Chi-Chih Kang

Investigation of spectral conversion of d(TTAGGG)4 and d(TTAGGG)13 upon potassium titration by a G-quadruplex recognizer BMVC molecule.

We have introduced a G-quadruplex-binding ligand, 3,6-bis(1-methyl-4-vinylpyridinium)carbazole diiodide (BMVC), to verify the major structure of d(T2AG3)4 (H24) in potassium solution and examine the structural conversion of H24 in sodium solution upon potassium titration. The studies of circular dichroism, induced circular dichroism, spectral titration and gel competition have allowed us to determine the binding mode and binding ratio of BMVC to the H24 in solution and eliminate the parallel form as the major G-quadruplex structure. Although the mixed-type form could not be eliminated as a main component, the basket and chair forms are more likely the main components of H24 in potassium solution. In addition, the circular dichroism spectra and the job plots reveal that a longer telomeric sequence d(T2AG3)13 (H78) could form two units of G4 structure both in sodium or potassium solutions. Of particular interest is that no appreciable change on the induced circular dichroism spectra of BMVC is found during the change of the circular dichroism patterns of H24 upon potassium titration. Considering similar spectral conversion detected for H24 and a long sequence H78 together with the G4 structure stabilized by BMVC, it is therefore unlikely that the rapid spectral conversion of H24 and H78 is due to structural change between different types of the G4 structures. With reference to the circular dichroism spectra of d(GAA)7 and d(GAAA)5, we suggest that the spectral conversion of H24 upon potassium titration is attributed to fast ion exchange resulting in different loop base interaction and various hydrogen bonding effects.

Improved diagnostic accuracy of malignant neck lumps by a simple BMVC staining assay

A handheld device based on fluorescence of 3,6-bis(1-methyl-4-vinylpyridinium)carbazole diiodide (BMVC) staining was established for the rapid, point-of-care screening of cancer cells (see Chang and co-workers, Analyst, 2007, 132, 745). Offering instant screening of cancer at low cost, here we apply this simple assay in clinical tests on fine needle aspirates of neck masses from 114 outpatients (115 specimens). The diagnostic accuracy of this simple method alone is ca. 80% (80/99). The combination of the BMVC test and the fine needle aspiration (FNA) cytology reduced the non-diagnosis from 17 cases in FNA cytology to 6 cases in the combined method. Moreover, an algorithm is proposed to improve the diagnostic accuracy of malignant neck lumps up to nearly 100%.

A Dual Selective Antitumor Agent and Fluorescence Probe: the Binary BMVC–Porphyrin Photosensitizer

Photodynamic therapy (PDT) is a promising modality for the treatment of localized tumors. To cause cell death during PDT, singlet oxygen can be generated by energy transfer from the triplet state of a photosensitizer to molecular oxygen. Among various photosensitizers, porphyrin derivatives have been widely studied. 6] However, porphyrin-based sensitizers have drawbacks to their application in PDT that include poor chemical selectivity toward intended tissue targets and a lack of specific light wavelengths that are optimal for tissue penetration and chromophore excitation. To overcome these shortcomings of the traditional porphyrin chromophore, a number of binary compounds have been recently designed and prepared with a component linked to the porphyrin photosensitizer. The resulting conjugates are highly selective for cancer cells and absorb at longer wavelengths more suitable for tissue penetration. For example, Drain and co-workers synthesized porphyrin–saccharide conjugates to increase the uptake of the photosensitizer by specific cancer cells. Dichtel et al. synthesized a binary compound to enhance singlet oxygen generation via fluorescence resonance energy transfer (FRET) by two-photon excitation of the donor chromophore to the central porphyrin acceptor. Selectivity is a key feature in the design of photosensitizers for killing cancer cells without damaging normal cells. We recently synthesized a novel 3,6-bis-(1-methyl-4-vinylpyridinium)carbazole diiodide (BMVC) compound with selectivity toward cancer cells over normal cells. 13] Specifically, the fluorescence of BMVC detected in cancer cells was found to be much stronger than that in normal cells. Moreover, BMVC has a large cross-section for two-photon absorption around 820 nm, a wavelength that is close to optimal for tissue penetration of the radiation needed to excite the photosensitizer. In addition, the windows of transparency for porphyrin derivatives in the range of 450–500 nm allow us to selectively excite BMVC. For the study reported herein, we combined these features into a binary porphyrin compound for selectivity in PDT. In this compound, two BMVC molecules are covalently linked to a central 5,10-bis-(4-hydroxyphenyl)-15,20-bis-(4-methoxyphenyl)porphyrin (PAP) molecule at the ortho positions, and we termed the resulting binary compound o-2B-P. The ortho position was chosen for covalent attachment, as it is known that compounds containing ortho-5,10-disubstituted water-soluble trimethylaminophenyl groups exhibit better PDT efficacy than those with para-5,15-disubstitution. The structure of o-2B-P is shown in Figure 1 A, and details of its synthesis can be found elsewhere (Supporting Information 1). We examined the efficiency of energy transfer from the excited state of BMVC to the porphyrin in o-2B-P, as well as the efficacy of singlet oxygen generation by the porphyrin in the binary compound. The absorption and fluorescence spectra of o-2B-P, PAP, BMVC, and a mixture of BMVC and PAP in DMSO are depicted in Figures 1 B and C, respectively. The absorption of o-2B-P is almost identical to that of a mixture of BMVC and PAP. The absorption is a linear sum of the absorbances of BMVC and PAP individually, indicating that there is no appreciable interaction between the two chromophores in the ground electronic states. In contrast, the fluorescence of BMVC in o-2BP is almost totally quenched, whereas the fluorescence of PAP is enhanced by at least fivefold upon excitation of o-2B-P at 470 nm. Evidently, there is efficient energy transfer from the excited state of BMVC to PAP. To evaluate the effect of photoinduced formation of singlet oxygen by o-2B-P, we applied a photochemical method using a singlet oxygen quencher, 1,3-diphenylisobenzofuran (DPBF), to verify the generation of singlet oxygen. Because the cationic meso-tetra-(4-N-methylpyridyl)porphyrin (TMPyP) molecule is itself a singlet oxygen photosensitizer, 20] we compared the yield of singlet oxygen upon irradiation of o-2B-P and TMPyP at various wavelengths. Figure 1 D shows a comparison of the absorbance of DPBF at 417 nm in the presence of o-2B-P and TMPyP in DMSO as a function of irradiation time at 470 10 nm as well as lex>580 nm selectively from a halogen lamp. Although singlet oxygen generation by o-2B-P is slightly lower than that of TMPyP at lex>580 nm, it is the singlet oxygen generated by o-2B-P, not by TMPyP, upon excitation at 470 10 nm that provides a more useful measure of the selectivity for PDT efficacy. Despite the large number of porphyrin derivatives in blood and tissues, the porphyrin absorbance at 470 10 nm is almost negligible. Accordingly, the 450–500 nm transparent windows of porphyrin derivatives [a] C.-C. Kang, Dr. C.-C. Cho, Y.-C. Lin, Prof. C.-C. Chang, Prof. T.-C. Chang Institute of Atomic and Molecular Sciences, Academia Sinica P.O. Box 23-166, Taipei 106 (Taiwan, ROC) Fax: (+ ) 886-2-23620200 E-mail : tcchang@po.iams.sinica.edu.tw [b] C.-C. Kang Taiwan International Graduate Program and Department of Chemistry National Tsing-Hua University and Academia Sinica (Taiwan, ROC) [c] Prof. C.-T. Chen Centers for Optoelectronic Biomedicine, College of Medicine National Taiwan University, Taipei (Taiwan, ROC) and Institute of Microbiology and Biochemistry, College of Life Science National Taiwan University, Taipei (Taiwan, ROC) [d] Prof. C.-C. Chang Department of Chemistry, National Chung-Hsing University Taichung (Taiwan, ROC) E-mail : ccchang555@dragon.nchu.edu.tw Supporting information for this article is available on the WWW under http://www.chemmedchem.org or from the author.

BMVC test, an improved fluorescence assay for detection of malignant pleural effusions

The diagnosis of malignant pleural effusions is an important issue in the management of malignancy patients. Generally, cytologic examination is a routine diagnostic technique. However, morphological interpretation of cytology is sometimes inconclusive. Here an ancillary method named BMVC test is developed for rapid detection of malignant pleural effusion to improve the diagnostic accuracy at low cost. A simple assay kit is designed to collect living cells from clinical pleural effusion and a fluorescence probe, 3,6‐Bis(1‐methyl‐4‐vinylpyridinium) carbazole diiodide (BMVC), is used to illuminate malignant cells. The fluorescence intensity is quantitatively analyzed by ImageJ program. This method yields digital numbers for the test results without any grey zone or ambiguities in the current cytology tests due to intra‐observer and inter‐observer variability. Comparing with results from double‐blind cytologic examination, this simple test gives a good discrimination between malignant and benign specimens with sensitivity of 89.4% (42/47) and specificity of 93.3% (56/60) for diagnosis of malignant pleural effusion. BMVC test provides accurate results in a short time period, and the digital output could assist cytologic examination to become more objective and clear‐cut. This is a convenient ancillary tool for detection of malignant pleural effusions.