Treatment Of Cancer Cells Research Articles

In silico study of the impact of oxidation on pyruvate transmission across the hVDAC1 protein channel

The overexpression of voltage dependent anion channels (VDACs), particularly VDAC1, in cancer cells compared to normal cells, plays a crucial role in cancer cell metabolism, apoptosis regulation, and energy homeostasis. In this study, we used molecular dynamics (MD) simulations to investigate the effect of a low level of VDAC1 oxidation (induced e.g., by cold atmospheric plasma (CAP)) on the pyruvate (Pyr) uptake by VDAC1. Inhibiting Pyr uptake through VDAC1 can suppress cancer cell proliferation. Our primary target was to study the translocation of Pyr across the native and oxidized forms of hVDAC1, the human VDAC1. Specifically, we employed MD simulations to analyze the hVDAC1 structure by modifying certain cysteine residues to cysteic acids and methionine residues to methionine sulfoxides, which allowed us to investigate the effect of oxidation. Our results showed that the free energy barrier for Pyr translocation through the native and oxidized channel was approximately 4.3 ± 0.7 kJ mol−1 and 10.8 ± 1.8 kJ mol−1, respectively. An increase in barrier results in a decrease in rate of Pyr permeation through the oxidized channel. Thus, our results indicate that low levels of CAP oxidation reduce Pyr translocation, resulting in decreased cancer cell proliferation. Therefore, low levels of oxidation are likely sufficient to treat cancer cells given the inhibition of Pyr uptake.

H3K4me3 remodeling induced acquired resistance through O-GlcNAc transferase.

Drivers of the drug tolerant proliferative persister (DTPP) state have not been well investigated. Histone H3 lysine-4 trimethylation (H3K4me3), an active histone mark, might enable slow cycling drug tolerant persisters (DTP) to regain proliferative capacity. This study aimed to determine H3K4me3 transcriptionally active sites identifying a key regulator of DTPPs. Deploying a model of adaptive cancer drug tolerance, H3K4me3 ChIP-Seq data of DTPPs guided identification of top transcription factor binding motifs. These suggested involvement of O-linked N-acetylglucosamine transferase (OGT), which was confirmed by metabolomics analysis and biochemical assays. OGT impact on DTPPs and adaptive resistance was explored in vitro and in vivo. H3K4me3 remodeling was widespread in CPG island regions and DNA binding motifs associated with O-GlcNAc marked chromatin. Accordingly, we observed an upregulation of OGT, O-GlcNAc and its binding partner TET1 in chronically treated cancer cells. Inhibition of OGT led to loss of H3K4me3 and downregulation of genes contributing to drug resistance. Genetic ablation of OGT prevented acquired drug resistance in in vivo models. Upstream of OGT, we identified AMPK as an actionable target. AMPK activation by acetyl salicylic acid downregulated OGT with similar effects on delaying acquired resistance. Our findings uncover a fundamental mechanism of adaptive drug resistance that governs cancer cell reprogramming towards acquired drug resistance, a process that can be exploited to improve response duration and patient outcomes.

I-CBP112 declines overexpression of ATP-binding cassette transporters and sensitized drug-resistant MDA-MB-231 and A549 cell lines to chemotherapy drugs

Despite extensive efforts and ongoing progress in personalized anticancer approaches, chemotherapy remains the first line or the only treatment for some tumors that may develop resistance to chemotherapeutics in time due to inter alia overexpression of ATP-binding cassette transporters. Using clinically-relevant resistant models of triple negative breast cancer (MDA-MB-231; TNBC) as well as non-small cell lung cancer (A549; NSCLC), we tested the efficacy of I-CBP112 – CBP/EP300 bromodomain inhibitor to overcome drug resistance by declining ABC gene transcription. I-CBP112 significantly reduced ABCB1, ABCC1, ABCC2, ABCC3, ABCC5 and ABCG2 in all resistant lines, as well as ABCC10 in TNBC and ABCC4 in paclitaxel-resistant NSCLC, thereby increasing intracellular drug accumulation and cytotoxicity in 2D and 3D cultures. This was phenocopied only by the joint effect of ABC inhibitors such as tariquidar (ABCB1 – P-glycoprotein and ABCG2) and MK-571 (ABCC), whereas single inhibition of ABCB1/ABCG2 or ABCC proteins did not affect drug accumulation, thereby implying the need of simultaneous deficiency in activity of majority of drug pumps for enhanced drug retention. I-CBP112 failed to directly inhibit activity of ABCB1, ABCG2 and ABCC subfamily members at the same time. Importantly, I-CBP112 treated cancer cells polarized human macrophages into proinflammatory phenotypes. Moreover, I-CBP112 remained non-toxic to primary cell lines, nor did it enhance anticancer drug toxicity to blood-immune cells. In silico assay of ADMET properties confirmed the desired pharmacokinetic features of I-CBP112. The results suggest that the CBP/p300 inhibitor is a promising co-adjuvant to chemotherapy in drug-resistant cancer phenotypes, capable of decreasing ABC transporter expression.

Modulation of EGFR Activity by Molecularly Imprinted Polymer Nanoparticles Targeting Intracellular Epitopes.

In recent years, molecularly imprinted polymer nanoparticles (nanoMIPs) have proven to be an attractive alternative to antibodies in diagnostic and therapeutic applications. However, several key questions remain: how suitable are intracellular epitopes as targets for nanoMIP binding? And to what extent can protein function be modulated via targeting specific epitopes? To investigate this, three extracellular and three intracellular epitopes of epidermal growth factor receptor (EGFR) were used as templates for the synthesis of nanoMIPs which were then used to treat cancer cells with different expression levels of EGFR. It was observed that nanoMIPs imprinted with epitopes from the intracellular kinase domain and the extracellular ligand binding domain of EGFR caused cells to form large foci of EGFR sequestered away from the cell surface, caused a reduction in autophosphorylation, and demonstrated effects on cell viability. Collectively, this suggests that intracellular domain-targeting nanoMIPs can be a potential new tool for cancer therapy.

Hapten/Myristoyl Functionalized Poly(propyleneimine) Dendrimers as Potent Cell Surface Recruiters of Antibodies for Mediating Innate Immune Killing.

Recruiting endogenous antibodies to the surface of cancer cells using antibody-recruiting molecules has the potential to unleash innate immune effector killing mechanisms against antibody-bound cancer cells. The affinity of endogenous antibodies is relatively low, and many currently explored antibody-recruiting strategies rely on targeting over-expressed receptors, which have not yet been identified in most solid tumors. Here, both challenges are addressed by functionalizing poly(propyleneimine) (PPI) dendrimers with both multiple dinitrophenyl (DNP) motifs, as anti-hapten antibody-recruiting motifs, and myristoyl motifs, as universal phospholipid cell membrane anchoring motifs, to recruit anti-hapten antibodies to cell surfaces. By exploiting the multivalency of the ligand exposure on the dendrimer scaffold, it is demonstrated that dendrimers featuring ten myristoyl and six DNP motifs exhibit the highest antibody-recruiting capacity in vitro. Furthermore, it is shown that treating cancer cells with these dendrimers in vitro marks them for phagocytosis by macrophages in the presence of anti-hapten antibodies. As a proof-of-concept, it is shown that intratumoral injection of these dendrimers in vivo in tumor-bearing mice results in the recruitment of anti-DNP antibodies to the cell surface in the tumor microenvironment. These findings highlight the potential of dendrimers as a promising class of novel antibody-recruiting molecules for use in cancer immunotherapy.

Green synthesized silver nanoparticles using the plant-based reducing agent Matricaria chamomilla induce cell death in colorectal cancer cells.

There is a need to treat cancer cells with safe and natural nanoparticles to avoid the side effects of chemotherapeutic agents. Chamomile is considered a safe, natural plant with anticancer activity. We synthesize simple, inexpensive, and eco-friendly silver nanoparticles (SNs) using Chamomile (CHM) to tune their anticancer properties. SN-CHM was synthesized by reducing 1 mM silver nitrate aqueous solution in 100 mL with the aqueous ethanolic flower extract of CHM (18 mg/mL, w/v). The reaction proceeded overnight at 600 rpm and 28°C. SN-CHM was characterized for their % yield, average diameter, charge, morphology, and silver release. Moreover, SN-CHM was investigated for its antioxidant and anticancer activities at 200 µg/mL and 5 mg/ mL, respectively. A 59.12% yield and a uniform SN-CHM size of 115 ± 3.1 nm with a ζ-potential of -27.67 ± (-3.92) mv were observed. The UV-visible absorption showed shifts from 379.5 to 383.5 nm for CHM and SN-CHM, respectively. Moreover, Ag+ was ultimately released from SN-CHM after 5 h. Fourier Transform Infrared Spectroscopy (FT-IR) showed characteristic absorption peaks of CHM and produced SN-CHM. Furthermore, SN-CHM showed moderate antioxidant activity. SN-CHM inhibited the % viability of SW620 and HT-29 cell lines at 20 μM. SN-CHM may also greatly upregulate the apoptotic gene BAX while considerably downregulating the anti-apoptotic genes BCL2 and BCL-Xl. CHM can be a safe soft drink, especially when conjugated with Ag ions as anticancer NPs. SN-CHM is considered potent anticancer activity against SW620, and HT-29 cell lines.

Investigation of Hydrogen Peroxide for 45MV 5-Aminolevulinic Acid-Mediated Radiodynamic Therapy

Cytotoxicity caused by the reactive oxygen species (ROS), such as singlet oxygen species (1O2), superoxide radical (O2-), and hydroxyl radical (HO), is a mechanism for treating cancer cells in radiation therapy. 5-aminolevulinic acid (5-ALA)-mediated radiodynamic therapy (RDT) is more effective in killing tumor cells than conventional radiation therapy. ROS is produced not only by ionizing radiation but also by Cherenkov light-activated protoporphyrin IX (PpIX), which is metabolized endogenously from 5-ALA. Moreover, PpIX also catalyzes hydrogen peroxide to generate 1O2, and an enhanced catalytic yield of 1O2 was observed in X-ray irradiation in vitro. Therefore, using an in-vivo mouse model, this study aimed to investigate the effect of hydrogen peroxide as a coenzyme catalyst on a novel 45MV 5-ALA-mediated RDT. A subcutaneous C57BL/6 mouse model of KP1 cell line was used. The tumors (n = 240) were randomized into six groups, consisting of untreated, conventional radiation treatment (RT), and RDT with or without hydrogen peroxide: 1. control (untreated), 2. hydrogen peroxide, 3. 45MV RT, 4. 45MV RT + hydrogen peroxide, 5. 45MV RDT, 6. 45MV RDT + hydrogen peroxide. For 45MV photon irradiation, a single fraction of 4 Gy was delivered to the tumors. 5-ALA was systemically injected at 100 mg/kg by tail-vein 4 hours before the treatment for endogenous PpIX accumulation in the tumor. Carbamide peroxide was used to deliver hydrogen peroxide to tissue and was administered at 60 mg/kg intratumorally into tumors ∼3-5 min before the treatment. The treatment effect of a single fraction of treatment was measured by calculating tumor growth, measured using a 1.5 T MR scanner on the day of treatment (prior to the treatment), 3 and 7 days post-treatment. Two-way repeated ANOVA with Bonferroni correction was used to compare each treatment group to determine the statistically significant difference in tumor growth. A total of 45MV RDT with hydrogen peroxide was shown to significantly delay the tumor growth for the mouse model and cell line investigated in this work. 45MV RDT with hydrogen peroxide group resulted in a decrease in tumor growth by 51.3 ± 4.1 % and 56.1 ± 5.1 % compared to the control group on 3 and 7 days post-treatment, respectively (P<0.001), and 43.4 ± 0.8 % and 50.9 ± 0.8 % compared to 45MV RT alone on 3 and 7 days post-treatment, respectively. Moreover, the enhancement effect of hydrogen peroxide on 45MV RDT was 2.2-4.7 times greater on 45MV RT alone (P<0.05). Hydrogen peroxide did not contribute to tumor growth when administered alone. A total of 45MV 5-ALA-mediated RDT with hydrogen peroxide resulted in the most significant tumor growth delay compared to the other groups. The catalytic effect of PpIX and hydrogen peroxide was observed in-vivo. These preliminary results demonstrate an effective cancer treatment modality.

Downregulation of AP1S1 causes the lysosomal degradation of EGFR in non-small cell lung cancer.

Matrix stiffness has been shown to play a critical role in cancer progression by influencing various cellular processes, including epidermal growth factor (EGF) signaling. However, the underlying molecular mechanisms are not fully understood. Here, we investigated the role of adaptor-related protein complex 1 subunit sigma 1 (AP1S1), a component of adaptor protein complex-1, in the regulation of EGF receptor (EGFR) intracellular trafficking during cancer cell progression. We found that AP1S1 expression was upregulated under stiff matrix conditions, resulting in the regulation of EGFR trafficking in non-small cell lung adenocarcinoma cells. Knockout of AP1S1 caused the lysosomal degradation of EGFR, leading to suppressed EGF-induced anaplastic lymphoma receptor tyrosine kinase phosphorylation. In addition, the downregulation of AP1S1 increased the sensitivity of H1975 cancer cells, which are resistant to tyrosine kinase inhibitors, to erlotinib. Collectively, our results suggest that AP1S1 could regulate EGFR recycling under stiff matrix conditions, and AP1S1 inhibition could be a novel strategy for treating cancer cells resistant to EGFR-targeted anticancer drugs.

A STING pathway-activatable contrast agent for MRI-guided tumor immunoferroptosis synergistic therapy

The immunotherapy efficiency of stimulator of interferon genes (STING)-activatable drugs (e.g., 7-ethyl-10-hydroxycamptothecin, SN38) is limited by their non-specificity to tumor cells and the slow excretion of the DNA-containing exosomes from the treated cancer cells. The efficacy of tumor ferroptosis therapy is always limited by the elimination of lipid peroxides (LPO) by the pathways of glutathione peroxidase 4 (GPX4), dihydroorotate dehydrogenase (DHODH) and ferroptosis suppressor protein 1(FSP1). To solve these problems, in this study, we developed a STING pathway-activatable contrast agent (i.e., FeGd-HN@TA-Fe2+-SN38 nanoparticles) for magnetic resonance imaging (MRI)-guided tumor immunoferroptosis synergistic therapy. The remarkable in vivo MRI performance of FeGd-HN@TA-Fe2+-SN38 is attributed to its high accumulation at tumor location, the high relaxivities of FeGd-HN core, and the pH-sensitive TA-Fe2+-SN38 layer. The effectiveness and biosafety of the immunoferroptosis synergistic therapy induced by FeGd-HN@TA-Fe2+-SN38 are demonstrated by the in vivo investigations on the 4T1 tumor-bearing mice. The mechanisms of in vivo immunoferroptosis synergistic therapy by FeGd-HN@TA-Fe2+-SN38 are demonstrated by measurements of in vivo ROS, LPO, GPX4 and SLC7A11 levels, the intratumor matured DCs and CD8+ T cells, the protein expresion of STING and IRF-3, and the secretion of IFN-β and IFN-γ.

Cholesterol crystals induce mechanical trauma, inflammation, and neo-vascularization in solid cancers as in atherosclerosis

Background and aimsCancer and atherosclerosis share common risk factors and inflammatory pathways that promote their proliferation via vascular endothelial growth factor (VEGF). Because CCs cause mechanical injury and inflammation in atherosclerosis, we investigated their presence in solid cancers and their activation of IL-1β, VEGF, CD44, and Ubiquityl-Histone H2B (Ub-H2B), that promote cancer growth. MethodsTumor specimens from eleven different types of human cancers and atherosclerotic plaques were assessed for CCs, free cholesterol content and IL1-β by microscopy, immunohistochemistry, and biochemical analysis. Breast and colon cancer cell lines were cultured with and without CCs to select for expression of VEGF, CD44, and Ub-H2B. Western blot and immunofluorescence were performed on cells to assess the effect of CCs on signaling pathways. ResultsCancers displayed higher CC content (+2.29 ± 0.74 vs +1.46 ± 0.84, p < 0.0001), distribution (5.06 ± 3.13 vs 2.86 ± 2.18, p < 0.001) and free cholesterol (3.63 ± 4.02 vs 1.52 ± 0.56 μg/mg, p < 0.01) than cancer free marginal tissues and similarly for atherosclerotic plaques and margins (+2.31 ± 0.51 vs +1.44 ± 0.79, p < 0.02; 14.0 ± 5.74 vs 8.14 ± 5.52, p < 0.03; 0.19 ± 0.14 vs 0.09 ± 0.04 μg/mg, p < 0.02) respectively. Cancers displayed significantly increased expression of IL1-β compared to marginal tissues. CCs treated cancer cells had increased expression of VEGF, CD44, and Ub-H2B compared to control. By microscopy, CCs were found perforating cancer tumors similar to plaque rupture. ConclusionsThese findings suggest that CCs can induce trauma and activate cytokines that enhance cancer growth as in atherosclerosis.

Preparation and characterization of ionic metal-based anticancer formulations of the type [bis-DACH tolfenamate metal {Co(II), Cu(II) and Zn(II)}] complexes: Binding studies with ct-DNA/tRNA, cleavage and cytotoxic activity against chemoresistant cancer cells

A series of ionic metallo-NSAID’s having the formulae [Co(DACH)2(H2O)2](tolf)21, [Cu(DACH)2(H2O)2](tolf)22 and [Zn(DACH)2(H2O)2](tolf)23 where DACH and tolf− are 1,2-diaminocyclohexane and tolfenamate anion, respectively were synthesized. Complexes 1–3 were characterized by FT–IR, EPR, 1H/13C NMR, UV–vis and ESI–MS spectrometry methods. The bonding features as well as the electronic structure of all the complexes were optimized using DFT studies. Binding affinity of parent NSAID and 1–3 complexes with ct–DNA/tRNA were studied by using various complementary biophysical methods. The corroborative results of these experimental studies revealed that they were capable to interact with the therapeutic targets, ct–DNA/tRNA by the electrostatic interactions, showing higher preference for tRNA as compared to ct–DNA as quantified by their binding strength (Kb and Ksv values). It was also observed that the complexes exhibited much higher binding affinity than the parent free NSAID. A concentration-dependent cleavage of the complexes was observed with tRNA which promoted the hydrolysis of RNA. The cytotoxicity of 1–3 was evaluated against chemo-resistant phenotypes of cancers, viz., breast cancer cell line (MCF- 7), cervical cancer cell line (SiHa), triple-negative breast cancer cell (MDA-MB-231), oral squamous cancer cell (AW13516) and lung cancer cell line (Hop-62) by SRB assay. The cytotoxicity activity results displayed that among all the ionic tolfenamate metal complexes, complex 2 showed a much superior cytotoxic profile against treated cancer cell lines (SiHa, MDA-MB-231, AW13516, MCF-7 and Hop-62) with a good GI50 value of (4.7 ± 0.03) µg/ml, (10.5 ± 0.36) µg/ml, (19.2 ± 0.02) µg/ml, (4.1 ± 0.05) µg/ml, and (18.2 ± 0.04) µg/ml, respectively, while the complexes 1 and 3 showed moderate activity. Cell morphology studies were carried out in AW13516, MDA–MB–231, MCF–7 and Hop–62 cells with the incubation of complexes for a period of 48 h which implicated that the cells exposed to complex 2 presented round features and were nonadherent in culture. These morphological changes indicated that complex 2 was capable of inducing cell death, probably by apoptosis suggesting copper complex 2 was promising potent anticancer agent.

Palladium Nanocapsules for Photothermal Therapy in the Near-Infrared II Biological Window.

Recent developments in nanomaterials with programmable optical responses and their capacity to modulate the photothermal effect induced by an extrinsic source of light have elevated plasmonic photothermal therapy (PPTT) to the status of a favored treatment for a variety of malignancies. However, the low penetration depth of near-infrared-I (NIR-I) lights and the need to expose the human body to a high laser power density in PPTT have restricted its clinical translation for cancer therapy. Most nanostructures reported to date exhibit limited performance due to (i) activity only in the NIR-I region, (ii) the use of intense laser, (iii) need of large concentration of nanomaterials, or (iv) prolonged exposure times to achieve the optimal hyperthermia state for cancer phototherapy. To overcome these shortcomings in plasmonic nanomaterials, we report a bimetallic palladium nanocapsule (Pd Ncap)─with a solid gold bead as its core and a thin, perforated palladium shell─with extinction both in the NIR-I as well as the NIR-II region for PPTT applications toward cancer therapy. The Pd Ncap demonstrated exceptional photothermal stability with a photothermal conversion efficiency of ∼49% at the NIR-II (1064 nm) wavelength region at a very low laser power density of 0.5 W/cm2. The nanocapsules were further surface-functionalized with Herceptin (Pd Ncap-Her) to target the breast cancer cell line SK-BR-3 and exploited for in vitro PPTT applications using NIR-II light. Pd Ncap-Her caused more than 98% cell death at a concentration of just 50 μg/mL and a laser power density of 0.5 W/cm2 with an output power of only 100 mW. Flow cytometric and microscopic analyses revealed that Pd Ncap-Her-induced apoptosis in the treated cancer cells during PPTT. Additionally, Pd Ncaps were found to have reactive oxygen species (ROS) scavenging ability, which can potentially reduce the damage to cells or tissues from ROS produced during PPTT. Also, Pd Ncap demonstrated excellent in vivo biocompatibility and was highly efficient in photothermally ablating tumors in mice. With a high photothermal conversion and killing efficiency at very low nanoparticle concentrations and laser power densities, the current nanostructure can operate as an effective phototherapeutic agent for the treatment of different cancers with ROS-protecting ability.

Hyperpolarized 13C NMR for longitudinal in-cell metabolism using a mobile 3D cell culture system

Hyperpolarization with the dissolution dynamic nuclear polarization (dDNP) technique yields > 10,000-fold signal increases for NMR-active nuclei (e.g. 13C). Hyperpolarized 13C-labeled metabolic tracer molecules thus allow real-time observations of biochemical pathways in living cellular systems without interfering background. This methodology lends itself to the direct observation of altered intracellular reaction chemistry imparted for instance by drug treatment, infections, or other diseases. A reoccurring challenge for longitudinal cell studies of mammalian cells with NMR and dDNP-NMR is maintaining cell viability in the NMR spectrometer. 3D cell culture methods are increasing in popularity because they provide a physiologically more relevant environment compared to 2D cell cultures. Based on such strategies a mobile 3D culture system was devised. The clinical drug etoposide was used to treat cancer cells (HeLa) and the resulting altered metabolism was measured using hyperpolarized [1–13C]pyruvate. We show that sustaining the cell cultivation in cell incubators and only transferring the cells to the NMR spectrometer for the few minutes required for the dDNP-NMR measurements is an attractive alternative to cell maintenance in the NMR tube. High cell viability is sustained, and experimental throughput is many doubled.

Scrophularia amplexicaulis increases anti-cancer potential of doxorubicin in gastric cancer cells.

Increased drug resistance has reduced efficiency of chemotherapic drugs such as Doxorubicin (Dox). Scrophularia amplexicaulis (Scr) is one of the most important medicinal plants in Iran that has anti-cancer activity. The aim of this study was to investigate a novel approach to enhance therapeutic efficacy of Dox (as a chemotherapeutic agent) by co-administration of Scr (as a bioactive herbal compound) in gastric cancer treatment. In the present study, effects of Dox, Scr, and their combinations (Scr-Dox) were evaluated on viability and proliferation of two gastric cancer cell lines (AGS and MKN28). Moreover, morphological changes, invasion, migration, colony formation, and apoptosis rate in the treated cancer cells were evaluated. Expression of BAX, BCL2, SAMC, SURVIVIN, CASP9, P53, MMP9, and MMP2 in the treated cancer cells and untreated controls were evaluated by Real-Time PCR method. Treatments of cancer cells by Scr, Dox, and Scr-Dox significantly decreased proliferation, invasion, migration, and colony formation of gastric cancer cells. Treatments of cancer cells by Scr, Dox, and Scr-Dox significantly increased apoptosis rate as well as decreased cells mobility through modification of apoptosis- and metastasis-related genes expression. However, anti-cancer activity of Scr-Dox combination was significantly more than Scr and Dox treatments alone. In general, we demonstrated that Scr-Dox combination therapy exerts more profound anti-cancer effects on AGS and MKN28 cell lines than Scr and Dox monotherapy.

A novel EGFR-specific recombinant ricin-panitumumab (scFv) immunotoxin against breast and colorectal cancer cell lines; in silico and in vitro analyses

The Epidermal Growth Factor Receptor (EGFR) has been of high importance as it is over expressed in a wide diversity of epithelial cancers, promoting cell proliferation and survival pathways. Recombinant immunotoxins (ITs) have emerged as a promising targeted therapy for cancer treatment. In this study, we aimed to investigate the antitumor activity of a novel recombinant immunotoxin designed against EGFR. Using an in silico approach, we confirmed the stability of the RTA-scFv fusion protein. The immunotoxin was successfully cloned and expressed in the pET32a vector, and the purified protein was analyzed by electrophoresis and western blotting. In vitro evaluations were conducted to assess the biological activities of the recombinant proteins (RTA-scFv, RTA, scFv). The novel immunotoxin demonstrated significant anti-proliferative and pro-apoptotic effects against cancer cell lines. The MTT cytotoxicity assay revealed a decrease in cell viability in the treated cancer cell lines. Additionally, Annexin V/Propidium iodide staining followed by flow cytometry analysis showed a significant induction of apoptosis in the cancer cell lines, with half maximal inhibitory concentration (IC50) values of 81.71 nM for MDA-MB-468 and 145.2 nM for HCT116 cells (P < 0.05). Furthermore, the EGFR-specific immunotoxin exhibited non-allergenic properties. The recombinant protein demonstrated high affinity binding to EGFR. Overall, this study presents a promising strategy for the development of recombinant immunotoxins as potential candidates for the treatment of EGFR-expressing cancers.

Peptide‐Modified AIEgen Probe for Selective Imaging and Killing of Drug‐Resistant Cancer Cells Based on Cell‐Cycle‐Dependent Uptake

AbstractDistinguishing the precise phases of cells in the cell cycle is important for elucidation of their regulatory mechanisms and exploiting cell‐cycle‐targeted drugs. Here, a simple but efficient strategy is established based on a peptide‐modified aggregation‐induced emission luminogen (AIEgen) probe PKK–TTP by utilizing its differential cellular uptake amounts for specific cell cycle recognition and targeted therapy. After incubating the probe in cells with different phases, maximal fluorescence and reactive oxygen species (ROS) generation are observed in S phase, middle in G0/G1 phase, and the minimum in G2/M phase. The cell‐cycle‐dependent fluorescent intensity is proven to be related to variable uptake routes of PKK–TTP. The maximized uptake of PKK–TTP in S phase relies on three simultaneously working endocytic routes, including macro‐pinocytosis and clathrin‐ and caveolin‐dependent endocytosis, while decreased uptake in the G0/G1 and G2/M phases mainly depends on the single route. Furthermore, the uptake of PKK–TTP is further verified in normal cells and drug‐resistant cancer cells (S). Bright fluorescence of PKK–TTP is observed in drug‐resistant cancer cells in S phase, while a little fluorescence is observed in paclitaxel (PTX)‐treated cancer cells due to G2/M phase. Collectively, PKK–TTP can be applied to distinguish cancer cells, evaluating the drug resistance and exerting its therapeutic efficiency downstream.

Anti-proliferative Effect of Satureja sahandica Extraction by Co-administration of Layered Double Hydroxide (LDH) Nanosheets on HepG2 Hepatocellular Carcinoma Cell Line

Background: Hepatocellular carcinoma (HCC) is a common solid tumor and the third cause of cancer death worldwide. Resistance to chemotherapeutic drugs is a serious challenge in the treatment of HCC. To address this challenge, therefore, attempts have been made to develop novel anticancer agents by using medicinal plants and producing novel natural chemical compounds. Objectives: This study aimed to investigate the anti-proliferative effect of the Satureja sahandica extraction combined with layered double hydroxide (LDH) nanosheets on the HepG2 HCC cell line. Methods: The anti-proliferative effects of S. sahandica extraction and LDH nanosheets on HCC cell line HepG2 were investigated by adopting the MTT method. The morphological alterations, migration, and apoptosis rates were also evaluated in the treated cancer cells. The apoptosis-related genes mRNA expressions (BAX and BCL2) were examined by performing real-time PCR. Results: According to the study results, the S. sahandica extraction treatments significantly decreased the viability and proliferation of the HCC cell line HepG2. However, the anticancer activity of the S. sahandica extraction and LDH nanosheet combination was significantly less than its monotherapy. Various morphological alterations and apoptosis rates increased in the cancer cells. However, the migration rate of the cancer cells significantly decreased. Furthermore, the expressions of BAX and BCL2 genes were upregulated and downregulated, respectively. Conclusions: In sum, the significant anti-proliferative activity of the S. sahandica extraction against the HCC cells was documented, which may have been due to the induction of apoptosis. However, LDH nanosheets decreased the anti-proliferative activity of S. sahandica extraction.

Oleanolic acid increases the anticancer potency of doxorubicin in pancreatic cancer cells.

Combination therapy is a novel cancer therapy approach that combines two or more chemotherapy drugs. This treatment modality enhances the efficacy of chemotherapy by targeting key pathways in an additive or synergistic manner. Therefore, we investigated the efficacy of combination therapy by widely used chemotherapy drug doxorubicin (DOX) and oleanolic acid (OA) to induction of apoptosis for pancreatic cancer (PC) therapy.The effects of DOX, OA, and their combination (DOX-OA) were investigated on proliferation and viability of PC cell line (PANC-1) by MTT assay. Moreover, migration and invasion of the cancer cells were evaluated by trans-well migration assay and wound healing assay. Flow cytometry and DAPI (4',6-diamidino-2-phenylindole) staining were employed to investigate apoptosis quantification and qualification of the treated cancer cells. Finally, mRNA expression of apoptosis-related genes was assessed by quantitative real-time polymerase chain reaction.Our results demonstrated that the proliferation and metastasis potential of PC cells significantly decreased after treatment by DOX, OA, and DOX-OA. Moreover, we observed an increase in apoptosis percentage in the treated cancer cells. The apoptosis-related gene expression was modified to increase the apoptosis rate in all of the treatment groups. However, the anticancer potency of DOX-OA combination was significantly more than that ofDOX and OA treatments alone.Our study suggested that DOX-OA combination exerts more profound anticancer effects against PC cell lines than DOX or OA monotherapy. This approach may increase the efficiency of chemotherapy and reduce unintended side effects by lowering the prescribed dose of DOX.

New phosphoramides containing 2-amino-1,4-naphthaquinone moiety as anticancer and antibacterial agents: Experimental and theoretical evaluations

In this study, two novel phosphoramides containing 2-Amino-1,4-naphthaquinone, O,O-diethyl (2-naphthaleneyl-1,4-dione)phosphoramidothioate (NPT) and N,N´- bis(2-naphthaleneyl-1,4-dione) phenylphosphonic diamide (NPA) as cytotoxic agents, were synthesized and characterized by using spectroscopic methods. The anticancer activities of the synthesized compounds were investigated against the human breast cancer cell line SK-BR-3 and normal cells PC12 using MTT assay. The resulting data showed that NPA with an IC50 value of 6.384 μg/ml had a better inhibitory effect against cancer cells than NPT (IC50 = 10.08 μg/ml). According to the annexin V‐FITC/PI results, after treating cancer cells with synthesized compounds for 24 h, the overall population of apoptotic cells increased from 14.96% in the control group to 97.45%, 75.58% in NPA and NPT-treated cells, respectively. Given these results, NPA and NPT have considerable toxicity and apoptosis-inducing ability against SK-BR-3. The cytotoxicity results were supported by a molecular docking study to compute the interactions between synthesized compounds and transcription factor DNA (PDB ID: 1bg1). In order to know the structure-activity relationship (SAR) of synthesized compounds, the values of the quantum-mechanical descriptors were investigated computationally by the DFT method, and the obtained results were then compared with Cyclophosphamide (CY) as a standard anticancer drug. These results were in good accordance with those of in vitro investigation and molecular docking study. Antibacterial study of phosphoramides against Gram-negative (Escherichia coli) and Gram-positive bacteria (Staphylococcus aureus) was carried out MIC, MBC and DDA methods. Additionally, the molecular docking study was performed for the interaction between E. coli beta-lactamase (PDB: 4HBT) and S. aureus beta-lactamase (PDB: 1MWU) with target compounds, the obtained results showed that these compounds are appropriate ligands for antibacterial activity against Staphylococcus aureus and Escherichia coli bacteria.

Anticancer activity of lactoferrin-coated biosynthesized selenium nanoparticles for combating different human cancer cells via mediating apoptotic effects

The present study aims to develop a novel nanocombination with high selectivity against several invasive cancer cells, sparing normal cells and tissues. Bovine lactoferrin (bLF) has recently captured the interest of numerous medical fields owing to its biological activities and well-known immunomodulatory effects. BLF is an ideal protein to be encapsulated or adsorbed into selenium nanocomposites (Se NPs) in order to produce stable nanocombinations with potent anticancer effects and improved immunological functions. The biosynthesis of the functionalized Se NPs was achieved using Rhodotorula sp. strain MZ312359 via a simultaneous bio-reduction approach to selenium sodium salts. The physicochemical properties of Se NPs using SEM, TEM, FTIR, UV Vis, XRD, and EDX confirmed the formation of uniform agglomerated spheres with a size of 18–40 nm. Se NPs were successfully embedded in apo-LF (ALF), forming a novel nanocombination of ALF-Se NPs with a spherical shape and an average nanosize of less than 200 nm. The developed ALF-Se NPs significantly displayed an effective anti-proliferation efficiency against many cancer cells, including MCF-7, HepG-2, and Caco-2 cell lines, as compared to Se NPs and ALF in free forms. ALF-Se NPs showed a significant selectivity impact (> 64) against all treated cancer cells at IC50 63.10 ≤ μg/mL, as well as the strongest upregulation of p53 and suppression of Bcl-2, MMP-9, and VEGF genes. Besides, ALF-Se NPs were able to show the maximum activation of transcrition of key redox mediator (Nrf2) with suppression in reactive oxygen species (ROS) levels inside all treated cancer cells. This study demonstrates that this novel nanocombination of ALF-Se NPs has superior selectivity and apoptosis-mediating anticancer activity over free ALF or individual form of Se NPs.