Apoptotic Activity Research Articles

Teratogenic Effects of Drugs on Primary Lymphocytes Assessed by Flow Cytometry.

Peripheral blood lymphocytes as primary cells can be isolated from human, animal, fetus, and placenta. These cells are an excellent cellular model for the assessment of cytotoxicity, genotoxicity, oxidative stress, and mitochondrial and lysosomal dysfunction induced by drug and chemicals. Moreover, peripheral blood lymphocytes are an easily available source of primary cells appropriate for basic research and in cellular studies regarding teratogenic, genotoxic, and cytotoxic effect of drugs and chemicals. Most drugs and other chemicals that produce birth defects, known as teratogenic agents, produce reactive oxygen species (ROS) formation and mitochondrial and lysosomal dysfunction. It seems that there is an important mechanistic link between oxidative stress, mitochondrial damages, lysosomal integrity, and teratogenic drug-induced birth defects. One of the most sensitive periods in the embryo is transition from an important developmental event to another such as transition from proliferation to differentiation. Mitochondria, lysosomes, and cellular ROS have an important role in proliferative, differentiative, and apoptotic activities during the development. Therefore, disruption of the function of mitochondria, lysosomes, oxidative stress, and redox imbalance leads to cellular dysfunctions and subsequently poor developmental outcomes in the fetus. In this chapter, we will focus on evaluation of mitochondrial/lysosomal functions and estimation of ROS formation using flow cytometry methods in isolated lymphocytes and their isolated mitochondria.

Phytochemical profiling and anticancer potential of gardenia latifolia extracts against arsenic trioxide induced liver fibrosis in rat model.

Arsenic trioxide (As2O3) is an environmental contaminant that may cause hepatic injuries. As2O3-induced liver injuries are detected as an underlying cause of hepatocellular carcinoma (HCC) around the globe. The present study aimed to investigate the potential of Gardenia latifolia (GL) extracts against oxidative stress and apoptotic activity in HCC-induced rats and to explore in silico molecular docking analysis of phytocompounds of G. latifolia. The present study was designed to investigate the hepato-protective effect of ethanol and n-hexane extract of G. latifolia. Phytochemical analysis was performed using gas-chromatography-mass spectrometry (GC-MS), and the identified metabolites were used for computational docking analysis. The binding potential and inhibitory effect of the identified metabolites against inflammatory markers were assessed. Fifty male albino rats were selected for the in vivo study and were randomly divided into five groups, with 10 rats in each group. Group I is the control group. Hepatotoxicity was induced in groups II, III, IV, and V with 350mg/kg/day of As2O3. Group II was taken as positive control, Group III and IV were treated with ethanol and n-hexane extract of G. latifolia, respectively, and Group V was treated with cisplatin 3.0mg/kg/day. At the end of treatment, different stress and liver biomarkers were also analyzed. The quantitative phytochemical profiling revealed a high content of total flavonoid and tannins found at 5.731 ± 0.1856mg quercetin equivalent (QE)/g and 86.31 ± 14.20mg tannic acid equivalent (TAE)/g in G. latifolia n-hexane extract, while a significant concentration of TFC was 276.821 ± 2.19mg gallic acid equivalent (GAE)/g, in ethanolic extract. GC-MS analysis resulted in the identification of 26 metabolites in ethanol extract while 32 metabolites in n-hexane extract, respectively. Both the extracts restored the abnormal levels of stress markers (p < 0.05) in Groups III and IV, and were comparable to the comparative control group V, which was given cisplatin as the standard drug. The histopathological examination revealed the regeneration of hepatocytes, dilated sinusoidal cells, necrosis, and distorted hepatic architecture observed in arsenic trioxide hepatotoxic liver. Among the top most identified metabolites from GC-MS analysis, stigmasterol exhibited -8.3 and -7.1kcal/mol in silico binding affinities against cyclooxygenase-2 (COX-2), and interleukin (IL-6), respectively, while Dasycarpidan-1-methanol exhibited the best binding affinities of -6.8 and -7.2kcal/mole against matrixmetalloprotinease (MMP)-3 and heat shock protein-90 (HSP-90), respectively. 6-AH-cAMP showed the best docking score of -7.5kcal/mol for the vascular endothelial growth factor (VEGF) macromolecule. Metabolite Dasycarpidan-1-methanol, acetate represented drug like properties so it was further analyzed by MD simulation and stable dynamic nature of protein ligand complex was evaluated. In conclusion, the effective therapeutic potential of G. latifolia extracts targeted oxidative stress, increasing antioxidant activities and inhibiting inflammation and liver complications at early stages. Further research on the molecular level may further explore the anticancer potential of this plant against various types of cancers.

Supported silver nanoparticles over Metronidazole modified-single walled carbon nanotubes for overcoming anti-microbial properties, anti-cancer effect, and apoptosis induction on human gastric cancer cell line

One of the most important goals of current research in the medical field is to identify treatments for antimicrobial resistance (AMR) and cancer. The latest developments in nanotechnology have provided opportunities to develop novel nanomaterials that can overcome the limitations of current treatments. In this regard, carbon nanotubes (CNTs) are considered potential nano-tools with attractive implications. The analysis of MTZ/SWCNT/AgNPs in this study was done using different techniques, including SEM, TEM, FTIR, XRD, and EDX. Antibacterial activity was assessed for MTN/SWCNT/AgNPs against Escherichia coli, Pseudomonas aeruginosa, Enterococcus faecalis, and Staphylococcus aureus. The effects of MTZ/SWCNTs/AgNPs on mitochondrial and apoptotic activities in human gastric cancer (AGS) were evaluated. Overall, new CNTs were produced efficiently. MTZ/SWCNTs/AgNPs are considered efficient antibacterial and anticancer agents for biological applications.

Design and synthesis of Meldrum's acid based 7-azaindole anchored 1,2,3-triazole hybrids as anticancer agents.

A series of Meldrum's acid, 7-azaindole and 1,2,3-triazole hybrids were synthesized and evaluated for their in vitro anticancer activity against five different cancer cell lines viz. MCF-7 (breast cancer), HeLa (cervical cancer), DU-145 (prostate cancer), HepG2 (liver cancer) and K562 (myelogenous leukemia cell). Among the series, compound 6b containing a 4-methyl substitution showed potent activity against HeLa cell line. Cell cycle analysis revealed that compound 6b induced cell cycle arrest at the G2/M phase and induced apoptosis. Apoptotic activity was further confirmed by Hoechst staining and Annexin V-FITC assay. Compound 6b has been found to exhibit higher activity in all four cell lines, with IC50 values of 6.67 ± 0.39 μM, 4.44 ± 0.32 μM, 12.38 ± 0.51 μM and 9.97 ± 0.25 μM against MCF-7, HeLa, DU-145 and HepG2 cell lines respectively. Compounds 6m (9.68 ± 0.10 μM) and 6n (9.52 ± 0.38 μM), which have dimethoxy and trimethoxy substitutions, respectively, have demonstrated significant anticancer activity against HeLa cells compared to the other cells. The molecular docking study of ligand 6b against the crystal structure of EGFR and Mcl-1 scored notable binding energy values and displayed important interactions like H-bond, π-cation and other hydrophobic interactions.

Investigation of Antiproliferative, Apoptotic, and Migration Activity of Combination of Paclitaxel and Aloe Vera in Non-Small Cell Lung Cancer and Breast Cancer

Investigation of Antiproliferative, Apoptotic, and Migration Activity of Combination of Paclitaxel and Aloe Vera in Non-Small Cell Lung Cancer and Breast Cancer

Design, synthesis, and biological evaluation of novel quinoline-based EGFR/HER-2 dual-target inhibitors as potential anti-tumor agents.

Dual targeting of EGFR and HER2 is a valid anti-cancer approach for treating solid tumors. We designed and synthesized a new series of EGFR/HER-2 dual-target inhibitors based on quinoline derivatives. The structure of the newly synthesized compounds was verified using 1H NMR, 13C NMR, and elemental analysis. The targeted compounds were tested for antiproliferative efficacy against four cancer cell lines. All the compounds had GI50s ranging from 25 to 82 nM, with breast (MCF-7) and lung (A-549) cancer cell lines being the most sensitive. Compound 5a demonstrated the most significant antiproliferative action. With inhibitory (IC50) values of 71 and 31 nM, respectively, compound 5a proved to be the most effective dual-target inhibitor of EGFR and HER-2, outperforming the reference erlotinib (IC50 = 80 nM) as an EGFR inhibitor but falling short of the clinically used agent lapatinib (IC50 = 26 nM) as a HER2 inhibitor. The apoptotic potential activity of 5a was examined, and the findings demonstrated that 5a promotes apoptosis by activating caspase-3, 8, and Bax while simultaneously reducing the expression of the anti-apoptotic protein Bcl-2. The docking studies provided valuable insights into the binding interactions of compounds 3e and 5a with EGFR, effectively rationalizing the observed SAR trends.

Anti-proliferative activity and apoptotic induction of tannins extracted from Quercus infectoria on oral cancer KB cell lines

The present study aimed to evaluate the phytochemicals in Quercus infectoria ethanolic extract and to test the cytotoxic effect on an oral cancer cell line KERATIN-HeLa cells (KB cells). The cytotoxic potential of tannins extracted from Q. infectoria was tested by cell cycle analysis, transforming growth factor (TGF)-beta expression, matrix metalloproteinase (MMP) by fluorescent activated cell sorte, Caspase 3, and Caspase 9 expression by ELISA in KB cell lines treated with the extract. Specific protein (Bcl-2) and the gene expression in the KB cell line upon tannin treatment were detected by western blot and RT-PCR technique. The Caspase 3 and 9 enzymatic activity was carried out using the ELISA method. Virtual screening and molecular docking analysis were conducted to know the binding affinity against the targets matrix metalloproteinase-2 (MMP-2), NF-kB, and RhoA, which are targets of oral cancer cells. Preliminary screening of Q. infectoria indicates it primarily contains tannins and glycosides. The IC50 value of the ethanolic extract was determined to be 76.82 μg/ml. Analysis of the cell cycle revealed that the extract induced dose-dependent arrest in the G0/G1 phase. It is also revealed that KB cell treatment with the extract led to downregulation of the anti-apoptotic protein Bcl-2. Moreover, TGF beta expression was downregulated, and Caspase 3 and Caspase 9 were up-regulated in a dose-dependent manner. In addition, the extract induced mitochondrial membrane potential and MMP induction. Virtual screening and molecular docking revealed that tannins have a favorable binding affinity against MMP-2, NF-kB p65, and RhoA. The study identified that “6-O-digalloyl-1,2,3,4 tetra-O-galloyl-β-D-glucose,” a gallotannin in Q. infectoria, is responsible for the anti-cancer activity. The results of our study found that Q. infectoria extracts have anti-proliferative and apoptotic induction activity, which can help in novel drug discovery and development to alleviate cancer.

Tuning of TRAIL clustering on the surface of nanoscale liposomes by phase separation.

Phase-separated liposomes were used to formulate tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), a protein that selectively kills cancer cells while sparing most healthy ones. By controlling the average number of TRAIL molecules per liposome, we demonstrate the ability to tune the formation of TRAIL clusters and their resulting apoptotic activity.

Integrating computational and experimental chemical biology revealed variable anticancer activities of phosphodiesterase isoenzyme 5 inhibitors (PDE5i) in lung cancer.

Phosphodiesterase 5 (PDE5), an enzyme responsible for catalyzing the degradation of cyclic guanosine monophosphate (cGMP), has been linked to the development of cancer. PDE5 inhibitors (PDE5i), such as sildenafil (Viagra) and tadalafil (Cialis), work by blocking the action of PDE5 and are used primarily as treatments for erectile dysfunction and arterial hypertension. Some studies suggested a potential link between PDE5i and increased cancer risk, while other studies showed preferable antitumor effects. The present study is attempting to shed light on the systems biology effects of PDE5i by applying an integrative informatics approach followed by experimental validation methods including cell viability, cell motility, and proliferation capacity. Cell cycle and apoptosis analyses were carried out using flow cytometry, while real-time polymerase chain reaction (PCR) and western blotting were used to determine the relative gene and protein expression respectively. Our results indicated that the examined PDE5i significantly inhibited the proliferation of lung cancer cells, in addition to reducing wound closure and the mean colony count and size. Furthermore, PDE5i increased the early and late apoptotic activities and suppressed the gene and protein expression of PDE5 in lung cancer cells. The combination of cisplatin and raloxifene with PDE5i resulted in a synergistic effect. This study provides solid evidence supporting the anti-tumorigenic effect of PDE5i in lung cancer cells.

Design, synthesis, and biological evaluation of novel thiazole derivatives as PI3K/mTOR dual inhibitors.

The development of anticancer drugs targeting both PI3K and mTOR pathways is recognized as a promising cancer therapeutic approach. In the current study, we designed and synthesized seventeen new thiazole compounds to investigate their effect on both PI3K and mTOR as well as their anti-apoptotic activity. All the synthesized thiazoles were investigated for their antiproliferative activity on a panel of 60 different cancer cell lines at the National Cancer Institute. Compounds 3b and 3e were selected for further investigation at five dose concentrations due to their effective growth inhibiting activity. Compounds 3b and 3e were further evaluated for their in vitro inhibitory activities against PI3Kα and mTOR compared to alpelisib and dactolisib, respectively as reference drugs. The inhibitory effect of compound 3b on PI3Kα was similar to alpelisib, but it showed weaker inhibitory activity on mTOR compared to dactolisib. Moreover, compound 3b exhibited significantly higher inhibitory activity compared to compound 3e against both PI3Kα and mTOR. The cell cycle analysis showed that compounds 3b and 3e induced G0-G1 phase cell cycle arrest in the leukemia HL-60(TB) cell line. Meanwhile, they significantly increased the total apoptotic activity which was supported by an increase in the level of caspase-3 in leukemia HL-60(TB) cell lines. Molecular docking experiments provided additional explanation for these results by demonstrating the ability of these derivatives to form a network of key interactions, known to be essential for PI3Kα/mTOR inhibitors. All these experimental results suggested that 3b and 3e are potential PI3Kα/mTOR dual inhibitors and could be considered promising lead compounds for the development of anticancer agents.

Development and validation of an RP-HPLC method to compare the apoptotic activity of quercetin found in marketed dietary supplements

Development and validation of an RP-HPLC method to compare the apoptotic activity of quercetin found in marketed dietary supplements

Enabling continuous immune cell recirculation on a microfluidic array to study immunotherapeutic interactions in a recapitulated tumour microenvironment.

The effects of immunotherapeutics on interactions between immune and cancer cells are modulated by multiple components in the tumour microenvironment (TME), including endothelium and tumour stroma, which provide both a physical barrier and immunosuppressive stimuli. Herein, we report a recirculating chip to enable continuous immune cell recirculation through a microfluidic cell array to include these crucial players. This system consists of a three-layered cell array (μFCA) spatially emulating the TME, with tailored fluidic circuits establishing T cell recirculation. This platform enables the study of dynamics among the TME, immune cells in a circulatory system and cancer cell responses thereof. Through this system, we found that tumour endothelium hindered T cell infiltration into the reconstructed breast cancer tumour compartment. This negative effect was alleviated when treated with anti-human PD-L1 (programmed cell death ligand 1) antibody. Another key stromal component - cancer associated fibroblasts - attenuated T cell infiltration, compared against normal fibroblasts, and led to reduced apoptotic activity in cancer cells. These results confirm the capability of our tumour-on-a-chip system in identifying some key axes to target in overcoming barriers to immunotherapy by recapitulating immune cell interactions with the reconstructed TME. Our results also attest to the feasibility of scaling up this system for high-throughput cancer immunotherapeutic screening.

A comparative study of Mentha longifolia var. asiatica and Zygophyllum arabicum ZnO nanoparticles against breast cancer targeting Rab22A gene.

Breast cancer is the most frequently diagnosed cancer worldwide, and the incidence rate has increased enormously over the last three decades. Rab proteins are members of the Rab GTPase superfamily. The aberrant function of these proteins leads to the development of tumors. Mentha longifolia var. asiatica and Zygophyllum arabicum have been known for their therapeutic potential for ages. The present study aimed to synthesize ZnO nanoparticles encapsulated with the extracts of M. longifolia var. asiatica and Z. arabicum and evaluating their therapeutic potential against breast cancer, targeting the Rab22A gene and its protein. UV-Vis spectrophotometer showed characteristic absorbance peaks at 295 nm and 345 nm for Z. arabicum and M. longifolia var. asiatica ZnONPs, respectively. The FTIR bands of Z. arabicum nanoparticles suggested the presence of aldehydes, alcohols, and polyols whereas bands of M. longifolia var. asiatica ZnONPs suggested the presence of carboxyl groups, hydroxyl groups, alkynes, and amines. SEM revealed the size of Z. arabicum ZnO NPs to be 25 ± 4 nm with a spherical shape as compared to nanoparticles of M. longifolia var. asiatica having a size of 35 ± 6 nm with a hexagonal shape. EDX determined the elemental composition of both particles. The cytotoxicity of both plant extracts and respective NPs was determined against the MCF-7 breast cancer cell line, which was found to be significant with an IC50 value of 51.68 μM for Z. arabicum and 88.02 μM for M. longifolia var. asiatica ZnO compared to plant extracts (64.01 μM and 107.9 μM for Z. arabicum and M. longifolia var. asiatica). The gene expression and protein levels of Rab22A were decreased in nanoparticle-treated cells as compared to the control group. The apoptotic role of synthesized nanoparticles against the MCF-7 cell line was also determined by the expression of apoptotic pathway genes and proteins (bax, caspase 3, caspase 8 and caspase 9). All samples showed significant apoptotic activity by activating intrinsic and extrinsic pathway genes. The activity of Z. arabicum was more eminent as compared to M. longifolia var. asiatica which was evident by the greater expression of studied genes and proteins as determined by Real-time qPCR and ELISA. This is the first-ever report describing the comparative analysis of the efficacy of Z. arabicum and M. longifolia var. asiatica ZnONPs against breast cancer.

In vitro anti-cancer potency of Mortierella elongata lipids against MCF 7 cells through induction of apoptosis and cell cycle arrest

Globally, cancer remains the second-whacking cause of mortality. Several studies, like in vitro cell studies, clinical trials, in vivo studies, and cohort studies, have authenticated the anticancer potency of omega-3 polyunsaturated fatty acids (PUFA) against various cancer types. The present study reports the in vitro anticancer potency of PUFA produced by Mortierella elongata (Accession No. OK402027) against Michigan Cancer Foundation-7 (MCF-7) breast cancer cell lines. The biocompatibility and cytotoxicity of M. elongata lipids were evaluated against human embryonic kidney and MCF-7 cells, respectively. The anti-proliferative activity of M. elongata lipids was examined at three different concentrations based on the inhibitory concentration (IC50) value. The apoptotic activity of M. elongata lipids was analyzed by fluorescence microscopy by implementing three different staining methods, such as acridine orange/ethidium bromide, 4,6-diamidino-2-phenylindole, and propidium iodide. Further, the biological activity of M. elongata on apoptosis induction, cell cycle progression, mitochondrial membrane potential (MMP), and reactive oxygen species (ROS) generation were evaluated. The MTT assay revealed the anti-proliferative activity of M. elongata lipids, and the IC50 value of M. elongata lipids at 24 h was found to be 28 ± 1.3 μg/mL. A significant decrease in the percentage of live cells with 57.36% of apoptotic and 14.10% of necrotic cells was revealed in cells treated with 100 μg/mL of M. elongata lipids. Anti-proliferative activity is associated with increased ROS generation and the loss of MMP. The lipids of M. elongata induced a dose-dependent G1 arrest and were found to be more effective at 100 μg/ml, accumulating 54.49% of MCF-7 cells in the G1 phase. Taken together, the present study has confirmed the in vitro anticancer potency of M. elongata lipids via apoptosis and cell cycle arrest.

Design and synthesis of novel multi-target tetrabromophthalimides as CBS and Topo-II inhibitors and DNA intercalators.

Microtubules are highly dynamic structures and constitute a crucial component of the cellular cytoskeleton. Besides, topoisomerases (Topo) play a fundamental role in maintaining the appropriate structure and organization of DNA. On the other hand, dual mechanism drug candidates for cancer treatment primarily aim to enhance the efficacy of cancer treatment and potentially overcome drug resistance. Hence, this work was tailored to design and synthesize new multi-target tetrabromophthalimide derivatives (2a-2k) that are capable of inhibiting the colchicine binding site (CBS) and topoisomerase II (Topo-II). The conducted in vitro studies showed that compound 2f showed the lowest IC50 value (6.7 μg mL-1) against the MDA-MB-468 cancer cell line. Additionally, compound 2f prompted upregulation of pro-apoptotic markers (caspases 3, 7, 8, and 9, Bax and p53). Moreover, some anti-apoptotic proteins (MMP2, MMP9, and BCL-2) were downregulated by compound 2f treatment. Besides, the colchicine binding assay showed that compounds 2f and 2k displayed promising inhibitory potential with IC50 values of 1.92 and 4.84 μg mL-1, respectively, in comparison with colchicine (1.55 μg mL-1). Furthermore, the Topo-II inhibition assay displayed the prominent inhibitory potential of compound 2f with an IC50 value of 15.75 μg mL-1, surpassing the IC50 of etoposide (20.82 μg mL-1). Cell cycle analysis revealed that compound 2f induced cell cycle arrest at both the G0-G1 and G2-M phases. The new candidates were docked against both the CBS (PDB ID: 5XIW) and Topo-II (PDB ID: 5CDP) targets to investigate their binding interactions and affinities as well. Accordingly, the synthesized compounds could serve as promising multi-target anticancer candidates with eligible apoptotic activity.

Mechanistic Insight into the Autophagic and Apoptotic Activity of Kaempferol on Liver Cancer Cells.

The accumulation of poorly folded protein in the endoplasmic reticulum (ER) promotes ER stress and contributes to the pathogenesis of hepatocellular carcinoma (HCC). Current therapies have various adverse effects, therefore, laying the need for an alternative approach. Kaempferol (KP), a naturally occurring flavonoid, possesses potent anti-proliferative properties against various cancer cells. Nevertheless, its involvement in HCC remains relatively unexplored, particularly regarding its influence on apoptosis and autophagy pathways. The effect of KP on cell viability, and motility of Hep3B cells was evaluated by MTT, and scratch assay, respectively. Hoechst staining and FACS analysis were done to check the effect of KP on apoptosis and cell cycle progression. qRTPCR was used to evaluate the expression of several apoptosis and autophagy-related genes. KP was docked with several ER stress-related proteins involved in HCC to gain further insights into molecular mechanisms. The results of docking studies were validated with MD simulation and in vitro studies. Treatment with KP at different time intervals showed dose- and time-dependent growth inhibition of liver cancer cells. KP decreased motility and arrested the cell cycle at the G0/G1 phase in Hep3B cells. Additionally, in the context of HCC, the relationship between KP, apoptosis, and autophagy is significant. It induced apoptosis and autophagy in Hep3B cells by downregulating the expression of Bcl-2 and upregulated Bax and Bid, Caspase-3, Beclin-1, and LC3. KP showed a better binding affinity with Nrf2, PERK, and IRE1α among all selected proteins. Further, it reversed the protective effect of 4-PBA (ER Stress inhibitor) by inducing apoptosis and autophagy in Hep3B cells. The study suggested KP as a potential chemopreventive agent for managing HCC by effectively inducing apoptosis and autophagy in Hep3B cells.

Biopreparation of cerium oxide nanoparticles using alginate: Characterization and estimation of antioxidant and its activity against breast cancer cell lines (MCF7)

Cerium is an important element whose various biomedical properties have been proven. The purpose of the current study was to fabricate cerium oxide nanoparticles (CeO-NPs) using alginate and to investigate their anti-oxidant properties and cytotoxicity against breast cancer cells and human skin fibroblasts. Cerium oxide nanoparticles were first synthesized using alginate, and their characteristics were then determined using XRD, FESEM, DLS, and FITR techniques. The antioxidant capacity of the prepared nanoparticles was measured using the DPPH biochemical method, and cytotoxicity was measured by the MTT method. Also, DAPI staining and annexin PI staining via flow cytometry were used to determine the type of cell death. Our findings indicated that the synthesized nanoparticles had IC50 values of 174, 32.5, and 16.07 µg/mL against the MCF-7 breast cancer cell line at 24, 48, and 72 h after treatment, respectively. Cell cycle assessment via flow cytometry revealed that the nanoparticles affected the ratio of the cells entering the apoptotic phase, supported by DAPI staining results showing the apoptotic activity of the synthesized nanoparticles. In conclusion, our results suggest that CeO-NPs can be considered as potential anti-cancer agents, and they are recommended to be further investigated in animal studies.

Multimodal layer-by-layer nanoparticles: a breakthrough in gene and drug delivery for osteosarcoma.

Osteosarcoma is one of the most common primary malignant bone tumours in children and adolescents, frequently arising from mesenchymal tissue in the distal femur. It is highly aggressive, often metastasising to the lungs. Current treatments, which include surgery combined with neoadjuvant chemotherapy and radiotherapy, are often unsatisfactory due to the inability of surgery to control metastasis and the side effects and drug resistance associated with chemotherapy. Thus, there is an urgent need for new treatment technologies. This study explored the use of nanoparticles for gene and drug delivery in osteosarcoma treatment. The nanoparticles were composed of biodegradable and biocompatible polymers, chitosan and PLGA, and were loaded with miRNA-34a, a short RNA molecule that functions as a tumour suppressor by inducing cell cycle arrest and apoptosis in osteosarcoma cells. Recognising that the co-delivery of multiple drugs can enhance treatment efficacy while reducing systemic toxicity and drug resistance, three additional classes of nanoparticles were developed by adding doxorubicin and resveratrol to the chitosan-PLGA-miRNA-34a core. A layer-by-layer technique was employed to create a bilayer nanocoating using pectin and chitosan as polyelectrolytes, for encapsulating the therapeutic payloads. The manufactured nanoparticles were tested on U2OS and Saos-2 cells to assess cell viability, metabolic activity, and morphology before and after treatment. Cells were treated in both two-dimensional cultures and three-dimensional osteosarcoma spheroids, creating a biomimetic cellular model. Increased apoptotic activity and disruption of cellular functions were primarily observed with nanoparticles co-delivering miRNA-34a and drugs, particularly those functionalised with the LbL nanocoating, as confirmed by PCR analysis.

The Effects of Tiger Milk Mushroom Lignosus rhinocerus TM02® (Agaricomycetes) on Leukemogenicity Tyrosine Kinase Cell Lines.

Leukemia can be a result of genetic changes associated with protein tyrosine kinase activity such as in MPL W515L and BCR/ABL genes. However, the current conventional treatment of leukemia produces severe side effects that urge the approach to use natural products. A medicinal mushroom, Lignosus rhinocerus shows potential as an anti-cancer treatment. To investigate the efficacy and mechanism of action of the L. rhinocerus cultivar (TM02®) extract on leukemogenic tyrosine kinase cell lines, a cold-water extract (CWE) was produced by using TM02® sclerotia powder at 4°C. The carbohydrate and protein contents were found to be 77.24% and 1.75% respectively. In comparison to the normal Ba/F3 cell, the CWE TM02® shows significant effects on exhibiting proliferation of Ba/F3 expressed MPL W515L and BCR/ABL, possibly due to the presence of phenolic compounds and antioxidant properties of TM02®, which contribute to act on various signaling pathways, and the reported apoptotic activity of CWE TM02®. In contrast, CWE TM02® significantly exhibited high scavenging activity of both Ba/F3 expressed MPL W515L and BCR/ABL. At concentrations of 125 μg/mL and 500 μg/mL of CWE TM02® decreased 49.5% and 67.5% of cell migration activity of Ba/F3 expressed MPL W515L and BCR/ABL respectively. Therefore, we postulate that CWE TM02® has the capability to mediate the migration route of the leukemogenic tyrosine kinase cell lines.

Novel Mitochondrial and DNA Damaging Fluorescent Calix[4]arenes Bearing Isatin Groups as Aromatase Inhibitors: Design, Synthesis and Anticancer Activity

Breast cancer causes a high rate of mortality all over the world. Therefore, the present study focuses on the anticancer activity of new lower rim-functionalized calix[4]arenes integrated with isatin and the p-position of calixarenes with 1,4-dimethylpyridinium iodine against various human cancer cells such as MCF-7 and MDA-MB-231 breast cancer cell lines, as well as the PNT1A healthy epithelial cell line. It was observed that compound 6c had the lowest values in MCF-7 (8.83 µM) and MDA-MB-231 (3.32 µM). Cell imaging and apoptotic activity studies were performed using confocal microscopy and flow cytometry, respectively. The confocal imaging studies with 6c showed that the compound easily entered the cell, and it was observed that 6c accumulated in the mitochondria.The Comet assay test was used to detect DNA damage of compounds in cells. It was found that treated cells had abnormal tail nuclei and damaged DNA structures compared with untreated cells. In vitro human aromatase enzyme inhibition profiles showed that compound 6c had a remarkable inhibitory effect on aromatase. Compound 6c displayed a significant inhibition capacity on aromatase enzyme with the IC50 value of 0.104 ± 0.004 µM. Thus, not only the anticancer activity of the new fluorescent derivatives, which are the subject of this study, but the aromatase inhibitory profiles have also been proven.