Article pubs.acs.org/jmc
Phthalocyanine Labels for Near-Infrared Fluorescence Imaging of Solid Tumors Ana C. S. Lobo,†,§ Alexandre D. Silva,†,§ Vanessa A. Tomé,† Sara M. A. Pinto,† Elsa F. F. Silva,† Mário J. F. Calvete,† Célia M. F. Gomes,‡ Mariette M. Pereira,† and Luis G. Arnaut*,† †
CQC, Chemistry Department, University of Coimbra, 3004 - 535 Coimbra, Portugal Laboratory of Pharmacology and Experimental Therapeutics, IBILI, Faculty of Medicine, University of Coimbra, 3000-548 Coimbra, Portugal
‡
S Supporting Information *
ABSTRACT: Diamagnetic metal complexes of phthalocyanines with n-butoxyl groups in all the α-benzo positions of the macrocycle skeleton, MPc(OBu)8, have strong near-infrared absorptions and intense fluorescences that are Stokes shifted by more than 15 nm. Interestingly, the silicon complex 6 is also remarkably photostable and nontoxic. The use of 6 in the fluorescence imaging of BALB/c mice bearing a 4T1-luc2 tumor in the mammary fat pad unambiguously revealed the presence of the tumor when it was only 1 mm in diameter and was not visible with the naked eye. Compound 6 has an intrinsic ability to accumulate in the tumor, adequate spectroscopic properties, and excellent stability to function as a NIR fluorescent label in the early detection of tumors.
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INTRODUCTION Imaging has become an indispensable tool in cancer research, clinical trials, and medical practice.1 Fluorescence imaging is minimally invasive, affordable, and portable, with the potential to become a widespread medical imaging technique. However, adequate contrast agents are still needed to attain the sensitivity and specificity required for early detection of tumors. Fluorophores for image-guided surgery, for induced fluorescence endoscopy, or for biomarkers require high molar absorption coefficients in the near-infrared (NIR), large fluorescence quantum yields (ΦF) and large Stokes shifts, low photodecomposition quantum yields (Φpd) and low toxicity. Strong absorption in the “imaging window” (750−900 nm) avoids endogenous tissue absorption and autofluorescence and allows for deep-tissue imaging.2,3 Intense fluorescence increases sensitivity, and large Stokes shifts enable the separation between excitation and fluorescence wavelengths, thus reducing background light. Most molecular fluorophores suffer from relatively fast bleaching in biological media,4 which poses an additional challenge to imaging. The use of bright nanoparticles, more photostable, such as semiconductor quantum dots,5,6 is still restricted by concerns of clearance and toxicity. Indocyanine green (ICG) remains the only FDA-approved long wavelength probe for medical diagnostics, namely for sentinel lymph node mapping. This has inspired many efforts to improve optical probes, and the majority of NIR chromophores that have been proposed as optical imaging agents are cyanine dyes.7−9 Cyanine dyes do not have specificity for tumors. This may be overcome by chemically conjugating the dye with a © XXXX American Chemical Society
tumor-specific vehicle or with a sensor that activates the dye in the tumor microenvironment. However, it would be preferable to devise molecular fluorophores with intrinsic tumor-localizing abilities that could be economically prepared in a large scale and could be used in simple, nontoxic, nonimmunogenic formulations. Tetrapyrrolic macrocycles, such as reduced porphyrins (e.g., chlorins, bacteriochlorins) and phthalocyanines (Pc), combine strong absorptions in the “imaging window” with a natural tendency to accumulate in tumors. For example, tumor:peritumoral tissue ratios in excess of 5 were reported for hydrophobic phthalocyanines.10 Some lipophilicity improves tumor:muscle ratios, but excessive lipophilicity leads to aggregation and requires the use of vehicles that represent an additional burden to the organism. We have shown that the tendency of tetraphenylporphyrin derivatives to aggregate is opposed by ortho-substitution of the phenyl rings.11 The twisted phenyl rings hinder the cofacial interaction of the macrocycles and the formation of aggregates. Although orthosubstituted tetraphenylchlorins and tetraphenylbacteriochlorins can be strongly fluorescent, their Stokes shifts are usually small (
