.3 ppm).


#

Conjugate 15

Synthesized according to the general procedure for compounds 9a–d from compound 12 (0.056 g, 0.11 mmol, 2.5 equiv), quinazoline 14 (0.044 g, 0.044 mmol), CuI (0.9 mg, 0.0045 mmol, 0.1 equiv), DMF (2 mL), and H2O (0.05 mL) as a deep-green solid; yield: 0.059 g (57%); mp 187 °C.

1H NMR (400 MHz, DMF-d 7): δ = 1.35–1.41 (m), 1.58–1.78 (m), 1.91–1.97 (m), 2.13 (s), 2.16–2.18 (m), 2.26–2.30 (m), 2.60–2.66 (m), 3.36–3.41 (m), 3.59 (s), 3.60–3.60 (m), 3.73–3.80 (m), 3.84–3.90 (m), 4.44–4.52 (m), 4.90–5.03 (m), 5.29 (s), 5.52–5.57 (m), 5.83 (s), 6.00–6.04 (m), 6.24–6.28 (m), 6.86 (s), 7.05 (s), 7.36–7.38 (m), 7.47–7.60 (m), 8.03–8.05 (m), 8.20–8.39 (m), 8.72–8.75 (m), 8.76 (s), 9.49 (s), 9.65–9.67 (m).

13C NMR (101 MHz, DMF-d 7): δ = 8.05, 11.55, 12.53, 12.97, 18.57, 20.12, 23.56, 25.92, 26.82, 33.72, 36.62, 37.03, 38.74, 40.01, 41.00, 47.72, 50.67, 52.08, 52.10, 52.51, 53.63, 55.88, 56.84, 62.84, 71.75, 94.31, 101.36, 102.56, 103.59, 108.58, 119.94, 120.26, 120.45, 125.65, 128.56, 128.59, 130.51, 130.53, 131.92, 133.81, 134.50, 135.09, 139.04, 139.90, 140.01, 142.36, 142.82, 144.90, 146.02, 148.22, 149.49, 150.45, 153.65, 154.53, 156.88, 158.89, 161.11, 171.98, 173.64, 173.74, 174.47, 174.55, 175.17.

HRMS (ESI): m/z [M + Na]+ calcd for C118H131BrFN21O16Zn2 + Na: 2346.3579; found: 2346.3571 (error 0.3 ppm).


#
#

Acknowledgment

We thank the Ministry of Education and Science of the Russian Federation (Project 4.619.2014/K), and the German Academic Exchange Service (DAAD fellowship to A.V.N.). The research is partly supported by grant number 02.B.49.21.0003 of The Ministry of Education and Science of the Russian Federation to N.I. Lobachevsky State University of Nizhny Novgorod. The study was carried out with support from the Ministry of Education and Science of the Russian Federation for the equipment of the Collective Usage Center ‘New Materials and Resource­-Saving Technologies’ of Lobachevsky State University of Nizhny Novgorod (Project RFMEFI59414X0005).

Supporting Information

    Supporting information for this article is available online at http://dx.doi.org.accesdistant.sorbonne-universite.fr/10.1055/s-0034-1378876.
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Scheme 1 Functionality-based design, synthesis strategy, and expected properties of hybrid molecules for photodynamic therapy
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Scheme 2 Synthesis of the EGFR/VEGFR ligand core (1) and its azide-functionalized derivative 3
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Scheme 3 Synthesis of alkyne-functionalized chlorin e 6 derivatives
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Scheme 4 Synthesis of conjugates 9a–c
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Scheme 5 Synthesis of conjugates 9d and 11
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Scheme 6 Synthesis of conjugate 15