Delocalized Lipophilic Cations Research Articles

Formulation of Boron Encapsulated Smart Nanocapsules for Targeted Drug Delivery to the Brain

Drug delivery through the Blood–Brain Barrier (BBB) represents a significant challenge. Despite the current strategies to circumvent the BBB, nanotechnology offers unprecedented opportunities for combining selective delivery, improved bioavailability, drug protection, and enhanced pharmacokinetics profiles. Chitosan nanocarriers allow for a more efficacious strategy at the cellular and sub-cellular levels. Boron Neutron Capture Therapy (BNCT) is a targeted chemo-radiotherapeutic technique that allows the selective depletion of cancer cells by means of selective tagging of cancer cells with 10B, followed by irradiation with low-energy neutrons. Consequently, the combination of a polymer-based nanodelivery system enclosing an effective BNCT pharmacophore can potentially lead to the selective delivery of the load to cancer cells beyond the BBB. In this work, synthesized novel boronated agents based on carborane-functionalized Delocalized Lipophilic Cations (DLCs) are assessed for safety and selective targeting of tumour cells. The compounds are then encapsulated in nanocarriers constituted by chitosan to promote permeability through the BBB. Additionally, chitosan was used in combination with polypyrrole to form a smart composite nanocapsule, which is expected to release its drug load with variations in pH. Results indicate the achievement of more selective boron delivery to cells via carboranyl DLCs. Finally, preliminary cell studies indicate no toxicity was detected in chitosan nanocapsules, further enhancing its viability as a potential delivery vehicle in the BNCT of brain tumours.

Cyanine mitochondrial dye with slightly selective cytotoxicity against A549 cancerous cells.

Delocalized lipophilic cations (DLCs) are known as mitochondria-addressed molecules. Mitochondria targeting may provide opportunities for tumor detection. DLCs may have antioxidant or anticancer properties. In this study, we focused on the toxicity and localization of 2-[(E)-2-(5-fluoro-2-methyl-1H-indol-3-yl)ethenyl]-1,6-dimethylpyridin-1-ium iodide (62E2), which has recently been found as a novel cytotoxic fluorescent compound. The excitation maximum of 62E2 is 452 ± 10 nm and its emission maximum is 579 ± 10 nm. It is accumulated in the cells and stains mitochondria in nanomolar concentrations. 62E2 is cytotoxic and mitotoxic in low micromolar concentrations, and it demonstrates some selectivity of cytotoxicity against A549 cancer cells. The closest analog of 62E2 is F16, which is the fluorescent mitotoxic agent that has been described earlier as a potential anticancer agent. We hope that 62E2 described here is useful in expanding the diversity of cyanine fluorescent mitochondrial dyes and the analysis of their structure-activity relationships.

Anionic Polymers Promote Mitochondrial Targeting of Delocalized Lipophilic Cations.

Mitochondria are therapeutic targets in many diseases including cancer, metabolic disorders, and neurodegenerative diseases. Therefore, strategies to deliver therapeutics of interest to mitochondria are important for therapeutic development. As delocalized lipophilic cations (DLCs) preferentially accumulate in mitochondria, DLC-conjugation has been utilized to facilitate therapeutic delivery systems with mitochondrial targeting capability. Here we report that upon DLC-conjugation, anionic polymers exhibit significantly improved mitochondrial targeting when compared to cationic polymers and charge-neutral polymers. Considering that the cell membrane generally bears a net negative charge, the observed phenomenon is unexpected. Notably, the DLC-conjugated anionic polymers circumvent endosomal entrapment. The rapid mitochondrial accumulation of DLC-conjugated anionic polymers is likely a membrane-potential-driven process, along with the involvement of the mitochondrial pyruvate carrier. Moreover, the structural variations on the side chain of DLC-conjugated anionic polymers do not compromise the overall mitochondrial targeting capability, widely extending the applicability of anionic macromolecules in therapeutic delivery systems.

Mitochondria-targeting fluorescent molecules for high efficiency cancer growth inhibition and imaging.

Fluorescent mitochondria-accumulating delocalized lipophilic cations (DLCs) for cancer therapy have drawn significant attention in the field of cancer theranostics. One of the most promising fluorescent DLCs, F16, can selectively trigger the apoptosis and necrosis of cancer cells, making it an attractive targeted theranostic drug candidate. However, it suffers from low clinical translation potential, largely due to its inefficient anti-cancer activity (IC50 in the μM range) and poorly understood structure-activity relationship (SAR). In this report, eleven indole-ring substituted F16 derivatives (F16s) were synthesized. Among these derivatives, 5BMF was identified as a highly effective theranostic agent, with in vitro studies showing a low IC50 of ∼50 nM (to H2228 cells) and high cancer to normal cell selectivity index of 225. In vivo studies revealed that tumors treated with 5BMF were significantly suppressed (almost no growth over the treatment period) compared to the PBS treated control group, and also no obvious toxicity to mice was found. In addition, the tumor imaging capability of 5BMF was demonstrated by in vivo fluorescence imaging. Finally, we report for the first time a proposed SAR for F16 DLCs. Our work lays down a solid foundation for translating 5BMF into a novel and highly promising DLC for cancer theranostics.

ROS‐Responsive N‐Alkylaminoferrocenes for Cancer‐Cell‐Specific Targeting of Mitochondria

AbstractMitochondrial membrane potential is more negative in cancer cells than in normal cells, allowing cancer targeting by delocalized lipophilic cations (DLCs). However, as the difference is rather small, these drugs affect also normal cells. Now a concept of pro‐DLCs is proposed based on an N‐alkylaminoferrocene structure. These prodrugs are activated by the reaction with reactive oxygen species (ROS) forming ferrocenium‐based DLCs. Since ROS are overproduced in cancer, the high‐efficiency cancer‐cell‐specific targeting of mitochondria could be achieved as demonstrated by fluorescence microscopy in combination with two fluorogenic pro‐DLCs in vitro and in vivo. We prepared a conjugate of another pro‐DLC with a clinically approved drug carboplatin and confirmed that its accumulation in mitochondria was higher than that of the free drug. This was reflected in the substantially higher anticancer effect of the conjugate.

ROS-Responsive N-Alkylaminoferrocenes for Cancer-Cell-Specific Targeting of Mitochondria.

Mitochondrial membrane potential is more negative in cancer cells than in normal cells, allowing cancer targeting by delocalized lipophilic cations (DLCs). However, as the difference is rather small, these drugs affect also normal cells. Now a concept of pro-DLCs is proposed based on an N-alkylaminoferrocene structure. These prodrugs are activated by the reaction with reactive oxygen species (ROS) forming ferrocenium-based DLCs. Since ROS are overproduced in cancer, the high-efficiency cancer-cell-specific targeting of mitochondria could be achieved as demonstrated by fluorescence microscopy in combination with two fluorogenic pro-DLCs in vitro and in vivo. We prepared a conjugate of another pro-DLC with a clinically approved drug carboplatin and confirmed that its accumulation in mitochondria was higher than that of the free drug. This was reflected in the substantially higher anticancer effect of the conjugate.

Photo-activation of the delocalized lipophilic cation D112 potentiates cancer selective ROS production and apoptosis

Delocalized lipophilic cations (DLCs) selectively accumulate in cancer cell mitochondria and have long been explored for therapeutic applications. Although targeted effects to cancer cells are demonstrated in vitro, non-specific toxicities in vivo have hampered clinical development. Identifying the molecular mechanisms of action and enhancing selectivity are thus necessary next steps to improve these compounds and evaluate their suitability for further drug development. D112 is one such DLC with promising properties. We previously demonstrated that D112 selectively induced intrinsic apoptosis in transformed versus non-transformed cell lines. Here we show that D112 preferentially entered transformed cells where it interacted with, and damaged mitochondrial DNA, inhibited Complex I respiration and induced reactive oxygen species (ROS). ROS production was critical for Bax activation and subsequent apoptosis. Importantly, photo-activation of D112 potentiated selective ROS production and increased the window of toxicity towards cancer cells over non-transformed cells. Thus photodynamic therapy would be an exciting adjunct to D112 studies and may be generally applicable for other DLCs that are currently under therapeutic investigation.

Boron-containing delocalised lipophilic cations for the selective targeting of cancer cells.

To limit the incidence of relapse, cancer treatments must not promote the emergence of drug resistance in tumour and cancer stem cells. Under the proviso that a therapeutic amount of boron is selectively delivered to cancer cells, Boron Neutron Capture Therapy (BNCT) may represent one approach that meets this requirement. To this end, we report the synthesis and pharmacology of several chemical entities, based on boron-rich carborane moieties that are functionalised with Delocalized Lipophilic Cations (DLCs), which selectively target the mitochondria of tumour cells. The treatment of tumour and cancer stem cells (CSCs) with such DLC-functionalized carboranes (DLC-carboranes) induces cell growth arrest that is both highly cancer-cell-selective and permanent. Experiments involving cultures of normal and cancer cells show that only normal cells exhibit recapitulation of their proliferation potential upon removal of the DLC-carborane treatment. At the molecular level, the pharmacological effect of DLC-carboranes is exerted through activation of the p53/p21 axis.

Membrane permeable lipophilic cations as mitochondrial directing groups.

The mitochondrion's negatively charged membrane potential has been well documented to drive the accumulation of membrane permeable delocalized lipophilic cations (DLC). DLC attachments to known bioactive compounds can direct organelle localization and improve drug exposure to targets within the mitochondria. Due to the mitochondria's essential function and its regulation of cell death, DLC targeted therapies are the focus of drug discovery projects altering cellular fate via mitochondrial targets. This review provides an update on recent developments for the two main applications of DLCs: cytoprotective therapies aimed at reducing oxidative stress and cytotoxic therapies aimed at initiating cell death for the treatment of various cancers. Both approaches have produced significant improvements using DLC conjugated compounds that include improved potency, pharmacokinetic properties, and the potential to overcome resistance mechanisms.

Abstract 1603: Synthesis and in vitro evaluation of lipophilic cation conjugated photosensitizers for targeting mitochondria.

Abstract Mitochondria-specific photosensitizers were designed by taking advantage of the preferential localization of delocalized lipophilic cations (DLCs) in mitochondria. Three conjugates, CMP-Rh, CMP-tPP, and CMP-(tPP)2, were successfully synthesized by linking the hydroxy core modified porphyrins (CMP-OH and CMP-(OH)2) to either rhodamine B (Rh B) or one or two triphenyl phosphine (tPP) cations respectively via a saturated hydrocarbon linker. Their ability for delivering photosensitizers to mitochondria was evaluated using dual staining fluorescence microscopy and staining pattern was compared with a mitochondrial specific probe. Fluorescence imaging study suggested that CMP-Rh specifically localized in mitochondria. On the other hand, CMP-tPP and CMP-(tPP)2 showed less significant mitochondrial localization. The conjugation of CMP to Rh and tPP did greatly improve intracellular uptake and in vitro photodynamic activity compared to CMP-OH. In addition, to evaluate the efficiency of the conjugates as photosensitizers, their photophysical properties and in vitro biological activities were studied in comparison to those of CMP-OH. All conjugates were capable of generating singlet oxygen at rates comparable to CMP-OH. This improved photodynamic activity might be primarily due to an enhanced cellular uptake. Our study suggests that Rh B cationic group is better at least for CMP than tPP as a mitochondrial targeting vector. Citation Format: Pallavi Rajaputra, Gregory Nkepang, Ryan Watley, Youngjae You. Synthesis and in vitro evaluation of lipophilic cation conjugated photosensitizers for targeting mitochondria. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1603. doi:10.1158/1538-7445.AM2013-1603

Synthesis and in vitro biological evaluation of lipophilic cation conjugated photosensitizers for targeting mitochondria

Mitochondria-specific photosensitizers were designed by taking advantage of the preferential localization of delocalized lipophilic cations (DLCs) in mitochondria. Three DLC-porphyrin conjugates: CMP-Rh (a core modified porphyrin-rhodamine B cation), CMP-tPP (a core modified porphyrin-mono-triphenyl phosphonium cation), CMP-(tPP)2 (a core modified porphyrin-di-tPP cation) were prepared. The conjugates were synthesized by conjugating a monohydroxy core modified porphyrin (CMP-OH) to rhodamine B (Rh B), or either one or two tPPs, respectively, via a saturated hydrocarbon linker. Their ability for delivering photosensitizers to mitochondria was evaluated using dual staining fluorescence microscopy. In addition, to evaluate the efficiency of the conjugates as photosensitizers, their photophysical properties and in vitro biological activities were studied in comparison to those of CMP-OH. Fluorescence imaging study suggested that CMP-Rh specifically localized in mitochondria. On the other hand, CMP-tPP and CMP-(tPP)2 showed less significant mitochondrial localization. All conjugates were capable of generating singlet oxygen at rates comparable to CMP-OH. Interestingly, all cationic conjugates showed dramatic increase in cellular uptake and phototoxicity compared to CMP-OH. This improved photodynamic activity might be primarily due to an enhanced cellular uptake. Our study suggests that Rh B cationic group is better at least for CMP than tPP as a mitochondrial targeting vector.

Synthesis of F16 conjugated with 5‐fluorouracil and biophysical investigation of its interaction with bovine serum albumin by a spectroscopic and molecular modeling approach

5-Fluorouracil (5-FU) has been widely used as a chemotherapy agent in the treatment of many types of solid tumors. Investigation of its antimetabolites led to the development of an entire class of fluorinated pyrimidines. However, the toxicity profile associated with 5-FU is significant and includes diarrhea, mucositis, hand-foot syndrome and myelosuppression. In aiming at reducing of the side effects of 5-FU, we have designed and synthesized delocalized lipophilic cations (DLCs) as a vehicle for the delivery of 5-FU. DLCs accumulate selectively in the mitochondria of cancer cells because of the high mitochondrial transmembrane potential (ΔΨm). Many DLCs exhibited anti-cancer efficacy and were explored as potential anti-cancer drugs based on their selective accumulation in the mitochondria of cancer cells. F16, the DLC we used as a vehicle, is a small molecule that selectively inhibits tumor cell growth and dissipates mitochondrial membrane potential. The binding of the conjugate F16-5-FU to bovine serum albumin (BSA) was investigated using spectroscopic and molecular modeling approaches. Fluorescence quenching constants were determined using the Stern-Volmer equation to provide a measure of the binding affinity between F16-5-FU and BSA. The activation energy of the interaction between F16-5-FU and BSA was calculated and the unusually high value was discussed in terms of the special structural block indicated by the molecular modeling approach. Molecular modeling showed that F16-5-FU binds to human serum albumin in site II, which is consistent with the results of site-competitive replacement experiments. It is suggested that hydrophobic and polar forces played important roles in the binding reaction, in accordance with the results of thermodynamic experiments.

Abstract 3466: Mitochondrial targeting photosensitizer-lipophilic cation conjugates for photodynamic therapy

Abstract Mitochondrion of a living cell plays a pivotal role in apoptosis. Targeting the mitochondria of cancer cell for the induction of apoptosis in photodynamic therapy (PDT) has gained importance. Cancer cells have higher plasma and mitochondrial membrane potential compared to normal cells, consequently delocalized lipophilic cations (DLCs) tend to accumulate more in them than in normal cells. In PDT photosensitizers can be specifically delivered to the mitochondria of cancerous cell by conjugating them with DLCs. Three conjugates: CMP-TPP (a porphyrin-triphenyl phosphene monocation conjugate) and CMP-(TPP)2 (a porphyrin- triphenyl phosphene dication conjugate), CMP-Rh (a porphyrin -rhodamine monocation conjugate) were synthesized as mitochondria targeting photosensitizers. The conjugates were synthesized by conjugating a core modified porphyrin (CMP-TPP) to either 1 or 2 triphenyl phosphene (TPP) molecules or rhodamine B (Rh B) respectively via a saturated hydrocarbon linker. To evaluate the efficiency of the conjugates, the photophysical properties like fluorescence resonance energy transfer (FRET in CMP-Rh) and singlet oxygen generation was compared to that of the unconjugated CMP. Potent in vitro photodynamic activity was observed in Colon 26 cells upon treatment with CMP conjugates and exposure to 690 nm diode laser light source (5.6 mW/cm2 for 30 min, 10.8 J/cm2). No significant darktotoxicity was observed in cells treated up to 5 μM of the conjugates. The conjugates showed approximately 12-14 times higher uptake in Colon 26 cells compared to unconjugated porphyrin. Further, sub-cellular localization, comparison of preferential uptake by cancerous cells versus normal cells, in vivo bio-distribution, PDT in BALB/c mice will be discussed. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3466. doi:1538-7445.AM2012-3466

Development of Novel Peptides for Mitochondrial Drug Delivery: Amino Acids Featuring Delocalized Lipophilic Cations

To create a new class of mitochondria-penetrating peptides (MPPs) that would facilitate drug delivery into the organelle through the inclusion of delocalized lipophilic cations (DLCs) in the peptide sequence. We synthesized two novel amino acids featuring DLCs and incorporated them into peptides. Systematic studies were conducted to compare peptides containing these residues to those with natural cationic amino acids. Diastereomers were compared to determine the most advantageous arrangement for these peptides. Peptide lipophilicity, cellular uptake and mitochondrial specificity were compared for a variety of peptides. Synthetic DLC residues were found to increase mitochondrial localization of MPPs due to higher overall hydrophobicity. MPP stereochemistry was important for cellular uptake rather than subcellular localization. This study reaffirmed the importance of uniform overall charge distribution for mitochondrial specificity. DLCs can be incorporated into synthetic peptides and facilitate mitochondrial drug delivery. Lipophilicity and charge distribution must be carefully balanced to ensure localization within mitochondria.

Delocalized lipophilic cations as a new therapeutic approach in cancer

Delocalized lipophilic cations (DLCs) penetrate plasma and mitochondrial membranes and accumulate in mitochondria. The higher mitochondrial membrane potentials of neoplastic vs normal cells, in general, account for greater uptake and may be a way to selectively target these cells since. Dequalinium (DQA) is a DLC and so our goal is to evaluate the therapeutic potential of DQA in cancer, namely in B-cell Chronic Lymphocytic Leukaemia (B-CLL), Acute Promyelocytic Leukaemia (APL) and Hepatocellular Carcinoma (HCC). For this we used 3 cell lines, EHEB (B-CLL), HL-60 (APL) and HUH-7 (HCC), to evaluate the effect of different concentrations of DQA either by single dose administration, by daily dose administration and by association with conventional anticarcinogenic agents. Cell viability and death was determined by the resazurin assay, optical microscopy and by flow cytometry. The latter was also used to evaluate the mitochondrial membrane potential, the levels of ROS (H2O2; O2•-) and the antioxidant defense, Reduced Glutathione (GSH), using fluorescent probes. We found that DQA induced a decrease in cell viability inducing cell death by late apoptosis/necrosis in a time, dose and cell type dependent manner, with and IC50 of 2.5, 4.7 and 7.5 μM at 48h of exposure, respectively to HL-60, HUH-7 and EHEB. These effects may be mediated by oxidative stress as we have observed and increase in ROS production and a decrease in GSH levels and in mitochondrial membrane potential. We also observed that if DQA is administered on a daily basis a much lower concentration is required to induce the same effect. On the other hand, the association of DQA with the conventional drug induces a synergistic effect, because lower concentration of both drugs is required to obtain the some effect.

From delocalized lipophilic cations to hypoxia: blocking tumor cell mitochondrial function leads to therapeutic gain with glycolytic inhibitors.

An unexpected similarity between cancer and cardiac muscle cells in their sensitivity to anthracyclines and delocalized lipophilic cations (DLC) prompted a series of studies in which it was shown that the positive charge of these compounds is central to their selective accumulation and toxicity in these two distinct cell types. An initial finding to explain this phenomenon was that cancer and cardiac muscle cells exhibit high negative plasma membrane potentials resulting in increased uptake of these agents. However, the p-glycoprotein efflux pump was shown to be another factor underlying differential accumulation of these compounds, since it recognizes positively charged drugs and thereby actively reduces their intracellular concentrations. The delocalized positive charge and lipophilicity of DLCs leads to their retention and inhibition of ATP synthesis in mitochondria. Years later it was realized that cancer cells in the hypoxic portions of solid tumors were similar to those treated with DLCs in relying mainly on anaerobic metabolism for survival and could thus be targeted with a glycolytic inhibitor, 2-deoxy-D-glucose (2-DG). This hypothesis has lead to a Phase I clinical trial in which 2-DG is used to selectively kill the hypoxic tumor cell population which are resistant to standard chemotherapy or radiation.

The selective in vitro cytotoxicity of carcinoma cells by AZT is enhanced by concurrent treatment with delocalized lipophilic cations.

This study assessed the selective growth inhibitory effect on cultured carcinoma cells by 3'-azido-3'-deoxythymidine (AZT), as a single agent, and in combination with delocalized lipophilic cations (DLCs) that are known to inhibit mitochondrial function. In cytotoxicity assays, treatment of cells with varying concentrations of AZT induced a dose-dependent inhibition of cell growth of the human carcinoma lines DU-145 (prostate; IC50 at 24 microM), MCF-7 (breast; IC50 at 22 microM), and CX-1 (colon; IC50 at 23 microM), yet caused no significant effect on the growth of the control epithelial cell line CV-1 (monkey kidney) at a concentration as high as 50 microM AZT. Combination treatment employing a constant concentration (1.25 microM) of the DLC dequalinium chloride (DECA) plus varying concentrations of AZT (0-50 microM) enhanced the AZT-induced cytotoxicity of carcinoma cells at least fourfold for MCF-7 and CX-1 cells (IC50 at 5 microM AZT), and twofold for DU-145 cells (IC50 at 11 microM AZT). Similar results were obtained in DU-145 cells using a constant concentration of the DLC MKT-077 (1.0 microM) and varying concentrations of AZT (IC50 at 12.5 microM). As expected, the drug combination of constant DLC and varying AZT had no significant effect on the growth of CV-1 cells. Clonogenic assays demonstrated up to 20-fold enhancement of selective carcinoma cell killing by combination vs. single agent treatment, depending on the specific drug combination and concentrations used. It is hypothesized that the efficacy of the AZT/DLC drug combination in carcinoma cell killing may be based on a dual selectivity involving inhibition of mitochondrial energy metabolism and inhibition of DNA synthesis due to limited deoxythymidine monophosphate availability.

Delocalized lipophilic cations selectively target the mitochondria of carcinoma cells

Traditional chemotherapies, aimed at DNA replication in rapidly dividing cells, have achieved only limited success in the treatment of carcinomas due largely to their lack of specificity for cells of tumorigenic origin. It is important, therefore, to investigate treatment strategies aimed at novel cellular targets that are sufficiently different between normal cells and cancer cells so as to provide a basis for selective tumor cell killing. Delocalized lipophilic cations (DLCs) are concentrated by cells and into mitochondria in response to negative inside transmembrane potentials. The higher plasma and/or mitochondrial membrane potentials of carcinoma cells compared to normal epithelial cells account for the selective accumulation of DLCs in carcinoma mitochondria. Since most DLCs are toxic to mitochondria at high concentrations, their selective accumulation in carcinoma mitochondria and consequent mitochondrial toxicity provide a basis for selective carcinoma cell killing. Several of these compounds have already displayed some degree of efficacy as chemotherapeutic agents in vitro and in vivo. The effectiveness of DLCs can also be enhanced by their use in photochemotherapy or combination drug therapy. Discovery of the biochemical differences that account for the higher membrane potentials in carcinoma cells is expected to lead to the design of new DLCs targeted specifically to those differences, resulting in even greater selectivity and efficacy for tumor cell killing.

Micellar Delivery System for Dequalinium—A Lipophilic Cationic Drug with Anticarcinoma Activity

Delocalized lipophilic cations (DLC) such as dequalinium (DQA) comprise a novel class of antitumor agents. They are more selectively toxic to carcinoma cells in comparison to non-transformed epithelial cells. In addition, DLC’s have also been shown to enhance the efficacy of existing anticancer modalities such as radiation and photodynamic therapies. A severe drawback of these attractive novel anticancer agents is their limited solubility in aqueous solutions. To overcome this obstacle, we tried to incorporate DQA into liposomes and micelles. We found that incorporation in liposomes does not seem to be possible at physiological salt concentrations. However, we succeeded in preparing micelles made of polyethylene glycol derivatives of phosphatidy-lethanolamine (PEG-PE) which stably bind up to 30 mol % DQA.

Synthesis and evaluation of novel rhodacyanine dyes that exhibit antitumor activity.

Rhodacyanine dyes and several analogous delocalized lipophilic cations (DLCs) were synthesized and evaluated as novel antitumor agents. Rhodacyanine dye consists of two heteroaromatic rings such as thiazoles at both termini of the conjugate systems and 4-oxothiazolidine (rhodanine) in the middle of it. Compounds with such a unique double-conjugate structure were found to inhibit the growth of several tumor cell lines, such as colon carcinoma CX-1, and to exhibit relatively low toxicity against normal kidney cell line CV-1 (e.g., IC50(CX-1) = 50 nM, IC50(CV-1) = 17.3 microM; selectivity index = 346 for compound 5). These compounds were also found to be efficacious in the tumor-bearing nude mice model (e.g., against human melanoma LOX; T/C (%) = 168 for compound 5). Structural modifications on rhodacyanine, including deletion of a heteroaromatic ring involved in the merocyanine conjugate system and replacement of rhodanine with a structurally related moiety such as 4-oxoimidazolidine or 4-oxo-1,3-dithiolane, resulted in a loss of the selectivity and/or the activity. Our current structure-activity studies imply that the double-conjugate system with a rhodanine moiety is essential for the selective activity of rhodacyanine dyes, and we find this class of compounds as unique antitumor agents candidates.