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Extraction from the herbs was performed using the Soxhlet method. Various formula was synthesized for niosomes containing the extracts through thin film synthesis technique, and the most efficient formulation was selected. Afterwards, physicochemical properties of niosomes, including size, polydispersity index (PDI), zeta potential, morphology, encapsulation efficacy (EE%), extract release rate, non-interactive action between the extracts and niosomes, antibacterial potential, and cellular uptake were assessed. Finally, the toxicity level of the niosomes to breast cancer cells was compared and their impact on the expression of p53 and MCL-1 was evaluated. Our data demonstrated that the synthesized niosomes were sensitive to the temperature and pH. Also, the niosomes containing Hedera Helix extract (Nio-HHE) sized 97.7 nm, with a zeta potential of −19.9 ± 6.7 mV, PDI of 0.35, and 58 ± 2.4% encapsulation efficacy showed more toxicity to the cancer cells than the niosomes of Glycyrrhiza glabra extract (Nio-GGE) with the size of 111 ± 8.5 nm, zeta potential of −23.5 ± 4.5 mV, PDI of 0.113, and 69 ± 1.2% encapsulation efficacy. The former system proved to have more antibacterial potential, and affect the expression of the oncogenes more than the latter. Meanwhile, both niosomal systems demonstrated an acceptable cellular uptake, and no chemical interaction with the extracts was observed. Furthermore, useful function of the synthesized niosomes was confirmed by morphological assessments. Our data confirmed that encapsulation of herbal extracts improves their anticancer and antibacterial potential. We concluded that Nio-HHE has more significant antitumor effects on breast cancer cells than Nio-GGE. Consequently, applying nano drug delivery systems based on herbal therapy could mitigate the side effects resulting from chemotherapy and radiotherapy, and offer promising perspectives for treatment of breast cancer.

Breast cancer has been recognized as the most common cancer affecting women. Extremely low-frequency electromagnetic field (ELF-EMF) exposure can influence cellular activities such as cell-cell junctions and metastasis. However, more research is required to determine these fields' underlying mechanisms of action. Since cadherin switching is an important process during EMT (epithelial-mesenchymal transition), in this study, cadherin switching was regarded as one of the probable mechanisms of the effect of ELF-EMFs on metastasis suppression. For five days, breast cells received a 1 Hz, 100mT ELF-EMF (2 h/day). Cell invasion and migration were assessed in vitro by the Scratch wound healing assay and Transwell culture chambers. The expression of E- and N-cadherin was assessed using real-time PCR, western blotting, and Immunocytochemistry. ELF-EMF dramatically reduced the migration and invasion of MDA-MB 231 malignant cells compared to sham exposure, according to the results of the scratch test and the Transwell invasion test. The mRNA and protein expression levels of E-cadherin showed an increase, while the N-cadherin expression was found with a decrease, in MDA-MB231 cells receiving 1 Hz EMF compared to sham exposure. E-cadherin's mRNA and protein expression levels were enhanced in MCF10A cells receiving 1 Hz EMF compared to sham exposure. ELF-EMF can be used as a method for the multifaceted treatments of invasive breast cancer.

Pathogenic mutations in BRCA1 are associated with an increased risk of hereditary breast, ovarian, and some other cancers; however, the clinical significance of many mutations in this gene remains unknown (Variants of Unknown Significance/VUS). Since mutations in intolerant regions of a protein lead to dysfunction and pathogenicity, identifying these regions helps to predict the clinical importance of VUSs. This study aimed to identify intolerant regions of BRCA1 and understand the possible root of this susceptibility. Intolerant regions appear to carry more pathogenic mutations than expected due to their lower tolerance to missense variations. Therefore, we hypothesized that among the BRCA1 regions, the higher the mutation density, the greater the intolerance. Thus, pathogenic mutation density and regional intolerance scores were calculated to identify BRCA1-intolerant regions. To investigate the pathogenic mechanisms of missense-intolerant regions in BRCA1 , transcription activation (TA) experiments and molecular dynamics (MD) simulations were also performed. The results showed that the RING domain, followed by the BRCT domain, has the highest density of pathogenic mutations. In the BRCT domain, a higher density of pathogenic mutations was observed in the inter-BRCT linker. Additionally, scores generated by Missense Tolerance Ratio-3D (MTR3D) and the Missense Tolerance Ratio consensus (MTRX) showed that the inter-BRCT linker is more intolerant than other regions of the BRCT domain. The MD results showed that mutations in the inter-BRCT linker led to cancer susceptibility, likely due to disruption of the interaction between BRCA1 and phosphopeptides. TA laboratory assays further supported the importance of the inter-BRCT linker.Communicated by Ramaswamy H. Sarma.

Objectives: Among the most promising antibody formats in terms of inhibiting carcinogenesis are single-stranded variable fragments, whose targeted binding to the Fzd7 receptor has been proven effective at suppressing tumorigenesis. In this study, we investigated the effectiveness of an anti-Fzd7 antibody fragment against both tumor growth and metastasis of breast cancer cells., Methods: To develop anti-Fzd7 antibodies, bioinformatics approaches were used and the antibodies were expressed recombinantly in E. coli BL21 (DE3). The expression of anti-Fzd7 fragments was verified by Western blotting. Analysis of the antibody's binding capacity to Fzd7 was conducted by flow cytometry. Cell death and apoptosis were assessed by MTT and Annexin V/PI assays. The transwell migration and invasion assays, as well as the scratch method, were used to evaluate cell motility and invasiveness., Results: The anti-Fzd7 antibody was expressed successfully as a single band of 31 kDa. It could bind to 21.5% of MDA-MB-231 cells, as opposed to only 0.54% of SKBR-3 cells as negative control. According to MTT assay, induced apoptosis was 73.7% in MDA-MB-231 cells, compared with 29.5% in SKBR-3 cells. Also, the antibody exerted a significant inhibitory effect of 76% and 58% on migration and invasion of MDA-MB-231 cells, respectively., Conclusion: The recombinantly developed anti-Fzd7 scFv of this study could exhibit significant antiproliferative and antimigratory properties, along with a high apoptosis-inducing potential, making it suitable for the immunotherapy of triple negative breast cancer., (© 2023. The Author(s), under exclusive licence to Federación de Sociedades Españolas de Oncología (FESEO).)

Introduction: One of the most pressing concerns associated with breast cancer-targeted therapies is resistance to Tamoxifen and Herceptin. Such drug resistance is usually characterized by reduced expression of certain cell surface receptors. Some biological regimens can induce perceptible overexpression of these receptors in favor of drug responsiveness., Material and Methods: In this research, drug-responsive MCF-7 and SKBR-3, along with drug-resistant MCF-7R (Tamoxifen resistant) and JIMT-1 (Herceptin resistant) breast cancer cell lines in 2D and 3D cultures were exposed to anti-MUC1 nanobody and then assessed for their ER, PR, and HER2 gene and protein expression using qRT-PCR and immunofluorescent staining analyses. Cell viability and the synergistic relationships of combination treatments were determined with MTT assay followed by CompuSyn software. Apoptotic cells were evaluated with Annexin V/propidium Iodide (PI) and acridine orange/ethidium bromide (AO/EB) staining., Results: Anti-MUC1 exposure elevated the expression levels of ER (42 folds), PR (18.5 folds), and HER2 (4.7 folds). As a result of co-treatment, the IC 50 levels for Tamoxifen and Herceptin were reduced by up to 10 and 3 folds, respectively. MCF-7R cells responded positively to Tamoxifen, as evidenced by a 5-fold reduction in the IC 50 and enhanced apoptosis., Conclusion: The ER, PR, and HER2 overexpression after MUC1 blocking could signal drug hypersensitization and facilitate drug resistance management., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Background: Trastuzumab is a humanized monoclonal antibody that targets site-specifically human epidermal growth factor-2 receptor (HER2) cell surface antigen overexpressed in approximately 20% of human breast carcinomas. Despite its positive therapeutic outcomes, a large proportion of individuals are unresponsive to the treatment with the trastuzumab or develop resistance to it., Objective: To evaluate a chemically synthesized trastuzumab-based antibody-drug conjugate (ADC) to improve the trastuzumab therapeutic index., Methods: The current study explored the physiochemical characteristics of the trastuzumab conjugated to a cytotoxic chemotherapy agent DM1 via Succinimidyl 4-(N-maleimidomethyl) cyclohexane-1-carboxylate (SMCC) linker, created in our earlier study, using SDS-PAGE, UV/VIS, and RP-HPLC analyses. The antitumor effects of the ADCs were analyzed using MDA-MB-231 (HER2-negative) and SK-BR-3 (HER2-positive) cell lines utilizing in vitro cytotoxicity, viability, and binding assays. Three different formats of a HER2-targeting agent: trastuzumab, synthesized trastuzumab-MCC-DM1, and commercially available drug T-DM1 (Kadcyla®) were compared., Results: UV-VIS spectroscopic analysis showed that the trastuzumab-MCC-DM1 conjugates, on average, entailed 2.9 DM1 payloads per trastuzumab. A free drug level of 2.5% was determined by RP-HPLC. The conjugate appeared as two bands on a reducing SDS-PAGE gel. MTT viability assay showed that conjugating trastuzumab with DM1 significantly improved the antiproliferative effects of this antibody in vitro. Importantly, the evaluations using LDH release and cell apoptosis assays confirmed that trastuzumab maintains its ability to induce cell death response while conjugating with the DM1. The binding efficiency of trastuzumab-MCC-DM1 was comparable to that of the naked trastuzumab., Conclusion: Trastuzumab-MCC-DM1 was found effective against HER2+ tumors. The potency of this synthesized conjugate brings it closer to the commercially available T-DM1.

Background: The utilization of monoclonal antibodies (moAbs), an issue correlated with the biopharmaceutical professions, is developing and maturing. Coordinated with this conception, we produced the appealingly modeled anti-EpCAM scFv for breast cancer tumors., Methods: Afterward cloning and expression of recombinant antibody in Escherichia coli bacteria, the correctness of the desired antibody was checked by western blotting. Flow cytometry was utilized to determine the capacity of the recombinant antibody to append to the desired receptors in the malignant breast cancer (BC)cell line. The recombinant antibody (anti-EpCAM scFv) was examined for preclinical efficacy in reducing tumor growth, angiogenesis, and invasiveness (in vitro- in vivo)., Findings: A target antibody-mediated attenuation of migration and invasion in the examined cancer cell lines was substantiated (P-value < 0.05). Grafted tumors from breast cancer in mice indicated significant and compelling suppression of tumor growth and decrement in blood supply in reaction to the recombinant anti-EpCAM intervention. Evaluations of immunohistochemical and histopathological findings revealed an enhanced response rate to the treatment., Conclusion: The desired anti-EpCAM scFv can be a therapeutic tool to reduce invasion and proliferation in malignant breast cancer., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The usage of monoclonal antibodies (mAbs) and antibody fragments, as a matter associated with the biopharmaceutical industry, is increasingly growing. Harmonious with this concept, we designed an exclusive modeled single-chain variable fragment (scFv) against mesenchymal-epithelial transition (MET) oncoprotein. This scFv was newly developed from Onartuzumab sequence by gene cloning, and expression using bacterial host. Herein, we examined its preclinical efficacy for the reduction of tumor growth, invasiveness and angiogenesis in vitro and in vivo. Expressed anti-MET scFv demonstrated high binding capacity (48.8%) toward MET-overexpressing cancer cells. The IC 50 value of anti-MET scFv against MET-positive human breast cancer cell line (MDA-MB-435) was 8.4 µg/ml whereas this value was measured as 47.8 µg/ml in MET-negative cell line BT-483. Similar concentrations could also effectively induce apoptosis in MDA-MB-435 cancer cells. Moreover, this antibody fragment could reduce migration and invasion in MDA-MB-435 cells. Grafted breast tumors in Balb/c mice showed significant tumor growth suppression as well as reduction of blood-supply in response to recombinant anti-MET treatment. Histopathology and immunohistochemical assessments revealed higher rate of response to therapy. In our study, we designed and synthetized a novel anti-MET scFv which could effectively suppress MET-overexpressing breast cancer tumors., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Safranal, as an aroma in saffron, is one of the cytotoxic compounds in saffron that causes cell death in triple-negative breast cancer cells. Our recent research reported the anti-cancer effects of safranal, which further demonstrated its impact on protein translation, mitochondrial dysfunction, and DNA fragmentation. To better understand the underlying mechanisms, we identified acetylated and phosphorylated peptides in safranal-treated cancer cells. We conducted a comprehensive phosphoproteomics and acetylomics analysis of safranal-treated MDA-MB-231 cells by using a combination of TMT labeling and enrichment methods including titanium dioxide and immunoprecipitation. We provide a wide range of phosphoproteome regulation in different signaling pathways that are disrupted by safranal treatment. Safranal influences the phosphorylation level on proteins involved in DNA replication and repair, translation, and EGFR activation/accumulation, which can lead the cells into apoptosis. Safranal causes DNA damage which is followed by the activation of cell cycle checkpoints for DNA repair. Over time, checkpoints and DNA repair are inhibited and cells are under a mitotic catastrophe. Moreover, safranal prevents repair by the hypo-acetylation of H4 and facilitates the transcription of proapoptotic genes by hyper-acetylation of H3, which push the cells to the brink of death.

Although predicting the effects of variants near intron-exon boundaries is relatively straightforward, predicting the functional Exon Splicing Enhancers (ESEs) and the possible effects of variants within ESEs remains a challenge. Considering the essential role of CYP2D6/CYP2C19 genes in drug metabolism, we attempted to identify variants that are most likely to disrupt splicing through their effect on these ESEs. ESEs were predicted in these two genes using ESEfinder 3.0, incorporating a series of filters (increased threshold and evolutionary conservation). Finally, reported mutations were evaluated for their potential to disrupt splicing by affecting these ESEs. Initially, 169 and 243 ESEs were predicted for CYP2C19/CYP2D6, respectively. However, applying the filters, the number of predicted ESEs was reduced to 26 and 19 in CYP2C19/CYP2D6, respectively. Comparing prioritized predicted ESEs with known sequence variants in CYP2C19/CYP2D6 genes highlights 18 variations within conserved ESEs for each gene. We found good agreement in cases where such predictions could be compared to experimental evidence. In total, we prioritized a subset of mutational changes in CYP2C19/CYP2D6 genes that may affect the function of these genes and lead to altered drug responses. Clinical studies and functional analysis for investigating detailed functional consequences of the mentioned mutations and their phenotypic outcomes is mostly recommended., (© 2022. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

As an intelligent disease, tumors apply several pathways to evade the immune system. It can use alternative routes to bypass intracellular signaling pathways, such as nuclear factor-κB (NF-κB), Wnt, and mitogen-activated protein (MAP)/phosphoinositide 3-kinase (PI3K)/mammalian target of rapamycin (mTOR). Therefore, these mechanisms lead to therapeutic resistance in cancer. Also, these pathways play important roles in the proliferation, survival, migration, and invasion of cells. In most cancers, these signaling pathways are overactivated, caused by mutation, overexpression, etc. Since numerous molecules share these signaling pathways, the identification of key molecules is crucial to achieve favorable consequences in cancer therapy. One of the key molecules is the mesenchymal-epithelial transition factor (MET; c-Met) and its ligand hepatocyte growth factor (HGF). Another molecule is the epithelial cell adhesion molecule (EpCAM), which its binding is hemophilic. Although both of them are involved in many physiologic processes (especially in embryonic stages), in some cancers, they are overexpressed on epithelial cells. Since they share intracellular pathways, targeting them simultaneously may inhibit substitute pathways that tumor uses to evade the immune system and resistant to therapeutic agents., (© 2022. The Author(s).)

Extracellular matrix (ECM) refers to the non-cellular components of the tumour microenvironment, fundamentally providing a supportive scaffold for cellular anchorage and transducing signaling cues that orchestrate cellular behaviour and function. The ECM integrity is abrogated in several cases of cancer, ending in aberrant activation of a number of mechanotransduction pathways and induction of multiple tumorigenic events such as extended proliferation, cell death resistance, epithelial-mesenchymal transition and most importantly the development of chemoresistance. In this regard, the present study mainly aims to elucidate how the ECM-stiffening process may contribute to the development of chemoresistance during cancer progression and what pharmacological approaches are required for tackling this issue. Hence, the first section of this review explains the process of ECM stiffening and the ways it may affect biochemical pathways to induce chemoresistance in a clinic. In addition, the second part focuses on describing some of the most important pharmacological agents capable of targeting ECM components and underlying pathways for overcoming ECM-induced chemoresistance. Finally, the third part discusses the obtained results from the application of these agents in the clinic for overcoming chemoresistance., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

BRCA1 and BRCA2 are two prominent genes that account for about 20-40% of inherited breast cancer. Mutations in these genes are often associated with clustering of especially early-onset cancers in the family. The spectrum of BRCA variants showed a significant difference between geographic regions and ethnicities. The frequency and spectrum of BRCA mutations in Iran, a country in southwest Asia, have not yet been thoroughly studied. Here, for the first time, all published and not published BRCA pathogenic variants are presented. Among 1040 high risk families (1258 cases) which were detected, 116 families were found to carry pathogenic variants in either BRCA1 or BRCA2. Altogether 89 distinct types of pathogenic variants have been detected in Iran, including 41 in BRCA1 and 48 in BRCA2. 16 out of 89 mutations had not been previously reported in Iran and are presented for the first time in this article, among which 4 mutations are novel worldwide. 20% of families had one of the seven most commonly observed mutations, including c.81-1G > C, c.66&#95;67delAG, c.4609C>T, c.1568delT, c.1961delA, in BRCA1 and: c.3751&#95;3752insA, c.8585dupT in BRCA2. Combining the data from published articles and our study which has not been published before, a comprehensive table is created as a reference for entire BRCA pathogenic variants and their frequencies in Iran., (© 2021. The Author(s), under exclusive licence to Springer Nature B.V.)

Although resistance is its major obstacle in cancer therapy, trastuzumab is the most successful agent in treating epidermal growth factor receptor 2 positive (HER2 +) breast cancer (BC). Some patients show resistance to trastuzumab, and scientists want to circumvent this problem. This review elaborately discusses possible resistance mechanisms to trastuzumab and introduces mucin 1 (MUC1) as a potential target efficient for overcoming such resistance. MUC1 belongs to the mucin family, playing the oncogenic/mitogenic roles in cancer cells and interacting with several other oncogenic receptors and pathways, such as HER2, β-catenin, NF-κB, and estrogen receptor (ERα). Besides, it has been established that MUC1- Cytoplasmic Domain (MUC1-CD) accelerates the development of resistance to trastuzumab and that silencing MUC1-C proto-oncogene is associated with increased sensitivity of HER2 + cells to trastuzumab-induced growth inhibitors. We mention why targeting MUC1 can be useful in overcoming trastuzumab resistance in cancer therapy., (© 2022. The Author(s).)

Mucin-1 (MUC-1) is a transmembrane glycoprotein, which bears many similarities between dogs and humans. Since the existence of animal models is essential to understand the significant factors involved in breast cancer mechanisms, canine mammary tumors (CMTs) could be used as a spontaneously occurring tumor model for human studies. Accordingly, this review assessed the comparison of canine and human MUC-1 based on their diagnostic and therapeutic aspects and showed how comparative oncology approaches could provide insights into translating pre-clinical trials from human to veterinary oncology and vice versa which could benefit both humans and dogs., (© 2022. The Author(s).)

Alteration in glycosylation pattern of MUC1 mucin tandem repeats during carcinomas has been shown to negatively affect adhesive properties of malignant cells and enhance tumor invasiveness and metastasis. In addition, MUC1 overexpression is closely interrelated with angiogenesis, making it a great target for immunotherapy. Alongside, easier interaction of nanobodies (single-domain antibodies) with their antigens, compared to conventional antibodies, is usually associated with superior desirable results. Herein, we evaluated the preclinical efficacy of a recombinant nanobody against MUC1 tandem repeats in suppressing tumor growth, angiogenesis, invasion, and metastasis. Expressed nanobody demonstrated specificity only toward MUC1-overexpressing cancer cells and could internalize in cancer cell lines. The IC50 values (the concentration at which the nanobody exerted half of its maximal inhibitory effect) of the anti-MUC1 nanobody against MUC1-positive human cancer cell lines ranged from 1.2 to 14.3 nm. Similar concentrations could also effectively induce apoptosis in MUC1-positive cancer cells but not in normal cells or MUC1-negative human cancer cells. Immunohistochemical staining of spontaneously developed mouse breast tumors prior to in vivo studies confirmed cross-reactivity of nanobody with mouse MUC1 despite large structural dissimilarities between mouse and human MUC1 tandem repeats. In vivo, a dose of 3 µg nanobody per gram of body weight in tumor-bearing mice could attenuate tumor progression and suppress excessive circulating levels of IL-1a, IL-2, IL-10, IL-12, and IL-17A pro-inflammatory cytokines. Also, a significant decline in expression of Ki-67, MMP9, and VEGFR2 biomarkers, as well as vasculogenesis, was evident in immunohistochemically stained tumor sections of anti-MUC1 nanobody-treated mice. In conclusion, the anti-MUC1 tandem repeat nanobody of the present study could effectively overcome tumor growth, invasion, and metastasis., (© 2021 The Authors. Molecular Oncology published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV2), causing the 2019 novel coronavirus disease (COVID-19), was introduced by WHO (World Health Organization) as "pandemic" in March 2020. According to WHO, thus far (23 November 2020) 58,425,681 infected cases including 1,385,218 deaths have been reported worldwide. In order to reduce transmission and spread of this lethal virus, attempts are globally being made to develop an appropriate vaccine. Intending to neutralize pathogens at their initial entrance site, protective mucosal immunity is inevitably required. In SARS-CoV2 infection and transmission, respiratory mucosa plays a key role; hence, apparently mucosal vaccination could be a superior approach to elicit mucosal and systemic immune responses simultaneously. In this review, the advantages of mucosal vaccination to control COVID-19 infection, limitations, and outcomes of mucosal vaccines have been highlighted. Considering the gut microbiota dysregulation in COVID-19, we further provide evidences on utilization of recombinant probiotics, particularly lactic acid bacteria (LAB) as vaccine carrier. Their intrinsic immunomodulatory features, natural adjuvanticity, and feasible expression of relevant antigen in the mucosal surface make them more appealing as live cell factory. Among all available platforms, bioengineered probiotics are considered as the most affordable, most practical, and safest vaccination approach to halt this emerging virus., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The crucial role of the immune system in the progression/regression of breast cancer (BC) should always be taken into account. Various immunotherapy approaches have been investigated for BC, including tumor-targeting antibodies (bispecific antibodies), adoptive T cell therapy, vaccines, and immune checkpoint blockade such as anti-PD-1. In addition, a combination of conventional chemotherapy and immunotherapy approaches contributes to improving patients' overall survival rates. Although encouraging outcomes have been reported in most clinical trials of immunotherapy, some obstacles should still be resolved in this regard. Recently, personalized immunotherapy has been proposed as a potential complementary medicine with immunotherapy and chemotherapy for overcoming BC. Accordingly, this review discusses the brief association of these methods and future directions in BC immunotherapy., (Copyright © 2021. Published by Elsevier B.V.)