A375 Human Melanoma Research Articles

Optimization of topical curcumin spanlastics for melanoma treatment

The aim of the study was to enhance curcumin skin permeability through the preparation of spanlastics. Spanlastics were prepared using the ethanol injection technique through a central composite design using Span 60 concentration (X1), edge activator type (X2), and its concentration (X3) as the independent variables. The spanlastics were characterized for particle size (PS), encapsulation efficiency (EE), and dissolution efficiency (%DE24h). Formulae with the highest desirability (FN1 and FN2) were prepared and further characterized. They were elastic, spherical, nonirritant, and compatible with the used excipients. They had particle sizes of 147 and 198 nm, encapsulation efficiencies of 84.00% and 89.63%, zeta potential values of −45.50 and −39.10 mV, permeation enhancement ratios of 11.51 and 8.34 folds, and amounts retained of 7.25 and 10.44 µg/cm2 after 24 h, respectively. Formulae FN1 and FN2 showed cytotoxic effects after 48 h on human melanoma A375 with IC50 of 10.9 and 75.6 μg/mL, respectively. They increased the apoptotic effect confirming the success of the spanlastics to be a potential delivery for melanoma treatment.

Metastatic Melanoma: A Preclinical Model Standardization and Development of a Chitosan-Coated Nanoemulsion Containing Temozolomide to Treat Brain Metastasis.

Melanoma is the most aggressive type of skin cancer. Brain metastasis is the worst scenario in metastatic melanoma and the treatment options for these patients are limited. Temozolomide (TMZ) is a chemotherapy agent used to treat primary central nervous system tumors. Our objective was to develop chitosan-coated nanoemulsion containing temozolomide (CNE-TMZ) for nasal route administration to melanoma brain metastasis treatment. A preclinical model of metastatic brain melanoma was standardized, and the efficiency of the developed formulation was further determined in vitro and in vivo. The nanoemulsion was done by spontaneous emulsification method and the formulation was characterized by size, pH, polydispersity index, and zeta potential. Culture assessments to determine cell viability were done in the A375 human melanoma cell line. To determine the safety of formulation, healthy C57/BL6 mice were treated with a nanoemulsion without TMZ. The model in vivo used B16-F10 cells implanted by stereotaxic surgery in C57/BL6 mice brains. The results demonstrate that the preclinical model used showed to be useful to analyze the efficiency of new candidate drugs to treat melanoma brain metastasis. The chitosan-coated nanoemulsions with TMZ showed the expected physicochemical characteristics and demonstrated safety and efficacy, reducing around 70% the tumor size compared to control mice, and presenting a tendency in mitotic index reduction, becoming an interesting approach to treat melanoma brain metastasis.

Abstract 2826: Electromagnetic fields and hyperthermia induce lysosomal permeabilization and death in different cancer cell models: in vitro and in vivo studies

Abstract Introduction: Non-ionizing radiations (NIR), e.g. tumor-treating fields, high intensity focused ultrasounds (HIFU) or high frequency microwaves, have potential applications in Oncotherapy. Nevertheless, as today, their efficacy is limited. We investigated whether externally applied magnetic fields and non-ablative hyperthermia affect cancer cell viability and growth. Methods: Under in vitro conditions, human A2058 melanoma, AsPC1 pancreas carcinoma, and MDA-MB-231 breast carcinoma cells were treated with electromagnetic fields (EMFs, using solenoids) and/or hyperthermia (HT, using a thermostated water bath). The effect of both energies was also studied in combination with standard chemotherapy and targeted therapy. An experimental system combining EMFs and HIFU-induced HT was developed and tested in vivo. Results: EMFs (25 μT, 4h) or HT (52ºC, 40 min) quite moderately affect cancer cell viability. However, their combination decreased viability to approx. 16%, 50%, and 21% of controls values in A2058, AsPC1, and MDA-MB-231 cells, respectively). Increased lysosomal permeability, release of cathepsins into the cytosol and activation of the mitochondria-dependent cell death are the underlying mechanisms. Cancer cells were completely eliminated combining EMFs, HT and standard chemotherapy or EMFs, HT and anti-Hsp70-targeted therapy. In vivo experiments, using pancreatic cancer xenografts growing in nude mice, show that combination of EMFs, HIFU-induced HT and gemcitabine induces a drastic cancer regression (to approx. 3-4% of control values). Conclusion: Non-invasive NIR (EMFs and HIFU-induced HT) can complement current Oncotherapies. This strategy, associated to current chemotherapy or anti-Hsp70-targeted therapy, may facilitate the elimination of tumors that can be previously located using standard imaging techniques (CT scan and NMR). These results may help to overcome the limitations in the use of different NIR in Oncotherapy. Citation Format: Elena Obrador, Ali Jihad-Jebbar, Rosario Salvador-Palmer, Rafael López-Blanch, María Oriol-Caballo, María Paz Moreno-Murciano, Enrique Navarro, Rosa Cibrián, Jose M. Estrela. Electromagnetic fields and hyperthermia induce lysosomal permeabilization and death in different cancer cell models: in vitro and in vivo studies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2826.

Abstract 4451: Developing a PROTAC-based NR4A1 degrader for melanoma cancer therapy

Abstract Introduction: Melanoma originates from melanocytes within the epidermis and is one of the most common cancers, with nearly 100000, new cases yearly. Despite numerous advancements in therapies to treat melanoma, a prevalent population of patients still do not respond to currently approved therapies. Recent studies have highlighted the role of transcription factor nuclear receptor subfamily 4 group A member 1 (NR4A1) in melanoma for cancer survival, invasion, and metastasis. NR4A1 is also involved in glucose metabolism. Our project aims to identify an effective degrader of NR4A1 using a PROTAC strategy and then validate its effectiveness in reducing the growth of melanoma. PROTACs consists of three domains: a warhead that binds to the protein of interest, a ligand to an E3 ligase, and a linker that brings both domains in proximity to one another. The PROTAC can recruit an E3 ligase to ubiquitinate NR4A1 and degrade it via the ubiquitin-proteasome system (UPS). Results: Our first goal was to identify valid PROTAC candidates that can effectively degrade NR4A1. We identified NR-V04, which demonstrated a dose-dependent degradation of NR4A1 in various melanoma cell lines. We further investigated time-dependent degradation, and NR-V04 was able to achieve in vitro degradation of NR4A1 16 hours after treatment. Additionally, we validated the mechanism of degradation via the UPS through various models. NR-V04 treatment of cells with a VHL knockout, which removes the E3 ligase recruited, experienced no degradation of NR4A1, and treatment of cells treated with MG132, a proteasome inhibitor, also showed no degradation. We observed degradation in vitro of numerous mouse and human cell lines. To investigate the cancer-killing effects of NR-V04, we completed MTS assays on the human melanoma cell lines CHL1 and A375, which showed an EC50 of 0.723 μM and 1.025 μM, respectively. When comparing NR-V04 treated and untreated CHL1 cells, NR-V04 was able to significantly decrease melanoma cell viability. NR4A1 knockout in CHL1 also showed decreased melanoma cell viability, and when comparing NR-V04 treated and untreated in NR4A1 knockout, there was no further decrease in melanoma cell viability. As for in vivo models, NR4A1 knockout in CHL1 and A375 exhibited slower tumor growth compared to the wild type. Furthermore, NR-V04 showed suppression of CHL1 and A375 tumor-bearing NSG mice melanoma growth at low dose concentrations of 1 mg/kg after seven days compared to vehicle and warhead treatment. Western blot analysis of tumor tissue provides support for the ability of NR-V04 to degrade tumor-intrinsic NR4A1. Conclusion: NR-V04 can selectively degrade NR4A1, in vitro and in vivo, to decrease melanoma cancer cell viability via the UPS. NR-V04 holds promising therapeutic potential as a cancer therapy for patients with melanoma. Citation Format: Rohan Master, Yuewan Luo, Yufeng Xiao, Daohong Zhou, Lei Wang, Guangrong Zheng, Weizhou Zhang. Developing a PROTAC-based NR4A1 degrader for melanoma cancer therapy. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4451.

Abstract 1604: Preclinical development of ADCT-211, a novel pyrrolobenzodiazepine dimer-based antibody-drug conjugate targeting solid tumors expressing IL13RA2

Abstract Interleukin 13 receptor subunit alpha 2 (IL13RA2) is one of the two major receptors for the cytokine interleukin 13 (IL-13). While IL13RA1 has low affinity for IL-13 and it is expressed ubiquitously in humans, IL13RA2 has high binding affinity to IL-13 and its expression in normal tissues is mainly restricted to the testes. IL13RA2 was initially regarded as a decoy receptor because it has a short cytoplasmic tail with no known signaling motifs, and its binding to IL-13 does not lead to activation of the JAK/STAT6 pathway. However, recent studies demonstrated that IL-13-mediated IL13RA2 signaling occurs via STAT6-independent pathways. IL13RA2 is reported to be expressed at a high frequency in glioblastoma multiforme (GBM) as well as in other solid tumors including melanoma, renal cell carcinoma and adrenocortical carcinoma and is correlated with poor prognosis. ADCT-211 is an antibody-drug conjugate composed of HuCl47, a humanized IgG1 antibody directed against human IL13RA2, site-specifically conjugated using GlycoConnectTM technology to PL1801, which contains HydraspaceTM, a valine-alanine cleavable linker and the PBD dimer cytotoxin SG2000 (drug to antibody ratio ~ 1.8). The purpose of this study was to characterize the in vitro and in vivo anti-tumor activity of ADCT-211 in human cancer cell lines and xenograft models, to determine its safety, tolerability and pharmacokinetic (PK) in the rat and to measure IL13RA2 expression by immunohistochemistry (IHC) in human tumor specimens. HuCl47 showed specific binding to recombinant IL13RA2, while it did not bind to IL13RA1. In vitro, ADCT-211 had highly potent and targeted cytotoxicity against various IL13RA2-expressing solid cancer cell lines. In vivo, a single dose of ADCT-211 at 10 mg/kg was able to completely eradicate tumors in the subcutaneous (sc) A375 human melanoma xenograft model and resulted in 8/8 tumor-free survivors (TFS) at the end of the study on day 57. In the sc U251 human glioblastoma xenograft model, a single dose of ADCT-211 at 3.75 mg/kg showed potent and durable antitumor activity and it resulted in 5/10 partial responders and 5/10 complete responders, 2 of which were TFS at the end of the study on day 51. In an MTD study in male rats, ADCT-211 was stable and tolerated up to 24 mg/kg single dose, with exposure data being indicative of a linear PK profile with a half-life of 12-19 days. Expression of membranous IL13RA2 was confirmed by IHC in a panel of primary and refractory GBM samples and malignant melanoma, highlighting them as potential indications for the clinical development of ADCT-211. In conclusion, ADCT-211 demonstrated potent and specific in vitro and in vivo anti-tumor activity and it was stable and well tolerated in the rat, warranting further development of ADCT-211 into the clinic in IL13RA2-expressing cancers. Citation Format: Francesca Zammarchi, Karin Havenith, Ben Leatherdale, Britanny Roberts, Lara Montolio, Afroze Patel, Narinder Janghra, Pedro Alves, Kristina Zaitseva, Cecile Oblette, Ian Kirby, Lolke de Haan, Patrick H van Berkel. Preclinical development of ADCT-211, a novel pyrrolobenzodiazepine dimer-based antibody-drug conjugate targeting solid tumors expressing IL13RA2 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1604.

Zanthoxylum bungeanum seed oil inhibits tumorigenesis of human melanoma A375 by regulating CDC25A/CyclinB1/CDK1 signaling pathways in vitro and in vivo.

Background: Zanthoxylum bungeanum seed oil (ZBSO) is extracted from the seeds of the traditional Chinese medicine Z. bungeanum Maxim, which has been shown to have anti-melanoma effects. However, the specific mechanisms are not illustrated adequately. Aims: To further investigate the mechanism by which ZBSO inhibits melanoma and to provide scientific evidence to support ZBSO as a potential melanoma therapeutic candidate. Methods: CCK-8 assays were used to detect the function of ZBSO on A375 cells. Based on transcriptomics analyses, Western blot analysis was applied to determine whether an association existed in ZBSO with the CDC25A/CyclinB1/CDK1 signaling pathway. In addition, RT-qPCR and immunohistochemistry analysis validated that ZBSO has the anti-melanoma effect in a nude mouse xenograft model of human melanoma. Then, 16S rRNA sequencing was used to detect the regulation of gut microbes. Results: Cellular assays revealed that ZBSO could inhibit A375 cell viability by regulating the cell cycle pathway. Further studies presented that ZBSO could constrain CDC25A/CyclinB1/CDK1 signaling pathway in vitro and in vivo models of melanoma. ZBSO did not produce toxicity in mice, and significantly reduced tumor volume in xenotransplants of A375 cells. Genome analysis indicated that ZBSO successfully altered specific gut microbes. Conclusion: ZBSO inhibited the growth of A375 cells by regulating CDC25A/cyclinB1/CDK1 signaling pathway both in vitro and in vivo, suggesting that ZBSO may be a novel potential therapeutic agent.

Silver Nanoparticles Synthesized from Abies alba and Pinus sylvestris Bark Extracts: Characterization, Antioxidant, Cytotoxic, and Antibacterial Effects.

In recent years, phytofunctionalized AgNPs have attracted great interest due to their remarkable biological activities. In the present study, AgNPs were synthesized using Abies alba and Pinus sylvestris bark extracts. The chemical profile of these bark extracts was analyzed by LC-HRMS/MS. As a first step, the synthesis parameters (pH, AgNO3 concentration, ratio of bark extract and AgNO3, temperature, and reaction time) were optimized. The synthesized AgNPs were characterized by ATR-FTIR spectroscopy, DLS, SEM, EDX, and TEM. Their antioxidant, cytotoxic, and antibacterial properties were evaluated by the DPPH, ABTS, MTT, and broth microdilution assays, respectively. Abies alba and Pinus sylvestris bark extract-derived AgNPs were well-dispersed, spherical, small (average particle size of 9.92 and 24.49 nm, respectively), stable (zeta potential values of -10.9 and -10.8 mV, respectively), and cytotoxic to A-375 human malignant melanoma cells (IC50 = 2.40 ± 0.21 and 6.02 ± 0.61 μg/mL, respectively). The phytosynthesized AgNPs also showed antioxidant and antibacterial effects.

LL-37 Might Promote Local Invasion of Melanoma by Activating Melanoma Cells and Tumor-Associated Macrophages

LL-37 can stimulate various skin-resident cells to contribute to tumor development. Since tumor (T) stage is determined by the vertical invasion of tumor cells in melanoma, we hypothesized that the LL-37 expression level is correlated with the T stage in melanoma patients. Immunohistochemical staining of LL-37 was performed in each stage of melanoma (Tis-T4), suggesting the ratio of LL-37-expressing cells correlate positively to T stage severity. Next, to examine pro-angiogenetic factors induced by LL-37 stimulation, the B16F10 melanoma model was used. Intra-tumorally administered CRAMP, the mouse ortologe of LL-37, significantly increased the mRNA expression of CXCL5, IL23A, MMP1a, and MMP9 in B16F10 melanoma. To confirm the induction of pro-angiogenic factors, A375 human melanoma cells were stimulated by LL-37 in vitro. The mRNA expression of CXCL5, IL23A, and MMP9, but not MMP1, were significantly increased by LL-37 stimulation. Moreover, LL-37-stimulated A375 culture supernatant promoted tube networks, suggesting that these tumor-derived factors promote the pro-angiogenic effect on tumor development. In contrast to melanoma cell lines, M2 macrophages stimulated by LL-37 in vitro significantly increased their expression and secretion of MMP-1, but not MMP-9 expression. Collectively, these results suggest that LL-37 stimulates both tumor cells and macrophages to promote melanoma invasion by the induction of pro-angiogenic factors.

Comparison of In Vitro Antimelanoma and Antimicrobial Activity of 2,3-Indolo-betulinic Acid and Its Glycine Conjugates

Malignant melanoma is one of the most pressing problems in the developing world. New therapeutic agents that might be effective in treating malignancies that have developed resistance to conventional medications are urgently required. Semisynthesis is an essential method for improving the biological activity and the therapeutic efficacy of natural product precursors. Semisynthetic derivatives of natural compounds are valuable sources of new drug candidates with a variety of pharmacological actions, including anticancer ones. Two novel semisynthetic derivatives of betulinic acid-N-(2,3-indolo-betulinoyl)diglycylglycine (BA1) and N-(2,3-indolo-betulinoyl)glycylglycine (BA2)-were designed and their antiproliferative, cytotoxic, and anti-migratory activity against A375 human melanoma cells was determined in comparison with known N-(2,3-indolo-betulinoyl)glycine (BA3), 2,3-indolo-betulinic acid (BA4) and naturally occurring betulinic acid (BI). A dose-dependent antiproliferative effect with IC50 values that ranged from 5.7 to 19.6 µM was observed in the series of all five compounds including betulinic acid. The novel compounds BA1 (IC50 = 5.7 µM) and BA2 (IC50 = 10.0 µM) were three times and two times more active than the parent cyclic structure B4 and natural BI. Additionally, compounds BA2, BA3, and BA4 possess antibacterial activity against Streptococcus pyogenes ATCC 19615 and Staphylococcus aureus ATCC 25923 with MIC values in the range of 13-16 µg/mL and 26-32 µg/mL, respectively. On the other hand, antifungal activity toward Candida albicans ATCC 10231 and Candida parapsilosis ATCC 22019 was found for compound BA3 with MIC 29 µg/mL. This is the first report of antibacterial and antifungal activity of 2,3-indolo-betulinic acid derivatives and also the first extended report on their anti-melanoma activity, which among others includes data on anti-migratory activity and shows the significance of amino acid side chain on the observed activity. The obtained data justify further research on the anti-melanoma and antimicrobial activity of 2,3-indolo-betulinic acid derivatives.

Multifunctional bismuth oxide (Bi2 O3 ) particles: Evidence for selective melanoma therapy.

The current study investigates the therapeutic and optical properties of bismuth oxide (Bi2 O3 ) particles for selective melanoma therapy and prevention. The Bi2 O3 particles were prepared using a standard precipitation method. The Bi2 O3 particles induced apoptosis in human A375 melanoma cells but not human HaCaT keratinocytes or CCD-1090Sk fibroblast cells. This selective apoptosis appears to be associated with a combination of factors: increased particle internalization (2.29 ± 0.41, 1.16 ± 0.08 and 1.66 ± 0.22-fold of control) and enhanced production of reactive oxygen species (ROS) (3.4 ± 0.1, 1.1 ± 0.1 and 2.05 ± 0.17-fold of control) in A375 cells compared to HaCaT and CCD-1090SK cells, respectively. As a high-Z element, bismuth is also an excellent contrast agent for computer tomography, which renders Bi2 O3 a theranostic material. Moreover, Bi2 O3 displays high UV absorption and low photocatalytic activity compared to other semiconducting metal oxides, which opens further potential fields of application as a pigment or as an active ingredient in sunscreens. Overall, this study demonstrates the multifunctional properties of Bi2 O3 particles surrounding the treatment and prevention of melanoma.

Gene Profiling of Cannabis-sativa-mediated Apoptosis in Human Melanoma Cells.

Malignant melanoma is an aggressive skin cancer, accounting for the majority of skin cancer deaths. Prognosis is often poor and finding effective treatment remains a challenge. Tetrahydrocannabinol (THC) and cannabidiol (CBD) are main bioactive components of Cannabis sativa plant extracts that have been shown to exert anti-tumor effects. In this study, we aimed to perform gene expression analysis of human melanoma A375 cells following stimulation with C. sativa extracts. Gene expression profiles of A375 human melanoma and Vero (control) cell lines were evaluated by RNA sequencing and quantitative real-time PCR. Flow cytometry showed that the THC+CBD cannabis fractions induced apoptosis on A375 cells. Induction of apoptosis was accompanied by a notable up-regulation of DNA damage inducible transcript 3 (DDIT), nerve growth factor receptor (NGFR), colony-stimulating factor 2 (CSF2), growth arrest and DNA damage inducible beta (GADD45B), and thymic stromal lymphopoietin (TSLP) genes and down-regulation of aryl hydrocarbon receptor nuclear translocator 2 (ARNT2), cyclin E2 (CCNE2), integrin subunit alpha 9 (ITGA9), proliferating cell nuclear antigen (PCNA) and E2F transcription factor 1 (E2F1) genes. Treatment of A375 cells with the THC+CBD fraction inhibited the phosphorylation of ERK1/2 signaling pathway, which regulates melanoma cell proliferation. We showed that the THC+CBD combination disrupted melanoma cell migration. Use of C. sativa-derived extracts containing equal amounts of THC and CBD is proposed as a potential treatment of melanoma.

Nanoemulsion and Nanogel Containing Cuminum cyminum L Essential Oil: Antioxidant, Anticancer, Antibacterial, and Antilarval Properties.

Cuminum cyminum L. is a widespread medicinal plant with a broad spectrum of biological activity. In the present study, the chemical structure of its essential oil was examined utilizing GC-MS analysis (gas chromatography-mass spectrometry). Then, a nanoemulsion dosage form was prepared with a droplet size and droplet size distribution (SPAN) of 121 ± 3 nm and 0.96. After that, the dosage form of the nanogel was prepared; the nanoemulsion was gelified by the addition of 3.0% carboxymethyl cellulose. In addition, the successful loading of the essential oil into the nanoemulsion and nanogel was approved by ATR-FTIR (attenuated total reflection Fourier transform infrared) analysis. The IC50 values (half maximum inhibitory concentration) of the nanoemulsion and nanogel against A-375 human melanoma cells were 369.6 (497-335) and 127.2 (77-210) μg/mL. In addition, they indicated some degrees of an antioxidant activity. Interestingly, after treatment of Pseudomonas aeruginosa with 5000 µg/mL nanogel, bacterial growth was completely (∼100%) inhibited. In addition, the growth of Staphylococcus aureus after treatment with the 5000 μg/ml nanoemulsion was decreased by 80%. In addition, nanoemulsion and nanogel LC50 values for Anopheles stephensi larvae were attained as 43.91 (31-62) and 123.9 (111-137) µg/mL. Given the natural ingredients and promising efficacy, these nanodrugs can be regarded for further research against other pathogens or mosquito larvae.

Effect of Extraction Solvent and Harvest Time on Recovery of Bioactive Com-pounds, Antioxidant and Cancer Cell Growth Inhibition Activities of Korean Ca-mellia mistletoe

This study aimed to determine the phytochemical profiles,in vitroantioxidant and antiproliferative properties of KoreanCamellia mistletoe(Korthalsella japon-ica(Thunb.) Engl.) depending on the harvest time (August and November) and sol-vent (100% methanol, 70% ethanol, and hot water). TheCamellia japonicaL. mistle-toe extracts were analyzed for total phenol, flavonoid, carotenoid, and L-ascorbic acid contents and antioxidant properties such as scavenging capacities (1,1-diphenyl-2-pic-rylhydrazyl (DPPH) and NO•),ferrous ion chelating and reducing power. Concur-rently, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) as-say was used to assess the antiproliferative properties against human cancer cell lines; MCF (human breast cancer cells), HeLa (human cervical cancer cells), A375 (human malignant melanoma cells), HCT116 (human colon cancer cells), HepG2 (human liver cancer cells) and A549 (human non-small cell lung adenocarcinoma cells). The results showed that extraction solvent and harvest time had significant impacts on antioxidant and anticancer activities and selectivity for free phenolic compounds inCamellia ja-ponicaL. mistletoe. Among all the tested extracts, the highest amounts of total phe-nolic and total flavonoids content were found in ethanol extracts ofCamellia japon-icaL. mistletoe harvested in November, while the methanol extracts ofCamellia ja-ponicaL. mistletoe collected in August showed the highest contents of total carote-noids and L-ascorbic acidscompared to the other tested extracts. Additionally, the highest NO•radical scavenging activity was found in ethanol extracts, whereas the strongest DPPH radical scavenging activity was found in methanol extracts ofCamel-lia japonicaL. mistletoe harvested in November. Furthermore, methanol extracts showed much higher antiproliferative activity against all human cancer cells than eth-anol extracts ofCamellia japonicaL. mistletoe harvested in November. In conclusion, the antioxidant and anticancer properties ofCamellia japonicaL. mistletoe showed significant dependence on the extraction solvent type and harvest time. With optimum harvest time and extraction solvent, they boast a wide range of promising medical, pharmaceutical, and food applications

Small Structural Differences Govern the Carbonic Anhydrase II Inhibition Activity of Cytotoxic Triterpene Acetazolamide Conjugates.

Acetylated triterpenoids betulin, oleanolic acid, ursolic acid, and glycyrrhetinic acid were converted into their succinyl-spacered acetazolamide conjugates. These conjugates were screened for their inhibitory activity onto carbonic anhydrase II and their cytotoxicity employing several human tumor cell lines and non-malignant fibroblasts. As a result, the best inhibitors were derived from betulin and glycyrrhetinic acid while those derived from ursolic or oleanolic acid were significantly weaker inhibitors but also of diminished cytotoxicity. A betulin-derived conjugate held a Ki = 0.129 μM and an EC50 = 8.5 μM for human A375 melanoma cells.

Biogeneration of silver nanoparticles from Cuphea procumbens for biomedical and environmental applications

Nanotechnology is one of the most important and relevant disciplines today due to the specific electrical, optical, magnetic, chemical, mechanical and biomedical properties of nanoparticles. In the present study we demonstrate the efficacy of Cuphea procumbens to biogenerate silver nanoparticles (AgNPs) with antibacterial and antitumor activity. These nanoparticles were synthesized using the aqueous extract of C. procumbens as reducing agent and silver nitrate as oxidizing agent. The Transmission Electron Microscopy demonstrated that the biogenic AgNPs were predominantly quasi-spherical with an average particle size of 23.45 nm. The surface plasmonic resonance was analyzed by ultraviolet visible spectroscopy (UV–Vis) observing a maximum absorption band at 441 nm and Infrared Spectroscopy (FT IR) was used in order to structurally identify the functional groups of some compounds involved in the formation of nanoparticles. The AgNPs demonstrated to have antibacterial activity against the pathogenic bacteria Escherichia coli and Staphylococcus aureus, identifying the maximum zone of inhibition at the concentration of 0.225 and 0.158 µg/mL respectively. Moreover, compared to the extract, AgNPs exhibited better antitumor activity and higher therapeutic index (TI) against several tumor cell lines such as human breast carcinoma MCF-7 (IC50 of 2.56 µg/mL, TI of 27.65 µg/mL), MDA-MB-468 (IC50 of 2.25 µg/mL, TI of 31.53 µg/mL), human colon carcinoma HCT-116 (IC50 of 1.38 µg/mL, TI of 51.07 µg/mL) and melanoma A-375 (IC50 of 6.51 µg/mL, TI of 10.89 µg/mL). This fact is of great since it will reduce the side effects derived from the treatment. In addition, AgNPs revealed to have a photocatalytic activity of the dyes congo red (10–3 M) in 5 min and malachite green (10–3 M) in 7 min. Additionally, the degradation percentages were obtained, which were 86.61% for congo red and 82.11% for malachite green. Overall, our results demonstrated for the first time that C. procumbens biogenerated nanoparticles are excellent candidates for several biomedical and environmental applications.

Nano-scaled emulsion and nanogel containing Mentha pulegium essential oil: cytotoxicity on human melanoma cells and effects on apoptosis regulator genes

BackgroundTopical drug delivery using nanoemulsions and nanogels is a promising approach to treating skin disorders such as melanoma.MethodsIn this study, the chemical composition of Mentha pulegium essential oil with five major compounds, including pulegone (68.11%), l-menthone (8.83%), limonene (2.90%), iso-pulegone (2.69%), and iso-menthone (1.48%) was first identified using GC-MS (Gas chromatography–Mass Spectrometry) analysis. Afterward, a nano-scaled emulsion containing the essential oil with a droplet size of 7.70 ± 1 nm was prepared. Nanogel containing the essential oil was then prepared by adding (2% w/v) carboxymethyl cellulose to the nano-scaled emulsion. Moreover, the successful loading of M. pulegium essential oil in the nano-scaled emulsion and nanogel was confirmed using ATR-FTIR (Attenuated total reflectance-Fourier Transform InfraRed) analysis. Then, human A375 melanoma cells were treated with different concentrations of samples, the MTT assay evaluated cell viability, and cell apoptosis was confirmed by flow cytometry. In addition, the expression of apoptotic and anti-apoptotic genes, including Bax and Bcl-2, was evaluated using the qPCR (quantitative Polymerase Chain Reaction) technique.ResultsThe results showed that cell viability was reduced by 90 and 45% after treatment with 300 μg/mL of the nanogel and nano-scaled emulsion. As confirmed by flow cytometry, this effect was mediated by apoptosis. Furthermore, gene expression analysis showed up-regulation of Bax and down-regulation of Bcl-2 genes. Therefore, the prepared nanogel, with high efficacy, could be considered a potent anticancer agent for supplementary medicine and in vivo research.

The Antimelanoma Biological Assessment of Triterpenic Acid Functionalized Gold Nanoparticles.

One of several promising strategies for increasing the bioavailability and therapeutic potential of high-lipophilic biologically active compounds is gold nanoparticle formulation. The current study describes the synthesis and biological antimelanoma evaluation of three triterpen-functionalized gold nanoparticles, obtained using our previously reported antimelanoma benzotriazole-triterpenic acid esters. Functionalized gold nanoparticle (GNP) formation was validated through UV-VIS and FTIR spectroscopy. The conjugate's cytotoxic effects were investigated using HaCaT healthy keratinocytes and A375 human melanoma cells. On A375 cells, all three conjugates demonstrated dose-dependent cytotoxic activity, but no significant cytotoxic effects were observed on normal HaCaT keratinocytes. GNP-conjugates were found to be more cytotoxic than their parent compounds. After treatment with all three GNP-conjugates, 4,6'-diamidino-2-phenylindole (DAPI) staining revealed morphological changes consistent with apoptosis in A375 melanoma cells. Quantitative real-time polymerase chain reaction (RT-qPCR) analysis revealed that the triterpene-GNP conjugate treated A375 melanoma cells had a fold change increase in Bcl-2-associated X protein (BAX) expression and a fold change decrease in B-cell lymphoma 2 (Bcl-2) expression. In A735 melanoma cells, high-resolution respirometry studies revealed that all three GNP-conjugates act as selective inhibitors of mitochondrial function. Furthermore, by examining the effect on each mitochondrial respiratory rate, the results indicate that all three conjugates are capable of increasing the production of reactive oxygen species (ROS), an apoptosis trigger in cancer cells.

SQUID PENS MEDIATED SILVER NANOPARTICLES SYNTHESIS, ITS CHARACTERISATION, AND BIOLOGICAL ACTIVITIES

This research scintillates the synthesis of silver nanoparticles from chitosan (C-AgNps) from squid pens as a reducing and stabilizing agent and assesses its antioxidant, antibacterial, and anticancer potentials. The nanoparticles were 240 to 290 nm with excellent stability of +98.2 mV. The C-AgNps possessed an excellent 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging activity with an IC50 of 6.65 µg/mL. The C-AgNps showed exceptional antibacterial properties against S. aureus, B. subtilis, P. aeruginosa, and E. coli using a good diffusion assay. The C-AgNps also showed outstanding anticancer properties against A375 human skin malignant melanoma. Thus, the C-AgNps could be used as antimicrobial and anticancer agents for novel therapeutics after lead optimization and in-vivo clinical trials.

Synthesis, molecular docking, and biological evaluation of [3,2-b]indole fused 18β-glycyrrhetinic acid derivatives against skin melanoma.

Melanoma, the most serious yet uncommon type of cancer, originates in melanocytes. Risk factors include UV radiation, genetic factors, tanning lamps and beds. Here, we described the synthesis and selective anti melanoma activity of [3,2-b]indole fused 18β-glycyrrhetinic acid, a derivative of 18β-glycyrrhetinic acid in murine B16F10 and A375 human melanoma cell lines. Among the 14 molecules, GPD-12 showed significant selective cytotoxic activity against A375 and B16F10 cell lines with IC50 of 13.38 μM and 15.20 μM respectively. GPD 12 induced the formation of reactive oxygen species in A375 cells that could trigger oxidative stress mediated cell death as is evident from the increased expression of apoptosis related proteins such as caspase-9 and caspase-3 and the increased ratio of Bax to Bcl2. The results showed that GPD 12 can be used as an effective therapeutic agent against melanoma.

A new amide alkaloid induces the apoptosis of human melanoma A375 cellsviainhibition of the STAT3 signaling pathway

A new amide alkaloid (ZYL-01) can inhibit the activity of A375 cells by inducing cell apoptosisviainhibiting STAT3 signaling.