Anticancer Research Articles

Physical, biochemical, and biological characterization of olive-derived lipid nanovesicles for drug delivery applications

AbstractExtracellular vesicles (EVs) have shown great promise as drug delivery system (DDS). However, their complex and costly production limit their development for clinical use. Interestingly, the plant kingdom can also produce EV-like nanovesicles that can easily be isolated and purified from a large quantity of raw material at a high yield. In this study, olive-derived nanovesicles (ODNVs) were isolated from raw fruits using serial centrifugations and their physical and biological features characterized to demonstrate their promising potential to be used as a DDS. Nanotracking particle analysis indicated an average size of 109.5 ± 3.0 nm and yield of 1012 ODNVs/mL for the purest fraction. Microscopy imaging, membrane fluidity assay and lipidomics analysis showed the presence of a rich lipid bilayer that significantly varied between different sources of ODNVs but showed a distinct signature compared to human EVs. Moreover, ODNVs were enriched in PEN1 and TET8 compared to raw fruits, suggesting an extracellular origin. Interestingly, ODNVs size and yield stayed unchanged after exposure to high temperature (70 °C for 1 h), wide pH range (5–10), and 50–100 nm extrusion, demonstrating high resistance to physical and chemical stresses. This high resistance allowed ODNVs to stay stable in water at 4 °C for a month, or with the addition of 25 mM trehalose for long-term freezing storage. Finally, ODNVs were internalized by both 2D and 3D cell culture without triggering significant cytotoxicity and immunogenicity. Importantly, the anticancer drug doxorubicin (dox) could be loaded by passive incubation within ODNVs and dox-loaded ODNVs decreased cell viability by 90% compared to only 70% for free dox at the same concentration, indicating a higher efficiency of drug delivery by ODNVs. In addition, this high cytotoxicity effect of dox-loaded ODNVs was shown to be stable after a 2-week storage at 4 °C. Together, these findings suggested that ODNVs represent a promising candidate as drug nanocarrier for various DDS clinical applications, as demonstrated by their biocompatibility, high resistance to stress, good stability in harsh environment, and improvement of anticancer drug efficacy.

Experimental and computational studies on possibility of using glucose diazacrown cryptand as a carrier for anticancer drugs busulfan and lomustine

AbstractDrug carriers play a very important role in pharmacy, especially in cancer therapy. Most drugs used in the treatment of cancer are characterized by poor solubility in water and lack of selectivity in their toxic effects on normal and cancer cells. Administration of the drug in the form of a complex with an appropriately selected carrier can significantly improve its therapeutic effect and reduce side effects. In this study, the possibility of using the cryptand L1, containing two diazacrown ethers and two saccharide groups, as a potential drug carrier is investigated. In order to determine whether it can form complexes with drugs, the cryptand L1 and its complexes with two anticancer drugs, busulfan (BSF) and lomustine (CCNU), were synthesized. Their selected structural and energetic properties were investigated using both experimental and computational methods. Additionally, water solubility and cytotoxicity tests were performed for all compounds. The measured 1H NMR spectra confirm that L1 forms complexes L1:BSF and L1:CCNU, the solubility of which in water appears to be much higher than that of the pure drugs. The results of DFT calculations made in water described with the implicit solvent model confirm high stability of L1:BSF and L1:CCNU and indicate that L1 forms with the drugs mainly non-inclusion complexes. However, additional tests with 20 H2O molecules explicitly included in the model suggest that both inclusion and non-inclusion forms can occur in a real solution. Cytotoxicity studies show that the macrocycle L1 is non-toxic towards both normal and cancer cells, and its complexes with drugs show greater selectivity towards cancer cells. Interestingly, while the cytotoxicity of the L1:BSF complex is stronger than that of pure BSF, the relationship is opposite in the case of L1:CCNU and CCNU. Therefore, L1 can be considered as a potential drug carrier, especially for those drugs that have weak activity on cancer cells.

Lycopene alleviates chemotherapy-induced mucositis via enhancing intestinal barrier function and attenuating TLR4/MyD88/NF-κB signaling cascade

5-Fluorouracil (5-FU) is one of the most common chemotherapeutic agents used in treating solid tumors. Lycopene is a natural exogenous carotenoid with antioxidant, anti-inflammatory, immunomodulatory, and anticancer activity. Intestinal mucositis is a commonly encountered complication for chemotherapy. However, effective or preventive treatment for this common complication is still need to be elucidated. The current study aimed to appraise the effect of lycopene on 5-FU-induced intestinal mucositis (FUIIM). Twenty-eight male rats were divided randomly and equally into four groups: control, lycopene, FUIIM, and 5-FU + lycopene groups. Parameters for mucosal barrier integrity, redox and inflammatory status, and apoptosis were immunoassayed in addition to histological assessment. Decreased body weight, significantly impaired redox status, increased caspase 3 level, toll-like receptor (TLR) 4 level, myd88 and NF-κB expression. However, decreased Bcl-2, and Ki-67 expression were detected in 5-FUIIM group. Furthermore, impairment in intestinal barrier integrity, represented by decreased Occludin and claudin1 levels with impaired histological features and ultrastructural features, were also detected. Meanwhile, lycopene co-administration protected against weight loss, heightened oxidative stress, apoptosis, inflammation, disturbed intestinal barrier integrity, and impaired histological and ultrastructure features. In conclusion, lycopene significantly improved intestinal mucositis, barrier integrity and gut toxicity induced by 5-FU in rats.

Synthesis of Self-Assembled Nanostructured Cisplatin Using the RESS Process

Background/Objectives: The primary goal of our research is to develop a process to prepare an aqueous dispersion of Cisplatin, an important anticancer drug, with increased solubility and storage stability. Method: In this context, we report the use of a customized RESS process for the synthesis of a novel, amber-colored and viscous aqueous cisplatin solution, an important anticancer drug, which we have denoted as “liquid” cisplatin. Results: Using specialized liquid cell in situ transmission electron microscopy (Liquid in situ TEM) and Raman spectroscopy, we demonstrated that “liquid” cisplatin comprises a bi-modal distribution of a highly solvated network of stable cisplatin nanoclusters in water and exhibited 27 times greater water solubility than standard cisplatin. More importantly, “liquid” cisplatin was stable at ambient conditions for over two years. Extensive analytical characterization of “liquid” cisplatin confirmed that it retained the original chemical identity of cisplatin. Cell viability and apoptosis studies on human lung adenocarcinoma A549 cells provided compelling evidence that “liquid” cisplatin demonstrated a more sustained anticancer effect compared to standard cisplatin. Conclusions: Aqueous cisplatin solubility was increased by 27X in the “liquid” cisplatin medium which retained its bio efficacy over a 2-year period. Our experimental results suggest the possibility of developing non-invasive and highly effective novel cisplatin drug-delivery platforms.

Preliminary phase results of GC-MS analysis and cytotoxic activity of Argemone mexicana leaves methanol extract against MCF-7 cell lines

Argemone mexicana leaves (AML) contain a variety of bioactive components such as fatty acids, amino acids, phenolics and alkaloids, including Berberine, Isoquinoline, Scoulerine, Stylopine, Thalifone, Benzylisoquinolines, Protopine, and Tetrahydroberberine. This study aimed to examine the gas chromatography and mass spectrometry (GC-MS) analysis of AML extract and its cytotoxic activity on MCF-7 cell lines to identify an alternative source for anticancer treatment. The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was utilized to assess the viability of AML extract-treated cells, while the apoptotic assay involved Acridine orange staining to determine the mode of cell death based on morphological and nuclear fragmentation. The GC-MS analysis was used to identify the biologically active compounds. The results showed that the percentage of cell viability after AML extract treatment ranged from 28.50% to 64.28% for concentrations of 1000 to 125 μg/ml (IC50 = 258.87 μg/ml). Statistical analysis revealed a significant difference in cell viability (p < 0.05). Exposure of cells to AML extract stimulated apoptosis through membrane blebbing, cell shrinkage, cytoplasm condensation, and nuclear fragmentation. Forty-one compounds were identified by GC-MS in the AML extract. Based on the peak area, some major compounds included n-Hexadecanoic acid (palmitic acid), 9,12-Octadecadienoic acid (Z, Z)-, Phytol, 9,12,15-Octadecatrienoic acid (Z, Z, Z)-, and 1-methyl piperidine. In conclusion, the bioactive compounds are apparently responsible for the cytotoxic effects of the AML extract, and the in-vitro results highlight the extract’s potential as a source of natural anticancer drugs. However, further investigation is necessary to confirm this.

A Model‐Based Trial Design With a Randomization Scheme Considering Pharmacokinetics Exposure for Dose Optimization in Oncology

ABSTRACTThe primary purpose of an oncology dose‐finding trial for novel anticancer agents has been shifting from determining the maximum tolerated dose to identifying an optimal dose (OD) that is tolerable and therapeutically beneficial for subjects in subsequent clinical trials. In 2022, the FDA Oncology Center of Excellence initiated Project Optimus to reform the paradigm of dose optimization and dose selection in oncology drug development and issued a draft guidance. The guidance suggests that dose‐finding trials include randomized dose–response cohorts of multiple doses and incorporate information on pharmacokinetics (PK) in addition to safety and efficacy data to select the OD. Furthermore, PK information could be a quick alternative to efficacy data to predict the minimum efficacious dose and decide the dose assignment. This article proposes a model‐based trial design for dose optimization with a randomization scheme based on PK outcomes in oncology. A simulation study shows that the proposed design has advantages compared to the other designs in the percentage of correct OD selection and the average number of patients assigned to OD in various realistic settings.

Streptomyces hygroscopicus and rapamycinicus Evaluated from a U.S. Marine Sanctuary: Biosynthetic Gene Clusters Encode Antibiotic and Chemotherapeutic Secondary Metabolites

Cancer remains a leading cause of death worldwide. Also threatening the public is the emergence of antibiotic resistance to existing medicines. Despite the challenge to produce viable natural products to market, there continues to be a need within public health to provide new chemotherapeutic drugs such as those exhibiting cytotoxicity and tumor cell growth-inhibitory properties. As marine genomic research advances, it is apparent that marine-derived sediment harbors uniquely potent bioactive compounds compared to their terrestrial counterparts. The Streptomyces genus in particular produces more than 30% of all secondary metabolites currently approved for human health, thus harboring unexplored reservoirs of chemotherapeutic and antibiotic agents to combat emerging disease. The present study identifies the presence of Streptomyces hygroscopicus and rapamycinicus in environmental sediment at locations within the U.S. Stellwagen Bank National Marine Sanctuary (SBNMS) from 2017 to 2022. Sequencing and bioinformatics methods catalogued biosynthetic gene clusters (BGCs) that drive cytotoxic and antibiotic biochemical processes in samples collected from sites permittable and protected to fishing activity. Poisson regression models confirmed that Sites 1 and 3 had significantly higher occurrences of rapamycinicus than other sites (p < 0.01). Poisson regression models confirmed that Sites 1, 2 and 3 had significantly higher occurrence for Streptomyces hygroscopicus across sites (p < 0.05). Interestingly, permitted fishing sites showed a greater prevalence of both species. Statistical analyses showed a significant difference in aligned hits with polyketide synthases (PKSs) and non-ribosomal peptide synthetases (NRPSs) by site and between species with hygroscopicus showing a greater quantity than rapamycinicus among Streptomyces spp. (p < 0.05; F = 4.7 > F crit).

4-(Pyrazolyl)benzenesulfonamide Ureas as Carbonic Anhydrases Inhibitors and Hypoxia-Mediated Chemo-Sensitizing Agents in Colorectal Cancer Cells.

Hypoxia in tumors contributes to chemotherapy resistance, worsened by acidosis driven by carbonic anhydrases (hCA IX and XII). Targeting these enzymes can mitigate acidosis, thus enhancing tumor sensitivity to cytotoxic drugs. Herein, novel 4-(pyrazolyl)benzenesulfonamide ureas (SH7a-t) were developed and evaluated for their inhibitory activity against hCA IX and XII. They showed promising results (hCA IX: KI = 15.9-67.6 nM, hCA XII: KI = 16.7-65.7 nM). Particularly, SH7s demonstrated outstanding activity (KIs = 15.9 nM for hCA IX and 55.2 nM for hCA XII) and minimal off-target kinase inhibition over a panel of 258 kinases. In NCI anticancer screening, SH7s exhibited broad-spectrum activity with an effective growth inhibition full panel GI50 (MG-MID) value of 3.5 μM and a subpanel GI50 (MG-MID) range of 2.4-6.3 μM. Furthermore, SH7s enhanced the efficacy of Taxol and 5-fluorouracil in cotreatment regimens under hypoxic conditions in HCT-116 colorectal cancer cells, indicating its potential as a promising anticancer agent.

Abstract C045: The extracellular matrix glycoprotein periostin supports chemotherapy resistance by modulating macrophage recruitment and phagocytosis in triple negative breast cancer

Abstract Triple-negative breast cancer (TNBC) accounts for approximately 15–20% of all breast cancer cases and is notoriously aggressive, often developing resistance to standard chemotherapy thus leading to a high rate of metastatic relapse. Evidence suggests that this resistance is linked to changes in the extracellular matrix (ECM) composition, particularly an increase in periostin (POSTN) expression, which is also associated with poor prognosis in breast cancer. Our recent publication showed that the ECM alone can influence macrophage phenotypes. Furthermore, another study from our lab has demonstrated that POSTN mediates resistance to anti-angiogenic therapy by recruiting macrophages in pancreatic neuroendocrine tumors. Notably, POSTN deletion reduced monocyte/macrophage recruitment and enhanced cytotoxic CD8+ T-cell infiltration. Based on these findings, we hypothesized that paclitaxel (PTX), a widely used chemotherapeutic drug for the management of TNBC, may stimulate POSTN production by fibroblasts leading to the recruitment of pro-tumoral macrophages that facilitate tumor growth resulting into therapy resistance. In this study, we showed that PTX induces increased POSTN expression in mouse models of TNBC through immunohistochemistry, and in both 2D and 3D in vitro models via interactions between TNBC cell lines and mammary fibroblasts, as demonstrated by immunofluorescence. An in vitro migration assay confirmed that PTX-induced POSTN enhances the recruitment of human monocytes and macrophages, which display a mixed immunosuppressive and inflammatory phenotype, as revealed by flow cytometry. Not only did these recruited macrophages exhibit a reduced phagocytic capacity against labelled TNBC cells in an in vitro phagocytosis assay, but we also found that POSTN plays a crucial role in inducing SIRPα expression, that may diminish macrophage phagocytic potential. Increased sialylation and Siglec-1 expression are known to be linked to poor prognosis and polarization towards a more immunosuppressive macrophage phenotype in TNBC. Interestingly in our model, we observed that higher concentrations of recombinant human POSTN induce increased Siglec-1 expression on macrophages, a receptor that interacts with sialic acids in the tumor ECM. Further work using a decellularized tissue model that preserves only the ECM from patient-derived tumor biopsies pre- and post-chemo, aims to further explore the correlation between PTX-induced POSTN, ECM sialylation, and Siglec-1 expression on macrophages. Additionally, we have developed POSTN-knockout murine cancer cells and fibroblasts using CRISPR/Cas9, allowing us to investigate how POSTN influences macrophage phenotypes and chemotherapy responses in vivo. Understanding the mechanisms behind chemotherapy resistance in TNBC is crucial for developing effective treatments for this aggressive cancer subtype. By elucidating the intricate interplay between POSTN, macrophages, and chemotherapy response, we aim to identify new therapeutic strategies to improve outcomes for TNBC patients. Citation Format: Ludovica Tarantola, Ioanna Keklikoglou, Oliver M. Pearce. The extracellular matrix glycoprotein periostin supports chemotherapy resistance by modulating macrophage recruitment and phagocytosis in triple negative breast cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr C045.

Abstract B003: In vitro 3D bone marrow niches differentially modulate survival, phenotype and drug responses of acute myeloid leukemia (AML) cells

Abstract Introduction: Acute myeloid leukemia (AML) is the deadliest type of hematopoietic cancer, with only about 50% of patients achieving overall survival [1]. This poor treatment outcome may be attributed to the significant heterogeneity of AML. Personalized cancer drug screening could be a solution, but primary AML cells undergo spontaneous apoptosis in ex vivo cultures [2], making personalized drug screening challenging. Research has indicated that stromal cells and extracellular matrix (ECM) create a pro-tumoral environment to support AML cells. We hypothesized that the bone marrow microenvironment mimicked by osteolineage cells encapsulated in ECM called 3D osteogenic niche (3DON) would support the survival of primary AML cells, and the presence of a biomimetic 3D cancer microenvironment would provide more reliable screening outcomes. Methods: Our laboratory has been developing a 3D microencapsulation platform using naturally occurring ECM to fabricate physiologically relevant and ECM-based 3D microtissues. 3D osteogenic niche (3DON) which mimics that in bone marrow is fabricated to support primary AML cell survival and phenotype maintenance in cultures. Specifically, 3DON derived from osteogenically differentiated mesenchymal stem cells (MSC) from healthy and AML donors are co-cultured with primary AML cells. Results: The AML cells co-cultured with the AML 3DON niche demonstrated improved viability, decreased apoptosis and preserved their CD33+ CD34− phenotype, which associated with the increased secretion of anti-apoptotic cytokines in the AML 3DON niche. In addition, AML cells under the AML 3DON niche displayed reduced sensitivity to FDA-approved chemotherapeutic drugs, doxorubicin and daunorubicin, while being more chemosensitive in the healthy 3DON niche. This further suggests the physiological relevance of the AML 3DON niche. Discussion and Conclusion: The AML 3DON maintained the viability and phenotypes of primary AML cells, suggesting that the AML 3DON environment may be more biomimetic and hence may achieve more reliable drug screening results. This study highlights the differential responses of AML cells to leukemic versus healthy bone marrow niches, suggesting the influence of native cancer cell niche in drug screening. It also suggests the potential of re-engineering healthy bone marrow niche to overcome AML chemoresistance.

A multicenter, phase II trial of triplet antiemetic therapy with palonosetron, aprepitant, and olanzapine for highly emetogenic chemotherapy in breast cancer (PATROL-II).

Dexamethasone is an antiemetic drug widely used to prevent nausea and vomiting caused by anticancer drugs. However, dexamethasone can cause several side effects even after short-term administration. Therefore, the development of dexamethasone-free antiemetic therapies has been recognized as an important challenge. The objective of this study was to investigate the efficacy and safety of palonosetron, aprepitant, and olanzapine. Patients who were chemotherapy-naïve and scheduled to receive highly emetogenic chemotherapy for breast cancer were enrolled and assessed for nausea and vomiting occurring within 120h after the start of chemotherapy. The primary endpoint was the total control (TC) rate of overall phases. Secondary endpoints included the complete response (CR) rate, which was evaluated during the acute, delayed, and overall phases. A total of 88 patients were enrolled from eight centers in Japan, of whom 84 were included in the analysis. The proportion of patients achieving TC throughout the overall period was 17.1%. Similarly, CR and CC rates for the overall period were 43.4% and 39.5%, respectively. Frequently reported adverse events were loss of appetite and constipation, with rates of 52.4% and 50.0%, respectively. The primary endpoint was not achieved. Therefore, antiemetic therapy without dexamethasone shows an inadequate effect on nausea, and it is generally advisable to avoid omitting dexamethasone. However, in the overall period, both CR and CC were comparable to conventional three-drug combination therapy. Thus, in patients unable to use dexamethasone, replacing it with olanzapine could be an option.Trial registration number: UMIN 000038644, November 20, 2019. The date of first trial registration: 13/03/2020.

Novel Dibenzoazepine-Substituted Triazole Hybrids as Cholinesterase and Carbonic Anhydrase Inhibitors and Anticancer Agents: Synthesis, Characterization, Biological Evaluation, and In Silico Studies

Novel Dibenzoazepine-Substituted Triazole Hybrids as Cholinesterase and Carbonic Anhydrase Inhibitors and Anticancer Agents: Synthesis, Characterization, Biological Evaluation, and <i>In Silico</i> Studies

Integrated proteomics and metabolomics analyses reveal new insights into the antitumor effects of valproic acid plus simvastatin combination in a prostate cancer xenograft model associated with downmodulation of YAP/TAZ signaling.

Despite advancements in therapeutic approaches, including taxane-based chemotherapy and androgen receptor-targeting agents, metastatic castration-resistant prostate cancer (mCRPC) remains an incurable tumor, highlighting the need for novel strategies that can target the complexities of this disease and bypass the development of drug resistance mechanisms. We previously demonstrated the synergistic antitumor interaction of valproic acid (VPA), an antiepileptic agent with histone deacetylase inhibitory activity, with the lipid-lowering drug simvastatin (SIM). This combination sensitizes mCRPC cells to docetaxel treatment both in vitro and in vivo by targeting the cancer stem cell compartment via mevalonate pathway/YAP axis modulation. Here, using a combined proteomic and metabolomic/lipidomic approach, we characterized tumor samples derived from 22Rv1 mCRPC cell-xenografted mice treated with or without VPA/SIM and performed an in-depth bioinformatics analysis. We confirmed the specific impact of VPA/SIM on the Hippo-YAP signaling pathway, which is functionally related to the modulation of cancer-related extracellular matrix biology and metabolic reprogramming, providing further insights into the molecular mechanism of the antitumor effects of VPA/SIM. In this study, we present an in-depth exploration of the potential to repurpose two generic, safe drugs for mCRPC treatment, valproic acid (VPA) and simvastatin (SIM), which already show antitumor efficacy in combination, primarily affecting the cancer stem cell compartment via MVP/YAP axis modulation. Bioinformatics analysis of the LC‒MS/MS and 1H‒NMR metabolomics/lipidomics results confirmed the specific impact of VPA/SIM on Hippo-YAP.

Unleashing the potential of Genistein and its derivatives as effective therapeutic agents for breast cancer treatment.

Breast cancer remains one of the leading causes of cancer-related deaths among women worldwide. Genistein (Gen), a phytoestrogen soy isoflavone, has emerged as a promising agent in the prevention and treatment of breast cancer due to its ability to function as a natural selective estrogen receptor modulator (SERM). This review explores the multifaceted mechanisms through which Gen and its derivatives exert their anticancer effects, including modulation of the PI3K/Akt signaling pathway, regulation of apoptosis, inhibition of angiogenesis, and impacts on DNA methylation and enzyme functions. We discuss the dual roles of Gen in both enhancing and inhibiting estrogen receptor (ER)-dependent pathways., highlighting its complex interactions with ERα and ERβ. Furthermore, the review examines the synergistic effect of combining Gen with conventional chemotherapeutic agents such as doxorubicin, cisplatin, and selenium, as well as other natural compounds like lycopene. Clinical studies suggest that while isoflavones may not significantly influence breast cancer progression in general, the high consumption of soy isoflavones is associated with reduced recurrence rates in breast cancer survivors. Importantly, Gen's ability to modulate key signaling pathways and enhance the efficacy of existing treatments improves its potential as a valuable adjunct in breast cancer therapy. In conclusion, Gen and its derivatives offer a novel and promising approach for treatment of breast cancer. Continued research into their mechanisms of action and clinical applications will be essential in optimizing their therapeutic potential and translating these findings into effective clinical interventions.

Site-Specific Quadruple-Functionalised Antibodies.

Antibody-drug conjugates (ADCs) are a growing class of chemotherapeutic agents that have yielded striking clinical successes. However, the efficacy of ADCs often suffers from issues associated with tumor heterogeneity and resistance. To overcome these problems, a new generation of ADCs comprising a single monoclonal antibody with multiple different payloads attached, termed multi-payload ADCs, have been developed. Here we deploy multiple orthogonal site-specific protein modification strategies to generate highly homogeneous multi-functionalised antibody conjugates comprising up to four different functionalities installed at four unique sites on the antibody. This work, which includes the use of a site-specific cyclopropenone (CPO)-based reagent, represents the first example of a homogeneous multi-payload ADC with a payload count greater than two, and thereby facilitates the development of the next generation of ADCs.

Site‐Specific Quadruple‐Functionalised Antibodies

AbstractAntibody–drug conjugates (ADCs) are a growing class of chemotherapeutic agents that have yielded striking clinical successes. However, the efficacy of ADCs often suffers from issues associated with tumor heterogeneity and resistance. To overcome these problems, a new generation of ADCs comprising a single monoclonal antibody with multiple different payloads attached, termed multi‐payload ADCs, have been developed. Here we deploy multiple orthogonal site‐specific protein modification strategies to generate highly homogeneous multi‐functionalised antibody conjugates comprising up to four different functionalities installed at four unique sites on the antibody. This work, which includes the use of a site‐specific cyclopropenone (CPO)‐based reagent, represents the first example of a homogeneous multi‐payload ADC with a payload count greater than two, and thereby facilitates the development of the next generation of ADCs.

Regulation of Apoptosis, Autophagy, and Metastasis by Luteolin in Human Bladder Cancer EJ138 Cells: An Experimental Study

Background: Chemotherapy remains a primary approach to cancer treatment, widely applied in bladder cancer (BC). However, the various side effects and resistance associated with chemotherapeutic drugs pose significant challenges in BC therapy, prompting interest in natural compounds like luteolin. Studies focus on its effects on key biological processes involved in BC, including metastasis, apoptosis, and autophagy. Objectives: This study investigated the regulation of mRNA expression of genes associated with apoptosis (BCL2, P53), autophagy (ULK1, ATG12), and metastasis (MMP2, MMP9) in malignant BC cells treated with luteolin. Methods: This was an in vitro experimental study. EJ138 BC cells were treated with various concentrations of luteolin, and its impact on cell viability, proliferation, and gene expression was assessed. The cytotoxic effect of luteolin on EJ138 BC cells was evaluated using the MTT assay after 24- and 48-hour treatments with different luteolin concentrations. Flow cytometry was performed to examine luteolin’s anti-proliferative effect, and RT-PCR was used to analyze mRNA expression of BCL2, P53, ULK1, ATG12, MMP2, and MMP9 genes. Results: MTT assay results confirmed that luteolin reduced the proliferation rate of BC cells. Flow cytometry indicated increased cell death in EJ138 BC cells following luteolin treatment. RT-PCR findings demonstrated that luteolin upregulated P53, ULK1, and ATG12 expression while downregulating BCL2 mRNA expression. However, luteolin treatment in EJ138 cells did not significantly alter MMP2 and MMP9 expression levels. Conclusions: These findings indicate that luteolin exerts cytotoxic effects on EJ138 BC cells by dysregulating mRNA expression of genes involved in apoptosis and autophagy. Therefore, luteolin shows potential as an effective anti-cancer agent for BC therapy.

Synthesis, Biological Activities, and Molecular Docking Studies of 15-Site Matrine Based Isatohydrazone Derivatives as Potential Anticancer Agents.

To acquire matrine derivatives with enhanced anticancer activity, we designed and synthesized twenty-one 15-site matrine based isatohydrazone derivatives were designed and synthesized. The anti-proliferative activity of all the compounds against human cervical cancer cells (HeLa), human colon cancer cells (HCT116), and non-small cell lung cancer cells (A549) was examined by the MTT method. The majority of the compounds exhibited superior anticancer activity compared to matrine. Among them, compound 5a displayed the most potent anti-proliferative activity, with IC50 values of 9.02±0.33 μM (HeLa), 10.49±1.09 μM (HCT116), and 15.23±0.12 μM (A549), respectively. Compound 5a can also induce cell cycle arrest in the G0/G1 phase and trigger apoptosis, as well as inhibit cell clonal formation and migration. Molecular docking experiments have demonstrated that compound 5a can form hydrogen bonds and hydrophobic interactions with EGFR-related protein 7AEI.

Unveiling the therapeutic potential of anthocyanin/cisplatin-loaded chitosan nanoparticles against breast and liver cancers

Abstract Background Liver and breast cancers are among the leading causes of cancer-related deaths worldwide, prompting researchers to seek natural anticancer agents and reduce chemotherapy side effects. Red beetroot (Beta vulgaris Linnaeus), rich in polyphenols and powerful antioxidants, has shown potential in cancer prevention. This study aimed to evaluate the impact of red beetroot-derived anthocyanin (Ant), Ant-loaded chitosan nanoparticles (Ant NPs), cisplatin (Cis), Cis-loaded chitosan (Cis NPs), and Cis + Ant-loaded chitosan NPs on human hepatoma HepG2 and breast adenocarcinoma MCF7 cell lines. Methods NPs preparation was evaluated by zeta potential, FTIR, and SEM. The cytotoxic, apoptotic, antioxidant, anti-inflammatory, anti-metastatic, and anti-angiogenic effects were assessed by MTT assay, qPCR, AO/EB staining, and flow cytometry. Results Treatment with Ant, Ant NPs, Cis, Cis NPs, and Cis + Ant NPs caused cytotoxicity in HepG2 and MCF7 with best effect in Cis-treated cells. The anticancer effects were attributed to mitochondrial-dependent apoptosis (with high Bax and low Bcl2 expression), chromatin disintegration, and cell cycle arrest in G2/M and S phases. All treatments inhibited migration by downregulating the migration-related gene MMP9 and upregulating the anti-migratory gene TIMP1 and decreased the angiogenesis-related gene VEGF and the inflammatory gene TNFα with best results in Cis NPs-treated cells. Interestingly, Ant, Ant NPs, and Cis + Ant NPs increased the antioxidant status (high GSH and upregulated expression of Nrf2 and OH-1) and decreased drug resistance-related MAPK1 and MDR1 genes compared to Cis and Cis NPs-treated cells. Conclusions Anthocyanin and cisplatin-loaded chitosan nanoparticles effectively combat breast and liver cancers by inducing cancer cell apoptosis, enhancing antioxidant defenses, and reducing inflammation. They also inhibit tumor spread and blood vessel formation through downregulation of MMP9 and VEGF, highlighting their therapeutic potential.

Pathology of doxorubicin-induced organopathies under different intravenous nicotinamide riboside administration modes

Doxorubicin (DOX) is a potent chemotherapeutic drug, but its clinical use is hindered by significant side effects on vital organs like the heart, kidneys, lungs, liver, and intestines. Currently, there is a lack of effective drugs that can provide simultaneous cardioprotection and organ protection during chemotherapy. Nicotinamide riboside (NR) holds promise as a pharmacological agent capable of offering comprehensive protection against the systemic toxicity caused by DOX. This study aimed to comprehensively evaluate the morphological characteristics of vital organs (heart, lungs, liver, kidneys) in Wistar rats with chronic doxorubicin-induced cardiomyopathy using various intravenous administration modes of NR as a protective agent. Sixty male SPF Wistar rats weighing 283 ± 22 g were divided into four groups: intact, control (DOX administered intraperitoneally), combined mode (the simultaneous use of DOX and NR) and preventive mode (the preliminary use of NR to realize the cumulative effect in cells, and its further joint use together with DOX) intravenous NR administration. Animal observation spanned two months after drug administration, followed by the collection of hearts, lungs, liver, and kidneys for morphological analysis. Echocardiographic assessment confirmed DOX cardiotoxicity. The study revealed that the hearts, kidneys and lungs exhibited more pronounced toxic effects of DOX compared to the liver. Both NR administration modes demonstrated protective effects, with the preventive regime showing the greatest efficacy in safeguarding vital organs.