CCRF-CEM Cancer Cell Research Articles

Novel triterpenoid pyrones, phthalimides and phthalates are selectively cytotoxic in CCRF-CEM cancer cells – Synthesis, potency, and mitochondrial mechanism of action

A series of triterpenoid pyrones was synthesized and subsequently modified to introduce phthalimide or phthalate moieties into the triterpenoid skeleton. These compounds underwent in vitro cytotoxicity screening, revealing that a subset of six compounds exhibited potent activity, with IC50 values in the low micromolar range. Further biological evaluations, including Annexin V and propidium iodide staining experiment revealed, that all compounds induce selective apoptosis in cancer cells. Measurements of mitochondrial potential, cell cycle analysis, and the expression of pro- and anti-apoptotic proteins confirmed, that apoptosis was mediated via the mitochondrial pathway. These findings were further supported by cell cycle modulation and DNA/RNA synthesis studies, which indicated a significant increase in cell accumulation in the G0/G1 phase and a marked reduction in S-phase cells, alongside a substantial inhibition of DNA synthesis. The activation of caspase-3 and the cleavage of PARP, coupled with a decrease in the expression of Bcl-2 and Bcl-XL proteins, underscored the induction of apoptosis through the mitochondrial pathway. Given their high activity and pronounced effect on mitochondria function, trifluoromethyl pyrones 1f and 2f, and dihydrophthalimide 2h have been selected for further development.

Synthesis and evaluation of cytotoxic and antimicrobial activity of some 3-aryl-6-phenyl-7H-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazines

3-aryl-6-phenyl-7H-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazines were obtained with good yields (81-95%) via the heterocyclization reaction of 1-phenyl-2-((5-aryl-1,3,4-oxadiazole-2-yl)thio)ethane-1-ones in acetic acid. The physicochemical characteristics of the synthesized compounds were established, the structures were confirmed by the data of IR, 1H and 13C NMR spectra, as well as the results of X-ray diffraction analysis. The cytotoxic, antibacterial and antifungal properties of these compounds were evaluated. In vitro screening results showed that compounds 8, 9 and 12 significantly inhibit (54-65%) the growth of HeLa, HBL-100 and CCRF-CEM cancer cell lines. It was found that the cytotoxicity of the synthesized compounds increases in the series of oxadiazoltiones (1-4) - S-derivatives (5-8) - triazolothiadiazines (9-12). Compounds 5-16 do not exhibit antimicrobial properties.

Design, synthesis, and anticancer evaluation of 1-benzo[1,3]dioxol-5-yl-3-N-fused heteroaryl indoles.

A series of 1-benzo[1,3]dioxol-5-yl-indoles bearing 3-N-fused heteroaryl moieties have been designed based on literature reports of the activity of indoles against various cancer cell lines, synthesized via a Pd-catalyzed C-N cross-coupling, and evaluated for their anticancer activity against prostate (LNCaP), pancreatic (MIA PaCa-2), and acute lymphoblastic leukemia (CCRF-CEM) cancer cell lines. A detailed structure-activity relationship study culminated in the identification of 3-N-benzo[1,2,5]oxadiazole 17 and 3-N-2-methylquinoline 20, whose IC50 values ranged from 328 to 644nM against CCRF-CEM and MIA PaCa-2. Further mechanistic studies revealed that 20 caused cell cycle arrest at the S phase and induced apoptosis in CCRF-CEM cancer cells. These 1-benzo[1,3]dioxol-5-yl-3-N-fused heteroaryl indoles may serve as a template for further optimization to afford more active analogs and develop a comprehensive understanding of the structure-activity relationships of indole anticancer molecules.

An Evaluation of the Novel Biological Properties of Diterpenes Isolated from Plectranthus ornatus Codd. In Vitro and In Silico.

Plectranthus ornatus Codd, the genus Plectranthus of the Lamiaceae family, has been used as traditional medicine in Africa, India and Australia. Pharmacological studies show the use of this plant to treat digestive problems. In turn, leaves were used for their antibiotic properties in some regions of Brazil to treat skin infections. The present study examines the anti-inflammatory, antioxidant and cytotoxic effects of the halimane and labdane diterpenes (11R*,13E)-11-acetoxyhalima-5,13-dien-15-oic acid (HAL) and 1α,6β-diacetoxy-8α,13R*-epoxy-14-labden-11-one (PLEC) and the forskolin-like 1:1 mixture of 1,6-di-O-acetylforskolin and 1,6-di-O-acetyl-9-deoxyforskolin (MRC) isolated from P. ornatus on lung (A549) and leukemia (CCRF-CEM) cancer cell lines, and on normal human retinal pigment epithelial (ARPE-19) cell line in vitro. Additionally, molecular docking and computational approaches were used. ADMET properties were analysed through SwissADME and proTox-II—Prediction. The results indicate that all tested compounds significantly reduced the viability of the cancer cells and demonstrated no cytotoxic effects against the non-neoplastic cell line. The apoptosis indicators showed increased ROS levels for both the tested A549 and CCRF-CEM cancer cell lines after treatment. Furthermore, computational studies found HAL to exhibit moderate antioxidant activity. In addition, selected compounds changed mitochondrial membrane potential (MMP), and increased DNA damage and mitochondrial copy number for the CCRF-CEM cancer cell line; they also demonstrated anti-inflammatory effects on the ARPE-19 normal cell line upon lipopolysaccharide (LPS) treatment, which was associated with the modulation of IL-6, IL-8, TNF-α and GM-CSF genes expression. Docking studies gave indication about the lowest binding energy for 1,6-di-O-acetylforskolin docked into IL-6, TNF-α and GM-CSF, and 1,6-di-O-acetyl-9-deoxyforskolin docked into IL-8. The ADMET studies showed drug-likeness properties for the studied compounds. Thus, halimane and labdane diterpenes isolated from P. ornatus appear to offer biological potential; however, further research is necessary to understand their interactions and beneficial properties.

Novel 7-Chloro-(4-thioalkylquinoline) Derivatives: Synthesis and Antiproliferative Activity through Inducing Apoptosis and DNA/RNA Damage.

A series of 78 synthetic 7-chloro-(4-thioalkylquinoline) derivatives were investigated for cytotoxic activity against eight human cancer as well as 4 non-tumor cell lines. The results showed, with some exceptions, that sulfanyl 5–40 and sulfinyl 41–62 derivatives exhibited lower cytotoxicity for cancer cell lines than those of well-described sulfonyl N-oxide derivatives 63–82. As for compound 81, the most pronounced selectivity (compared against BJ and MRC-5 cells) was observed for human cancer cells from HCT116 (human colorectal cancer with wild-type p53) and HCT116p53−/− (human colorectal cancer with deleted p53), as well as leukemia cell lines (CCRF-CEM, CEM-DNR, K562, and K562-TAX), lung (A549), and osteosarcoma cells (U2OS). A good selectivity was also detected for compounds 73 and 74 for leukemic and colorectal (with and without p53 deletion) cancer cells (compared to MRC-5). At higher concentrations (5 × IC50) against the CCRF-CEM cancer cell line, we observe the accumulation of the cells in the G0/G1 cell phase, inhibition of DNA and RNA synthesis, and induction of apoptosis. In addition, X-ray data for compound 15 is being reported. These results provide useful scientific data for the development of 4-thioalkylquinoline derivatives as a new class of anticancer candidates.

Six New Diterpene Glycosides from the Soft Coral Lemnaliabournei.

A chemical study on the extracts of soft coral Lemnalia bournei resulted in the isolation and identification of six new bicyclic diterpene glycosides including three new lemnaboursides E–G (1–3), and three new lemnadiolboursides A–C (4–6), along with three known lemnaboursides (7–9). Their structures were elucidated by detailed spectroscopic analysis, ECD analysis, chemical methods, and comparison with the literature data. Lemnadiolboursides A–C (4–6) contained a lemnal-1(10)-ene-7,12-diol moiety compared with the lemnaboursides. All these compounds were evaluated for antibacterial activity; cell growth inhibition of A549, Hela, HepG2, and CCRF-CEM cancer cell lines; and inhibition of LPS-induced NO production in RAW264.7 macrophages. The results indicated that compounds 1, 2, and 4–6 exhibited antibacterial activity against Staphylococcus aureus and Bacillus subtilis (MIC 4–16 μg/mL); compounds 1–9 displayed low cytotoxicity on the CCRF-CEM cell lines (IC50 10.44–27.40 µM); and compounds 1, 2, and 5 showed weak inhibition against LPS-induced NO production (IC50 21.56–28.06 μM).

Antimicrobial Terpenoids from South China Sea Soft Coral Lemnalia sp.

Chemical investigation of the South China Sea soft coral Lemnalia sp. afforded 13 structurally diverse terpenoids, including three new neolemnane sesquiterpene lineolemnenes E–G (1–3); a new aristolane-type sesquiterpenoid, 2-acetoxy-aristolane (4); four new decalin-type bicyclic diterpenes, named biofloranates A−D (5−8); a new serrulatane, named euplexaurene D (9); and a new aromadendrane-type diterpenoid cneorubin K (10), together with three known related compounds (11−13). The structures of the new compounds were elucidated by NMR spectroscopy, the Mosher’s method, and ECD analysis. Compounds 1–13 were tested in a wide panel of biological assays. Lineolemnene J (3) showed weak cytotoxicity against the CCRF-CEM cancer cell line. The isolated new diterpenes, except euplexaurene D (9), demonstrated moderate antimicrobial activity against Bacillus subtilis and Staphylococcus aureus with a MIC of 4−64 μg/mL. Compound 2 exhibited a mild inhibitory effect against influenza A H1N1 virus (IC50 = 5.9 µM).

Platelet-Rich and Platelet-Poor Plasma Might Play Supportive Roles in Cancer Cell Culture: A Replacement for Fetal Bovine Serum?

Platelet-Rich (PRP) and Platelet-Poor plasma (PPP) are widely used in research and clinical platforms mainly due to their capacities to enhance cell growth. Although the short half-life (5 days) and the high price of platelet products pose challenges regarding their usage, they maintain the growth regulatory functions for weeks. Thus, we aimed to assess the supplementary values of these products in human CCRF- CEM cancer cells. Mechanistically, we also checked if the PRP/PPP treatment enhances YKL-40 expression as a known protein regulating cell growth. The PRP/PPP was prepared from healthy donors using manual stepwise centrifugation and phase separation. The viability of the cells treated with gradient PRP/PPP concentrations (2, 5, 10, and 15%) was measured by the MTT assay. The YKL-40 mRNA and protein levels were assessed using qRT-PCR and western blotting. The data were compared to FBS-treated cells. Our findings revealed that the cells treated by PRP/PPP not only were morphologically comparable to those treated by FBS but also showed greater viability at the concentrations of 10 and 15%. Moreover, it was shown that PRP/PPP induce cell culture support, at least in part, via inducing YKL-40 expression at both mRNA and protein levels in a time- and dose-dependent manner. Collectively, by showing cell culture support comparable to FBS, the PRP/PPP might be used as good candidates to supplement the cancer cell culture and overcome concerns regarding the use of FBS as a non-human source in human cancer research.

Antimicrobial and cytotoxic activity of (thio)alkyl hexopyranosides, nonionic glycolipid mimetics

A series of 19 synthetic alkyl and thioalkyl glycosides derived from d-mannose, d-glucose and d-galactose and having C10–C16 aglycone were investigated for cytotoxic activity against 7 human cancer and 2 non-tumor cell lines as well as for antimicrobial potential on 12 bacterial and yeast strains. The most potent compounds were found to be tetradecyl and hexadecyl β-d-galactopyranosides (18, 19), which showed the best cytotoxicity and therapeutic index against CCRF-CEM cancer cell line. Similar cytotoxic activity showed hexadecyl α-d-mannopyranoside (5) but it also inhibited non-tumor cell lines. Because these two galactosides (18, 19) were inactive against all tested bacteria and yeast strains, they could be a target-specific for eukaryotic cells. On the other hand, β-D-glucopyranosides with tetradecyl (11) and hexadecyl (12) aglycone inhibited only Gram-positive bacterial strain Enterococcus faecalis. The studied glycosides induce changes in the lipid bilayer thickness and lateral phase separation at high concentration, as derived from SAXS experiments on POPC model membranes. In general, glucosides and galactosides exhibit more specific properties. Those with longer aglycone show high cytotoxicity and therefore, they are more promising candidates for cancer cell line targeted inhibition.

Ferrocenylchalcone–uracil conjugates: synthesis and cytotoxic evaluation

Huisgen’s azide–alkyne cycloaddition reaction was employed to synthesize a series of 1H-1,2,3-triazole-tethered uracil–ferrocenyl chalcone conjugates with the aim of evaluating their in vitro anti-proliferative efficacy on human leukemia (CCRF-CEM) and human breast adenocarcinoma (MDA-MB-468) cell lines. Cytotoxic evaluation studies identified a number of synthesized conjugates that inhibited the proliferation of leukemia cancer cells by ~70% after 72 h. The selected synthesized conjugates were found to be significantly less cytotoxic against normal kidney cell line (LLC-PK1) when compared with CCRF-CEM cancer cells.

Flow injection amperometric sandwich-type aptasensor for the determination of human leukemic lymphoblast cancer cells using MWCNTs-Pdnano/PTCA/aptamer as labeled aptamer for the signal amplification

In this research, we demonstrated a flow injection amperometric sandwich-type aptasensor for the determination of human leukemic lymphoblasts (CCRF-CEM) based on poly(3,4-ethylenedioxythiophene) decorated with gold nanoparticles (PEDOT-Aunano) as a nano platform to immobilize thiolated sgc8c aptamer and multiwall carbon nanotubes decorated with palladium nanoparticles/3,4,9,10-perylene tetracarboxylic acid (MWCNTs-Pdnano/PTCA) to fabricate catalytic labeled aptamer. In the proposed sensing strategy, the CCRF-CEM cancer cells were sandwiched between immobilized sgc8c aptamer on PEDOT-Aunano modified surface electrode and catalytic labeled sgc8c aptamer (MWCNTs-Pdnano/PTCA/aptamer). After that, the concentration of CCRF-CEM cancer cells was determined in presence of 0.1 mM hydrogen peroxide (H2O2) as an electroactive component. The attached MWCNTs-Pdnano nanocomposites to CCRF-CEM cancer cells amplified the electrocatalytic reduction of H2O2 and improved the sensitivity of the sensor to CCRF-CEM cancer cells. The MWCNT-Pdnano nanocomposite was characterized with transmission electron microscopy (TEM) and energy dispersive X-ray (EDX). The electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) were used to confirm the stepwise changes in the electrochemical surface properties of the electrode. The proposed sandwich-type electrochemical aptasensor exhibited an excellent analytical performance for the detection of CCRF-CEM cancer cells ranging from 1.0 × 101 to 5.0 × 105 cells mL−1. The limit of detection was 8 cells mL−1. The proposed aptasensor showed high selectivity toward CCRF-CEM cancer cells. The proposed aptasensor was also applied to the determination of CCRF-CEM cancer cells in human serum samples.

Cytotoxic Lanostanoids from Poria cocos.

Six new and 16 known lanostanoids were isolated from the sclerotia of Poria cocos. The structures of the new isolates were elucidated to be 16α-hydroxy-3-oxo-24-methyllanosta-5,7,9(11),24(31)-tetraen-21-oic acid (1), 3β,16α,29-trihydroxy-24-methyllanosta-7,9(11),24(31)-trien-21-oic acid (2), 3β,16α,30-trihydroxy-24-methyllanosta-7,9(11),24(31)-trien-21-oic acid (3), 3β-acetoxy-16α,24β-dihydroxylanosta-7,9(11),25-trien-21-oic acid (4), 3β,16α-dihydroxy-7-oxo-24-methyllanosta-8,24(31)-dien-21-oic acid (5), and 3α,16α-dihydroxy-7-oxo-24-methyllanosta-8,24(31)-dien-21-oic acid (6), based on extensive spectroscopic analyses. The absolute configuration of 4 was determined using Mosher's method. The antiproliferative activity of the isolated compounds (except 3 and 4) was evaluated against four leukemic cell lines (Molt 4, CCRF-CEM, HL 60, and K562). Dehydropachymic acid (9), dehydroeburicoic acid (12), pachymic acid (14), and lanosta-7,9(11),24-trien-21-oic acid (20) exhibited an antiproliferative effect on the CCRF-CEM cancer cell line with IC50 values of 2.7, 6.3, 4.9, and 13.1 μM, respectively. Both dehydropachymic acid (9) and dehydroeburicoic acid (12) showed antiproliferative effects against Molt 4 (IC50 13.8 and 14.3 μM) and HL 60 (IC50 7.3 and 6.0 μM) leukemic cell lines. Primary computational analysis using a chemical global positioning system for natural products (ChemGPS-NP) on the active lanostanoids from P.cocos suggested that targets other than topoisomerases may be involved in the antiproliferative activity.

Anticancer and Antibacterial Activity Studies of Gold(I)-Alkynyl Chromones.

Three gold(I) complexes of alkynyl chromones were synthesized and characterized. The single-crystal X-ray structure analysis of a dinuclear compound and of a flavone derivative exhibit a typical d10 gold(I)-alkynyl linear arrangement. All complexes were evaluated as anticancer and antibacterial agents against four human cancer cell lines and four pathogenic bacterial strains. All compounds show antiproliferative activity at lower micromolar range concentrations. Complex 4 showed a broad activity profile, being more active than the reference drug auranofin against HepG2, MCF-7 and CCRF-CEM cancer cells. The cellular uptake into MCF-7 cells of the investigated complexes was measured by atomic absorption spectroscopy (AAS). These measurements showed a positive correlation between an increased cellular gold content and the incubation time of the complexes. Unexpectedly an opposite effect was observed for the most active compound. Biological assays revealed various molecular mechanisms for these compounds, comprising: (i) thioredoxin reductase (TrxR) inhibition, (ii) caspases-9 and -3 activation; (iii) DNA damaging activity and (iv) cell cycle disturbance. The gold(I) complexes were also bactericidal against Gram-positive methicillin-sensitive Staphylococcus aureus (MSSA) and methicillin-resistant S. aureus (MRSA) bacterial strains, while showing no activity against the Gram-negative Escherichia coli bacterial strain.

One-step synthesis of gradient gadolinium ironhexacyanoferrate nanoparticles: a new particle design easily combining MRI contrast and photothermal therapy.

A one-step synthesis of Prussian blue nanoparticles possessing a concentration gradient of Gd3+ counterions, g-Gd-PB, has been developed, and the potential for the particles to perform as both MRI positive contrast agents and photothermal therapy agents is demonstrated. The synthesis of potassium/gadolinium ironhexacyanoferrate is performed under increasing concentration of Gd3+ ions forming particles with a higher concentration of gadolinium toward the outer layers. The proton relaxivity (r1) measured for the particles is 12.3 mM(-1) s(-1), and T1 weighted images of phantoms containing the particles show their potential as MRI contrast agents. In addition, the Prussian blue host can rapidly and efficiently convert energy from near-IR light into thermal energy, allowing g-Gd-PB to be used as a photothermal therapy agent. The photothermal properties are demonstrated by measuring temperature changes of particle suspensions under irradiation and by photothermal ablation of CCRF-CEM cancer cells.

Aptamer-functionalized Near-infrared Quantum Dots Combined with Flow Cytometry for Rapid Detection of Leukemia Cells†

As a haematological malignancy,leukemia poses a great threat to human health and life. Its early and rapid diagnosis is crucial for the improvement of the cure and survival rate of patients. In this paper,a novel leukemia cell assay strategy has been proposed based on aptamer-functionalized quantum dots( QDs)combined with flow cytometry. In this strategy,a biotin-labeled cancer-specific aptamer was adopted as the recognition molecule,and avidin-modified QDs with near-infrared fluorescence emission were utilized as the signal generator. Through the "biotin-avidin"interaction,QDs could be functionalized with aptamers to construct a novel aptamer-QDs fluorescent nano-probe. This probe could specifically bind to target cell surface via the interaction between aptamers and receptors on cell membrane,thus indicating the presence of the target after analysis with a flow cytometer. As proof of concept,the detection of human acute lymphoblastic leukemia CCRF-CEM cells was performed using the specific aptamer,Sgc8 c,as a demonstration. Results showed that the modification with Sgc8 c did not markedly influence the fluorescence emission and size of QDs. With a simple incubation with cell samples for just 30 min,this Sgc8c-QDs nano-probe could successfully achieve the highly selective detection of CCRF-CEM cancer cells both in buffer and in serum. By comparison with the traditional fluorescent dye labeling method,this Sgc8c-QDs-based strategy exhibited a substantial enhancement in analysis sensitivity for CCRF-CEM cells in buffer,which realized about 4. 3 folds signal-to-background ratio of the FAM-labeled Sgc8c( FAM-Sgc8c) strategy. In particular,when used for serum sample analysis,the Sgc8c-QDs nano-probe still reserved a perfect applicability and displayed a relatively high signal-to-background ratio of about 9,while FAM-Sgc8 c nearly lost the detection efficiency at the same concentration. It has been clearly verified that this aptamer-QDs strategy is facile,fast,washing-free,specific and sensitive,which might hold a great potential as a versatile technique for diagnosis and prognosis applications in cancer researches.

Inhibition Effect of Mahkota Dewa (Phaleria macrocarpa) on Benzo(a)pyrene Induced Cytotoxicity in CCRF-CEM Cell Lines

Mahkota Dewa as a traditional plant has been commonly used as traditional cancer medication. However, the mechanism of USAge is not yet clear. The objective of this study is to know the mechanism of the protection effect of Mahkota Dewa on Benzo(a)pyrene (BaP) induced cytotoxicity in CCRF-CEM cell line. The result show BaP induced cell death with in CCRF-CEM cell line is dose-dependent but not based on time-course. Exposure of this cell for 24 h with variation of dose between 5-20 mM increased the percentage of apoptosis to about 15%. On the other hand, Mahkota Dewa itself has dose-dependently induced cytotoxicity and has no effect in the inhibition of BaP exposure. Phosphorylation of p38 MAPK in both BaP and Mahkota Dewa induced cytotoxicity has been seen but the involvement of oxidative stress is unclear. However, in other cancer cell line SH-SY5Y human neuroblastoma cells, the inhibition efffect of Mahkota Dewa in BaP exposure has been seen and no cytotoxicity effect appeared in this cell line. In conclusion, Mahkota Dewa has induced apoptosis in CCRF-CEM cancer cell line but not in SH-SY5Y cell line, so it has a potential anticancer effect; Mahkota Dewa, however, requires more researches on DNA level using other type of cancer to observe the mechanism.

A versatile activatable fluorescence probing platform for cancer cells in vitro and in vivo based on self-assembled aptamer/carbon nanotube ensembles.

Activatable aptamer probes (AAPs) have emerged as a promising strategy in cancer diagnostics, but existing AAPs remain problematic due to complex design and synthesis, instability in biofluids, or lack of versatility for both in vitro and in vivo applications. Herein, we proposed a novel AAP strategy for cancer cell probing based on fluorophore-labeled aptamer/single-walled carbon nanotube (F-apt/SWNT) ensembles. Through π-stacking interactions and proximity-induced energy transfer, F-apt/SWNT with quenched fluorescence spontaneously formed in its free state and realized signal activation upon targeting surface receptors of living cells. As a demonstration, Sgc8c aptamer was used for in vitro analysis and in vivo imaging of CCRF-CEM cancer cells. It was found that self-assembled Cy5-Sgc8c/SWNT held robust stability for biological applications, including good dispersity in different media and ultralow fluorescence background persistent for 2 h in serum. Flow cytometry assays revealed that Cy5-Sgc8c/SWNT was specifically activated by target cells with dramatic fluorescence elevation and showed improved sensitivity with as low as 12 CCRF-CEM cells detected in mixed samples containing ~100,000 nontarget cells. In vivo studies confirmed that specifically activated fluorescence was imaged in CCRF-CEM tumors, and compared to "always on" probes, Cy5-Sgc8c/SWNT greatly reduced background signals, thus resulting in contrast-enhanced imaging. The general applicability of the strategy was also testified by detecting Ramos cells with aptamer TD05. It was implied that F-apt/SWNT ensembles hold great potential as a simple, stable, sensitive, specific, and versatile activatable platform for both in vitro cancer cell detection and in vivo cancer imaging.

Label-Free and Turn-on Aptamer Strategy for Cancer Cells Detection Based on a DNA–Silver Nanocluster Fluorescence upon Recognition-Induced Hybridization

We present here a label-free and turn-on aptamer strategy for cancer cell detection based on the recognition-induced conformation alteration of aptamer and hybridization-induced fluorescence enhancement effect of DNA-silver nanoclusters (DNA-Ag NCs) in proximity of guanine-rich DNA sequences. In this strategy, two tailored DNA probes were involved. One is designed as a hairpin-shaped structure consisting of a target specific aptamer sequence at the 3'-end, a guanine-rich DNA sequence, and an arm segment at the 5'-end (denote as recognition probe). The other, serving as a signal probe, contains a sequence for Ag NCs templated synthesis and a link sequence complementary to the arm segment of the recognition probe. Recognizing and binding of the aptamer to cancer cells enforces the recognition probe to undergo a conformational alteration and then initiates hybridization between the arm segment of the recognition probe and the link sequence of the signal probe. The Ag NCs are then close to the guanine-rich DNA, leading to an enhanced fluorescence readout. As proof-of-concept, the CCRF-CEM cancer cell detection were performed by using the specific aptamer, sgc8c. It was demonstrated that this strategy could specially image the CCRF-CEM cells. Determination by flow cytometry allowed for detection of as low as 150 CCRF-CEM cells in 200 μL binding buffer. The general applicability of the strategy is also achieved in the successful detection of Ramos cells. These results implied that this strategy holds considerable potential for simple, sensitive, universal, and specific cancer cell detection with no required washing and separation steps.

Activatable aptamer probe for contrast-enhanced in vivo cancer imaging based on cell membrane protein-triggered conformation alteration

Aptamers have emerged as promising molecular probes for in vivo cancer imaging, but the reported "always-on" aptamer probes remain problematic because of high background and limited contrast. To address this problem, we designed an activatable aptamer probe (AAP) targeting membrane proteins of living cancer cells and achieved contrast-enhanced cancer visualization inside mice. The AAP displayed a quenched fluorescence in its free state and underwent a conformational alteration upon binding to target cancer cells with an activated fluorescence. As proof of concept, in vitro analysis and in vivo imaging of CCRF-CEM cancer cells were performed by using the specific aptamer, sgc8, as a demonstration. It was confirmed that the AAP could be specifically activated by target cancer cells with a dramatic fluorescence enhancement and exhibit improved sensitivity for CCRF-CEM cell analysis with the cell number of 118 detected in 200 μl binding buffer. In vivo studies demonstrated that activated fluorescence signals were obviously achieved in the CCRF-CEM tumor sites in mice. Compared to always-on aptamer probes, the AAP could substantially minimize the background signal originating from nontarget tissues, thus resulting in significantly enhanced image contrast and shortened diagnosis time to 15 min. Furthermore, because of the specific affinity of sgc8 to target cancer cells, the AAP also showed desirable specificity in differentiating CCRF-CEM tumors from Ramos tumors and nontumor areas. The design concept can be widely adapted to other cancer cell-specific aptamer probes for in vivo molecular imaging of cancer.

Cytotoxic activity of mammea type coumarins from Mammea siamensis flowers

Mammea siamensis (Miq.) T. Anderson (local name: sarapi) is a Thai medicinal plant in the family Clusiaceae and used in indigenous medicine as a heart tonic. The n-hexane fraction from M. siamensis flowers CH2Cl2-CH3OH (1:1) crude extract yielded coumarins of the mammea type: mammea A/AA, mammea A/AA cyclo D, mammea A/AB cyclo D, mammea A/AC cyclo D, deacetylmammea E/BA, and deacetylmammea E/BB. The isolated compounds were examined in an in-vitro XTT assay against the human MDA-MB-231 (breast adenocarcinoma), U-251 (central nervous system), HCT-116 (colon cancer), as well as the CCRF-CEM (leukemia) cancer cell lines. Only mammea A/AA and the mixture of deacetylmammea E/BA and deacetylmammea E/BB were found to possess significant cytotoxic activities, at 10µg/mL in CCRF-CEM with values of 78.1±0.8% and 95.9±1.1%; in MDA-MB-231 with values of 58.9±0.8% and 82.4±0.9%; in U-251 with values of 27.7±3.2% and 78.8±1.6%; and in HCT-116 with inhibition values of 73.5±4.9% and 97.6±0.6% in the four human cancer cell lines, respectively, comparable to vinblastine at 0.01µg/mL, with values of 44.2±8.5%, 51.5±12.9%, 71.0±2.5%, and 54.8±9.4%, respectively. Mammea A/AA and the mixture of deacetylmammea E/BA and deacetylmammea E/BB showed significant cytotoxic activities against the human MDA-MB-231, U-251, and HCT-116 as well as the CCRF-CEM cancer cell lines with IC50 values of 7.2±1.0, 5.2±1.0; nd, 19.1±1.0; 16.6±1.1, 6.5±1.0; and 20.9±1.4, 4.9±1.0µM, respectively.