Synthesis and Evaluation of a Macrocyclic Actinium-225 Chelator,Quality Control and In Vivo Evaluation of 225Ac-crown-αMSH Peptide

Targeted alpha-therapy (TAT) has great potential for treating a broad range of late-stage cancers by delivering a focused and lethal radiation dose to tumors. Actinium-225 (²²⁵Ac) is an emerging alpha emitter suitable for TAT; however, the availability of chelators for Ac remains limited to a small number of examples (DOTA and macropa). Herein, we report a new Ac macrocyclic chelator named ‘ crown’ , which binds quantitatively and rapidly (<10 min) to Ac at ambient temperature. We synthesized ²²⁵Ac- crown -αMSH, a peptide targeting the melanocortin 1 receptor (MC1R), specifically expressed in primary and metastatic melanoma. Biodistribution of ²²⁵Ac- crown -αMSH showed favorable tumor-to-background ratios at 2 h post injection in a preclinical model. In addition, we demonstrated dramatically different biodistrubution patterns of ²²⁵Ac- crown -αMSH when subjected to different latency times before injection. A combined quality control methodology involving HPLC, gamma spectroscopy and radioTLC is recommended.     

超声激活血卟啉对B16小鼠黑色素瘤细胞杀伤作用的研究初报

采用MTT法检测,经超声、超声 血卟啉分别作用后细胞线粒体内琥珀酸脱氢酶的活性,研究超声激活血卟啉对体外培养的B16小鼠黑色素瘤细胞的杀伤作用。结果表明:1.6MHz超声对B16小鼠黑色素瘤细胞辐照5min可表现出明显的杀伤效应,而超声协同被激活的血卟啉对B16细胞的杀伤作用比单纯超声的杀伤效应更加明显,且有显著性差异。     

Fluorescence Studies of Melanin by Stepwise Two-Photon Femtosecond Laser Excitation

Fluorescence of synthetic melanin in the solvents H₂O, KOH, ethylene glycol monomethyl ether, and dimethyl sulfoxide has been excited by two-photon absorption at 800 nm, using 120-fs pulses with photon flux densities of 10²⁷ cm^–2.S^–1. Compared to the one-photon (400-nm)-induced fluorescence of melanin, the overall spectral shape is red-shifted and shows a strong environment sensitivity. The decay of the two-photon-induced fluorescence (TPF) of melanin is three-exponential, with a shortest main component of about 200 ps. The results of the TPF studies in line with the unique light absorption property of melanin of a monotonously decreasing absorption spectrum between the near UV-region and the near infrared region indicate that the TPF is realized via stepwise absorption of two 800-nm photons. In comparison to the simultaneous absorption of two photons, the stepwise process needs lower photon flux densities to get a sufficient population of the fluorescent level. This stepwise process offers new possibilities of selective excitation of melanin in skin tissue in a spectral region where there is no overlap with any absorption of another fluorescent tissue component. The first results with different samples of excised human skin tissue (healthy, nevus cell nevi, malignant melanoma) suggest that fluorescence excited in this way yields information on malignant transformation.     

Synthesis of <Emphasis Type="Italic">O</Emphasis>-(2-[<Superscript>18</Superscript>F]fluoroethyl)-<Emphasis Type="Italic">L</Emphasis>-tyrosine and its biological evaluation in B16 melanoma-bearing mice as PET tracer for tumor imaging

O-(2-[¹⁸F]fluoroethyl)-L-tyrosine ([¹⁸F]FET), a fluorine-18 labeled analogue of tyrosine, has been synthesized and biologically evaluated in tumor-bearing mice. The whole synthesis procedure is completed within 50 min. The radiochemical yield is about 40% (no decay corrected) and radiochemical purity more than 97% after simplified solid phase extraction. [¹⁸F]FET shows rapid, high uptake and long retention in the tumor as well as low uptake in the brain. The ratios of tumor-to-muscle (T/M) and tumor-to-blood (T/B) of [¹⁸F]FET are similar to those of [¹⁸F]FDG, but the ratios of tumor-to-brain (T/Br) are 2–3 times higher than that of [¹⁸F]FDG. Autoradiography of [¹⁸F]FET demonstrates a remarkable accumulation in melanoma with high contrast. It appears to be a probable competitive candidate for melanoma imaging with PET. Supported by the Knowledge Innovation Project of Chinese Academy of Sciences (No. KJCX1-SW-08) and the National Natural Science Foundation of China (Grant No. 30371634)     

Proliferation and Invasion of Melanoma Are Suppressed by a Plant Protease Inhibitor,Leading to Downregulation of Survival/Death-Related Proteins

Cell adhesion and migration are crucial for cancer progression and malignancy. Drugs available for the treatment of metastatic melanoma are expensive and unfit for certain patients. Therefore, there is still a need to identify new drugs that block tumor cell development. We investigated the effects of Enterolobium contortisiliquum trypsin inhibitor (EcTI), a protease inhibitor, on cell viability, cell migration, invasion, cell adhesion, and cell death (hallmarks of cancer) in vitro using human melanoma cells (SK-MEL-28 and CHL-1). Although EcTI did not affect non-tumor cells, it significantly inhibited the proliferation, migration, invasion, and adhesion of melanoma cells. Investigation of the underlying mechanisms revealed that EcTI triggered apoptosis and nuclear shrinkage, increased PI uptake, activated effector caspases-3/7, and produced reactive oxygen species (ROS). Furthermore, EcTI disrupted the mitochondrial membrane potential, altered calcium homeostasis, and modified proteins associated with survival and apoptosis/autophagy regulation. Acridine orange staining indicated acidic vesicular organelle formation upon EcTI treatment, demonstrating a cell death display. Electronic microscopy corroborated the apoptotic pattern by allowing the visualization of apoptotic bodies, mitochondrial cristae disorganization, and autophagic vesicles. Taken together, these results provide new insights into the anti-cancer properties of the natural EcTI protein, establishing it as a promising new therapeutic drug for use in melanoma treatment.     

Soapwort (Saponaria officinalis L.) Extract vs. Synthetic Surfactants—Effect on Skin-Mimetic Models

Our skin is continuously exposed to different amphiphilic substances capable of interaction with its lipids and proteins. We describe the effect of a saponin-rich soapwort extract and of four commonly employed synthetic surfactants: sodium lauryl sulfate (SLS), sodium laureth sulfate (SLES), ammonium lauryl sulfate (ALS), cocamidopropyl betaine (CAPB) on different human skin models. Two human skin cell lines were employed: normal keratinocytes (HaCaT) and human melanoma cells (A375). The liposomes consisting of a dipalmitoylphosphatidylcholine/cholesterol mixture in a molar ratio of 7:3, mimicking the cell membrane of keratinocytes and melanoma cells were employed as the second model. Using dynamic light scattering (DLS), the particle size distribution of liposomes was analyzed before and after contact with the tested (bio)surfactants. The results, supplemented by the protein solubilization tests (albumin denaturation test, zein test) and oil emulsification capacity (using olive oil and engine oil), showed that the soapwort extract affects the skin models to a clearly different extent than any of the tested synthetic surfactants. Its protein and lipid solubilizing potential are much smaller than for the three anionic surfactants (SLS, ALS, SLES). In terms of protein solubilization potential, the soapwort extract is comparable to CAPB, which, however, is much harsher to lipids.     

Inhibitory effects of schisantherin F from Schisandra propinqua subsp. sinensis on human melanoma A375 cells through ROS-induced mitochondrial dysfunction and mitochondria-mediated apoptosis

Abstract

Schisandra propinqua subsp. sinensis is a traditional medicinal plant used in Chinese folk medicine. Melanoma is the most dangerous form of skin cancer. To discover bioactive phytochemicals for preventing human melanoma, we have investigated the inhibitory effects of schisantherin F in Schisandra propinqua subsp. sinensis on human melanoma A375 cells and relevant mechanisms. The results showed that schisantherin F can inhibit A375 cells through inducing apoptosis. Further investigations have demonstrated schisantherin F attenuated the overproduction of ROS, depolarization of MMP, and mPTP opening. Meanwhile, schisantherin F inhibited the activity of Caspase-3 and up-stream Caspase-9, down-regulated Bcl-2 and up-regulated Bax. These findings propose the inhibitory mechanisms of schisantherin F in A375 cells include induction of mitochondrial dysfunction and mitochondria-mediated apoptosis.     

Confocal Raman microspectroscopy discriminates live human metastatic melanoma and skin fibroblast cells

Confocal Raman microspectroscopy (CRM) continues to develop as a promising technique with possible clinical applications for the diagnosis and treatment of skin cancers. CRM studies of single cells can provide information on the biochemical content of cancer cells in situ, potentially providing new biochemical signatures or markers of cancer cells. Here, we report a CRM study of single, living human metastatic melanoma cells (SK‐Mel‐2) and normal skin fibroblast cells (BJ) cultured and examined under identical experimental conditions. A total of almost 1200 Raman spectra were measured from more than 120 BJ and SK‐Mel‐2 cells using an inverted microscope with 647 nm laser excitation. Raman spectra were measured from within three distinct intracellular regions of the cells – cytoplasm, nucleoplasm, and nucleolus. When Raman spectra from each cell type were compared using principal components analysis (PCA) and linear discriminant analysis with leave‐one‐dish‐out cross‐validation (LDA‐CV), the two cell types were discriminated with 93% (cytoplasm), 98% (nucleolus), and 96% (nucleoplasm) accuracy. The main biochemical differences identified between the two cell types were higher RNA levels in the nucleoli of BJ cells and high amounts of lipid and collagen in the cytoplasm of SK‐Mel‐2 cells. For both cell types, higher levels of RNA were detected in the nucleoli versus the nucleoplasm. PCA with LDA‐CV was 98% (cytoplasm), 93% (nucleoplasm), and 73% (nucleolus) accurate in identifying the intracellular region based on the Raman spectra from both cell types. No significant trend was observed when the data were analyzed with respect to cell passage number. Thus, CRM with PCA and LDA‐CV successfully discriminated two skin cancer‐relevant cell lines while detecting different amounts of nucleic acids, lipids, and proteins in distinct intracellular regions, further underscoring its potential as a clinical diagnostic tool. Copyright © 2013 John Wiley & Sons, Ltd.     

A Self-Assembled ATP Probe for Melanoma Cell Imaging

In this investigation, a new terpyridine metal complex was developed as a probe for selective detection of ATP and imaging of melanoma cells. The probe takes advantage of the ability of the metal complex to be transformed to its imaging competent turn-on state through assembly with ATP.