HER2-positive Breast Cancer Cells Research Articles

Abstract PO5-04-02: Combination of trastuzumab competing and non-competing fully human internalizing anti-Her2 monoclonal antibodies drug conjugates increases their inhibitory effect on the growth of HER2 positive metastatic breast cancer cells

Abstract The human epidermal growth factor receptor (HER) family of receptors plays a key role in proliferation stimulator for several human cancers. In particular, HER2 is overexpressed in 15-30% of breast cancers with or without gene amplification with prognostic and predictive implications. HER2 overexpression is associated with shorter disease-free and overall survivals, resistance to hormonal agents and increased risk of brain metastasis. Trastuzumab is a humanized anti-HER2 monoclonal antibody that binds to domain IV of HER2 extracellular domain and blocks its signaling. More recently, two antibody drug conjugates using trastuzumab have been approved: Ado-Trastuzumab-Emtansine (Kadcyla) consisting of trastuzumab conjugated to the drug mertansine DM1 and fam-trastuzumab-deruxtecan-nhki (Enhertu to deliver the topoisomerase inhibitor deruxtecan. We are reporting here the development of fully human internalizing anti-HER2 antibodies with distinct epitopes on the HER2protein which compete or not with trastuzumab for binding to HER2. These monoclonal antibodies have been developed by immunizing fully human (TC-Mab mouse) mice with recombinant HER2 protein. After production of hybridoma secreting fully human immunoglobulins, the screening process included competition with trastuzumab for binding to Her2 by enzyme linked immunoassay and Octet epitope binning and affinity determination as well as internalization assay. Several internalizing antibodies able to compete or not with trastuzumab and with high affinity (Kd ranging from 10−9 M to 10−12 M) were selected. Their ability to deliver a cytotoxic payload in HER2 overexpressing breast cancer cells such as SKBR3, BT474 and AU565 was next investigated. Two groups of antibodies: one competing with trastuzumab and the other one non-competing were further characterized. Data related to these antibodies’ biochemical characteristics as well as their ability to inhibit proliferation in vitro and in vivo in mouse xenografts studies will be presented here. In conclusion, the use of fully human mice to develop monoclonal antibodies provides a powerful and attractive approach to develop fully human monoclonal antibodies against cancer targets by-passing the need for humanization and affinity maturation of antibodies. This work is supported by 5R44CA224718 SBIR grant from the National Cancer Institute. Citation Format: Ginette Serrero, Binbin Yue, Jianping Dong, Jun Hayashi. Combination of trastuzumab competing and non-competing fully human internalizing anti-Her2 monoclonal antibodies drug conjugates increases their inhibitory effect on the growth of HER2 positive metastatic breast cancer cells [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-04-02.

Abstract PO3-24-05: Cysteine metabolism related ferroptosis sensitivity in trastuzumab resistant HER2 positive breast cancer

Abstract Background Trastuzumab has shown great effectiveness in HER2 positive breast cancer treatment, but about 50% patients would undergo resistance during or after treatment. Although previous research has suggested several potential reasons for trastuzumab resistance, the metabolic reprogramming during resistance formation remains largely unclear. Here we identified aberrant ferroptosis associated cysteine metabolism in trastuzumab resistant HER2 positive breast cancer, which might become a novel target for overcoming resistance. Methods Trastuzumab sensitive HER2 positive breast cancer cell SKBR3 and resistant JITM1 were obtained for transcriptomics, proteomics, metabolomics and epigenomics analysis. Gene silencing was mediated by siRNAs. CUT&Tag was applied to compare H3K4me3 and H3K27me3 binding regions. DNA methylation levels and different methylated regions were evaluated by WGBS-seq. CRISPRi with dCas9-DNMT3A was applied to regulate specific DNA methylation in CpG islands. Lipid ROS was measured by flow cytometry with BODIPY-C11. Results Joint analyses of transcriptomics and proteomics according to ferroptosis pathways revealed downregulated glutathione metabolism, glutamate transmembrane and homocysteine metabolism processes, as well as upregulated fatty acid metabolism and iron metabolism pathways in JIMT1. Metabolomics verified that JIMT1 increased cysteine metabolism and decreased glutathione metabolism. SLC7A11 expression and GSH/GSSG ratio were increased in JIMT1, while no difference was observed in free cysteine. JIMT1 featured significant higher UGC codon usage bias and increased cysteinyl-tRNA synthetase. The abundance of H3K4me3 other than H3K27me3 in SLC7A11 promoter region was found increased in JIMT1, and the 5-mC level of CpG islands in SLC7A11 promoter region was shown decreased. Using dCas9-DNMT3A, the methylation of SLC7A11 promoter was enhanced and SLC7A11 expression was reduced in JIMT1. Inhibition of SLC7A11 by siRNAs, CRISPRi or Erastin all indicated a higher ferroptosis sensitivity in JIMT1. Conclusion Trastuzumab resistant HER2 positive breast cancer features aberrant cysteine metabolism resulting from altered H3K4me3 modification and DNA methylation in SLC7A11 promoter region. This might provide novel targets for further anti-HER2 treatment. Citation Format: Yongmei Yin, Yijia Hua, Ningjun Duan, Chunxiao Sun, Fan Yang. Cysteine metabolism related ferroptosis sensitivity in trastuzumab resistant HER2 positive breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-24-05.

Innovative tumor interstitial fluid-triggered carbon dot-docetaxel nanoassemblies for targeted drug delivery and imaging of HER2-positive breast cancer

In this study, we have developed an innovative pH-triggered nanomedicine delivery system, targeting HER2-positive breast cancer cells for effective low-cost, imaging-guided drug delivery and precise therapy. The key feature of this system lies in its unique tumor interstitial fluid microenvironment-responsive drug release behavior which achieved tumor site-specific drug delivery. Our in vitro experiments demonstrated that the carbon dot-integrated material achieves more efficient DTX release (96.13 % at 72 h) in the tumor interstitial fluid microenvironment (pH 6.5), thereby boosting drug concentration at the tumor site and enhancing therapeutic efficacy. Further cell experiments confirmed the system's significant inhibitory effect on HER2-positive tumor cells SKBR3 in a pH 6.5 environment, and apoptosis assays indicating a notable increase in early cell apoptosis (from 8.39 % to 24.61 % compared with pH 7.4). Furthermore, the integration of HER2 aptamer within the carbon dot-based system enables targeted recognition and binding to tumor cells, ensuring more precise delivery of DTX while minimizing potential side effects. Crucially, the carbon dots in this system emit superior red fluorescence (the QY = 47.64 % excited at 535 nm compared with Rodamine 6G), enabling real-time visualization of the drug delivery process. This feature provides valuable feedback on treatment effectiveness, facilitating necessary adjustments. The small size (1.88 ± 0.48 nm) of carbon dots significantly improved their ability to penetrate biological barriers, while their low toxicity (no significant cell toxicity under 350 μg/mL) contributed to the formulation’s outstanding biocompatibility. Overall, this carbon dot-enhanced drug delivery system offers immense potential for enhancing drug efficacy, minimizing side effects, and providing real-time treatment monitoring, thus proposing a innovate strategy for breast cancer therapy.

Liver X Receptor Ligand GAC0001E5 Downregulates Antioxidant Capacity and ERBB2/HER2 Expression in HER2-Positive Breast Cancer Cells.

The HER2-positive subtype accounts for approximately one-fifth of all breast cancers. Insensitivity and development of acquired resistance to targeted therapies in some patients contribute to their poor prognosis. HER2 overexpression is associated with metabolic reprogramming, facilitating cancer cell growth and survival. Novel liver X receptor (LXR) ligand GAC0001E5 (1E5) has been shown to inhibit cancer cell proliferation by disrupting glutaminolysis and inducing oxidative stress. In this study, HER2-positive breast cancer cells were treated with 1E5 to determine their potential inhibitory effects and mechanisms of action in HER2-positive breast cancers. Similar to previous observations in other cancer types, 1E5 treatments inhibited LXR activity, expression, and cancer cell proliferation. Expression of fatty acid synthesis genes, including fatty acid synthase (FASN), was downregulated following 1E5 treatment, and results from co-treatment experiments with an FASN inhibitor suggest that the same pathway is targeted by 1E5. Treatments with 1E5 disrupted glutaminolysis and resulted in increased oxidative stress. Strikingly, HER2 transcript and protein levels were both significantly downregulated by 1E5. Taken together, these findings indicate the therapeutic potential of targeting HER2 overexpression and associated metabolic reprogramming via the modulation of LXR in HER2-positive breast cancers.

Exploring Anticancer Properties of Medicinal Plants against Breast Cancer by Downregulating Human Epidermal Growth Factor Receptor 2.

Plants have a history of being employed in managing breast cancer. However, no scientific evidence supports the idea that these plants can effectively reduce the level of HER2 expression. In this study, extracts from 10 medicinal plants were evaluated for their anticancer properties against HER2-positive breast cancer cells through various methods, including the SRB assay, comet assay, annexin V-FITC dual staining, and immunoblotting. All extracts exerted antiproliferative activity against HER2-positive breast cancer cells. Furthermore, Terminalia chebula (T. chebula), Berberis aristata (B. aristata), and Mucuna pruriens (M. pruriens) reduced HER2 expression in tested cell lines. In addition, an increased Bax/Bcl-2 ratio was observed after the treatment. A comparative proteomics study showed modulation in the proteome profile of breast cancer cells after treatment with T. chebula, B. aristata, Punica granatum, M. pruriens, and Acorus calamus. Metabolic profiling of lead plants revealed the existence of multiple anticancer compounds. Our study demonstrates the considerable potential of the mentioned plants as innovative therapies for HER2-positive breast cancer.

Maitake Beta-Glucan Enhances the Therapeutic Effect of Trastuzumab via Antibody-Dependent Cellular Cytotoxicity and Complement-Dependent Cytotoxicity.

Trastuzumab, an anti-HER2 monoclonal antibody, is the mainstay treatment for of HER2-positive breast cancer. However, trastuzumab resistance is often observed during treatment. Therefore, new therapeutic strategies are needed to enhance the clinical benefits of trastuzumab. Maitake β-glucan MD-Fraction, isolated from Grifola frondosa, inhibits tumor growth by enhancing immune responses. In this study, we examined the effect of MD-Fraction on trastuzumab treatment of HER2-positive breast cancer. MD-Fraction did not directly inhibit the survival of HER2-positive breast cancer cells, alone or in the presence of trastuzumab in vitro. In HER2-positive xenograft models, the combination of MD-Fraction and trastuzumab was more effective than trastuzumab alone. Peripheral blood lymphocytes and splenic natural killer cells isolated from BALB/c nu/nu mice treated with MD-Fraction showed enhanced trastuzumab-induced antibody-dependent cellular cytotoxicity (ADCC) ex vivo. MD-Fraction-treated macrophages and neutrophils did not show enhanced trastuzumab cytotoxicity in the presence of heat-inactivated serum, but they showed enhanced cytotoxicity in the presence of native serum. These results suggest that MD-Fraction-treated macrophages and neutrophils enhance trastuzumab-induced complement-dependent cellular cytotoxicity (CDCC). Treatment of HER2-positive breast cancer cells with MD-Fraction in the presence of trastuzumab and native serum increased C3a release and tumor cell lysis in a dose-dependent manner, indicating that MD-Fraction enhanced trastuzumab-induced complement-dependent cytotoxicity (CDC) by activating the complement system. This study demonstrates that the combination of trastuzumab and MD-Fraction exerts a greater antitumor effect than trastuzumab alone by enhancing ADCC, CDCC, and CDC in HER2-positive breast cancer.

IRX4204 Induces Senescence and Cell Death in HER2-positive Breast Cancer and Synergizes with Anti-HER2 Therapy.

Rexinoids, agonists of nuclear retinoid X receptor (RXR), have been used for the treatment of cancers and are well-tolerated in both animals and humans. However, the usefulness of rexinoids in treatment of breast cancer remains unknown. This study examines the efficacy of IRX4204, a highly specific rexinoid, in breast cancer cell lines and preclinical models to identify a biomarker for response and potential mechanism of action. IRX4204 effects on breast cancer cell growth and viability were determined using cell lines, syngeneic mouse models and primary PDX tumors. In vitro assays of cell cycle, apoptosis, senescence, and lipid metabolism were used to uncover a potential mechanism of action. Standard anti-HER2 therapies were screened in combination with IRX4204 on a panel of breast cancer cell lines to determine drug synergy. IRX4204 significantly inhibits the growth of HER2-positive breast cancer cell lines, including trastuzumab and lapatinib resistant JIMT-1 and HCC1954. Treatment with IRX4204 reduced tumor growth rate in the MMTV-ErbB2 mouse and HER2-positive PDX model by 49% and 44% respectively. Mechanistic studies revealed IRX4204 modulates lipid metabolism and induces senescence of HER2-positive cells. In addition, IRX4204 demonstrates additivity and synergy with HER2 targeted monoclonal antibodies, tyrosine kinase inhibitors, and antibody drug conjugates. These findings identify HER2 as a biomarker for IRX4204 treatment response and demonstrate a novel use of RXR agonists to synergize with current anti-HER2 therapies. Furthermore, our results suggest that RXR agonists can be useful for the treatment of anti-HER2 resistant and metastatic HER2-positive breast cancer.

Trastuzumab-conjugated liposomes for co-delivery of paclitaxel and anti-abcb1 siRNA in HER2-positive breast cancer: In vitro and in vivo evaluations

Chemotherapeutic drugs often face challenges such as non-specific binding, undesired toxicity, and limited blood circulation, leading to reduced therapeutic efficacy. This study introduces HER2-specific targeted nanoparticles designed by synthesizing cationic liposomes. These liposomes encapsulate paclitaxel and ABCB1-siRNA in their core and feature a therapeutic monoclonal antibody, trastuzumab, on the surface. This design aims for precise targeting and synergistic treatment of HER2-positive breast cancer cells in both in vitro and in vivo models. The optimized trastuzumab-conjugated liposomes exhibited a particle size of 229 ± 4 nm and a zeta potential of 43.46 ± 0.61 mV. Their spherical morphology was confirmed using Scanning Electron Microscopy (SEM). These liposomes demonstrated enhanced drug retention in in vitro release studies. Furthermore, trastuzumab-conjugated liposomes displayed markedly higher cellular uptake than their non-trastuzumab-conjugated counterparts. Their anticancer efficacy in BT-474 cells significantly surpassed that of non-targeted liposomes and unencapsulated paclitaxel. In xenograft Nude mice models derived from the BT-474 cell line, trastuzumab-conjugated liposomes showed superior tumor distribution and preclinical effectiveness compared to both non-targeted liposomes and plain paclitaxel.

Abstract 5540: Single cell multiomic analysis of human brain metastasis reveals conserved dormancy population

Abstract This study explores the complex realm of brain metastasis, pivotal in crafting effective anti-tumor strategies. Leveraging single-cell multi-omics sequencing, we profiled the transcriptomes and chromatin accessibility landscapes of tumor cells and their surrounding microenvironment in human brain metastases that originated from across a spectrum of primary cancers. Our investigations have identified distinct, yet uniformly present, tumor cell subpopulations, each defined by unique gene expression patterns, chromatin accessibility, and the interactions between tumor and stromal cells. Notably, one such subpopulation exhibits a dormancy signature, characterized by enhanced interferon response, oxidative phosphorylation, and upregulated HLA class molecules, alongside alterations in circadian rhythms. Our trajectory analysis charts a course from these dormant cells to rapidly proliferating ones, with notable shifts in circadian rhythm genes, underscoring their potential role in brain metastasis. This aligns with recent findings that disruptions in circadian rhythms can modify stem cell characteristics in glioblastomas. This insight led us to focus on a specific circadian rhythm gene, highly active in the dormant cells and located on the HER2 amplicon—an area linked to high brain metastasis relapse rates in patients. We've developed inducible models for both overexpression and knockdown to scrutinize this gene's impact in her2 positive breast cancer cell lines, examining gene expression and proliferation in vitro. Complementing this, we are conducting spatial transcriptomic analysis to evaluate the microenvironmental regulation of tumor dormancy in human brain metastases. This multi-omics approach reveals a nuanced regulatory landscape of brain metastasis, highlighting a novel and often overlooked factor in metastasis progression and brain tumor relapse: the interplay between circadian rhythms and tumor dormancy. This discovery not only deepens our understanding of brain metastases but also opens potential new avenues for therapeutic interventions. Citation Format: Remi Klotz, Yiru Wang, Anu Sunkara, Frank Attenelo, Min Yu. Single cell multiomic analysis of human brain metastasis reveals conserved dormancy population [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5540.

Abstract 480: Nanotherapeutic strategies to improve targeted radionuclide therapy

Abstract Cancer is a leading cause of death worldwide. Several multidisciplinary approaches exist for cancer treatment, including radiotherapy and chemotherapy. Radiotherapy uses high-energy radiation to kill cancer cells; chemotherapy inhibits cancer cell proliferation and often kills cells by targeting the cell cycle. Resistance to radiotherapy and chemotherapy is a key determining factor in the outcome of therapeutic efficacy. Conventional nontargeted radiotherapy also affects distant nonirradiated cells, leading to DNA damage and changes in the cell cycle that are often linked to secondary carcinogenesis in patients. In recent years, targeted radionuclide therapy (TRT) has emerged as a promising personalized treatment strategy that delivers cytotoxic levels of radiation directly and specifically to cancer cells. Among radioisotopes,225Ac exhibits desirable properties for TRT: multiple α-particle emission and high cytotoxicity. One of the challenges with 225Ac is the nonspecific toxicity caused by the release and relocation of its decay daughters. Different approaches have been proposed to prevent the relocation of decay daughters, including nanoparticles. Nanoparticles have been pursued as a promising delivery vehicle of α-emitting radioisotopes to the tumor site. Here, we show specific targeting of Her2-positive breast cancer cells with 225Ac-radiolabeled lanthanum orthovanadate (LaVO4) nanoparticles. Nanoparticles’ surfaces were functionalized to improve biocompatibility and conjugated to target cancer cells, respectively. Cellular uptake and localization of these engineered nanoparticles were analyzed by a Confocal microscope and IncuCyte live cell analysis system. Results confirm its localization in the nucleus following the endolysosomal path, enhancing the nanoparticles’ effectiveness as a delivery vehicle for α-emitting radioisotopes with the potential of increased treatment efficacy. Upon addition of cancer-targeting ligands, these nanoparticles can achieve higher efficiency by delivering the radioisotope to the tumor cells with a potential of safe encapsulation of all α-emitters in the decay chain. The anticancer efficiency of 225Ac-radiolabeled nanoparticles was demonstrated by a dose-dependent effective killing of 3D breast cancer spheroids using both IncuCyte live imaging and the cell survival assays. Ultimately, the results of this study will be crucial in determining the future use of targeted radiolabeled nanoparticle-based delivery systems as an approach for more efficacious cancer treatment. Citation Format: Debjani Pal, Miguel Toro Gonzáleza, Amber N. Bibleb, Brian Sanders, Anna Plechaty, Owee Kirpekar, Mircea Podar, Sandra M. Davern. Nanotherapeutic strategies to improve targeted radionuclide therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 480.

Abstract 536: Breaking through ADCs resistance: Constructing preclinical enhertu-induced resistant model

Abstract Antibody-drug conjugates (ADCs) are compounds that combine the specificity of monoclonal antibodies (mAbs) with the cytotoxic payload of toxic substances. In recent years, ADCs have rapidly developed in the field of cancer therapy, particularly in the treatment of HER2-positive breast cancer. One representative example is Trastuzumab deruxtecan (Enhertu), which has become a new benchmark for anti-HER2 ADCs due to its excellent performance in clinical trials. Despite the impressive clinical outcomes of ADCs, some tumors in patients may still develop resistance to these drugs. Therefore, it is crucial to understand the mechanisms of resistance and develop strategies to overcome ADCs resistance. To achieve these goals, it is essential to establish and utilize preclinical models to study these resistance mechanisms. Preclinical models of resistance to ADCs have mainly been generated using human cultured cell lines. JIMT-1 is a HER2-positive breast cancer cell line that exhibits primary resistance to the anti-HER2 antibody trastuzumab and also shows resistance to trastuzumab-derived ADC T-DM1. However, we found that Enhertu significantly inhibited tumor growth in JIMT-1 xenograft models. We subjected JIMT-1 tumor tissues to continuous passage in mice and administered long-term drug induction. The tumor growth rate gradually increased, and the drug sensitivity decreased, resulting in the acquisition of an Enhertu-resistant tumor model. The obtained resistant tumor tissues not only include tumor cells themselves but also comprise fibroblasts, extracellular matrix, and other cellular components, enabling a more comprehensive investigation into the interactions between different cells within the tumor microenvironment (TME). We believe that the establishment of preclinical Enhertu-resistant models helps predict resistance that has not yet emerged clinically and provides references for future drug development. Citation Format: Yuan Fang, Huiyi Wang, Jun Xing, Lu Yang, Jing Zhao, Xiang Gao, Cunxiang Ju. Breaking through ADCs resistance: Constructing preclinical enhertu-induced resistant model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 536.

Protecting the Brain: Novel Strategies for Preventing Breast Cancer Brain Metastases through Selective Estrogen Receptor β Agonists and In Vitro Blood-Brain Barrier Models.

Breast cancer brain metastasis (BCBM) is a challenging condition with limited treatment options and poor prognosis. Understanding the interactions between tumor cells and the blood-brain barrier (BBB) is critical for developing novel therapeutic strategies. One promising target is estrogen receptor β (ERβ), which promotes the expression of key tight junction proteins, sealing the BBB and reducing its permeability. In this study, we investigated the effects of 17β-estradiol (E2) and the selective ERβ agonist diarylpropionitrile (DPN) on endothelial and cancer cells. Western blot analysis revealed the expression patterns of ERs in these cell lines, and estrogen treatment upregulated claudin-5 expression in brain endothelial cells. Using in vitro models of the BBB, we found that DPN treatment significantly increased BBB tightness about suppressed BBB transmigration activity of representative Her2-positive (BT-474) and triple-negative (MDA-MB-231) breast cancer cell lines. However, the efficacy of DPN treatment decreased when cancer cells were pre-differentiated in the presence of E2. Our results support ERβ as a potential target for the prevention and treatment of BCBM and suggest that targeted vector-based approaches may be effective for future preventive and therapeutic implications.

Efficacy and delivery strategies of the dual Rac/Cdc42 inhibitor MBQ-167 in HER2 overexpressing breast cancer

Trastuzumab and trastuzumab-based treatments are the standard of care for breast cancer patients who overexpress the human epidermal growth factor receptor 2 (HER2). However, patients often develop resistance to trastuzumab via signaling from alternative growth factor receptors that converge to activate guanine nucleotide exchange factors (GEFs) that in turn activate the Rho GTPases Rac and Cdc42. Since Rac and Cdc42 have been implicated in high tumor grade and therapy resistance, inhibiting the activity of Rac and Cdc42 is a rational strategy to overcome HER2-targeted therapy resistance. Therefore, our group developed MBQ-167, a dual Rac/Cdc42 inhibitor with IC50s of 103 nM and 78 nM for Rac and Cdc42, respectively, which is highly effective in reducing cell and tumor growth and metastasis in breast cancer cell and mouse models. Herein, we created a trastuzumab resistant variant of the SKBR3 HER2 positive breast cancer cell line and show that Rac activation is a central mechanism in trastuzumab resistance. Next, we tested the potential of targeting MBQ-167 to HER2 overexpressing trastuzumab-resistant cell lines in vitro, and show that MBQ-167, but not trastuzumab, reduces cell viability and induces apoptosis. When MBQ-167 was targeted to mammary fatpad tumors established from HER2 overexpressing cells via immunoliposomes functionalized with trastuzumab, MBQ-167 and MBQ-167-loaded liposomes show equal efficacy in reducing the viability of trastuzumab-resistant cells, inhibiting tumor growth in mouse xenografts, and reducing metastasis to lungs and liver. This study demonstrates the efficacy of MBQ-167 as an alternative therapeutic in HER2 overexpressing cancers, delivered either in free form or in liposomes.

Targeted dual degradation of HER2 and EGFR obliterates oncogenic signaling, overcomes therapy resistance, and inhibits metastatic lesions in HER2-positive breast cancer models

AimsHuman epidermal growth factor receptor 2 (HER2) is an oncogenic receptor tyrosine kinase amplified in approximately 20% of breast cancer (BC). HER2-targeted therapies are the linchpin of treating HER2-positive BC. However, drug resistance is common, and the main resistance mechanism is unknown. We tested the hypothesis that drug resistance results mainly from inadequate or lack of inhibition of HER2 and its family member epidermal growth factor receptor (EGFR). MethodsWe used clinically relevant cell and tumor models to assess the impact of targeted degradation of HER2 and EGFR on trastuzumab resistance. Trastuzumab is the most common clinically used HER2 inhibitor. Targeted degradation of HER2 and EGFR was achieved using recombinant human protein PEPDG278D, which binds to the extracellular domains of the receptors. siRNA knockdown was used to assess the relative importance of EGFR and HER2 in trastuzumab resistance. ResultsBoth HER2 and EGFR are overexpressed in all trastuzumab-resistant HER2-positive BC cell and tumor models and that all trastuzumab-resistant models are highly vulnerable to targeted degradation of HER2 and EGFR. Degradation of HER2 and EGFR induced by PEPDG278D causes extensive inhibition of oncogenic signaling in trastuzumab-resistant HER2-positive BC cells. This is accompanied by strong growth inhibition of cultured cells, orthotopic patient-derived xenografts, and metastatic lesions in the brain and lung of trastuzumab-resistant HER2-positive BC. siRNA knockdown indicates that eliminating both HER2 and EGFR is necessary to maximize therapeutic outcome. ConclusionsThis study unravels the therapeutic vulnerability of trastuzumab-resistant HER2-positive BC and shows that an agent that targets the degradation of both HER2 and EGFR is highly effective in overcoming drug resistance in this disease. The findings provide new insights and innovations for advancing treatment of drug-resistant HER2-positive breast cancer that remains an unmet problem.

Evaluating a targeted Palbociclib-Trastuzumab loaded smart niosome platform for treating HER2 positive breast cancer cells

In this study, we present a targeted and pH-sensitive niosomal (pHSN) formulation, incorporating quantum dot (QD)-labeled Trastuzumab (Trz) molecules for the specific delivery of Palbociclib (Pal) to cells overexpressing human epidermal growth factor receptor 2 (HER2). FTIR analyses confirmed the successful preparation of the pHSNs and their bioconjugation. The labeled Trz-conjugated Pal-pHSNs (Trz-Pal-pHSNs) exhibited a size of approximately 170 nm, displaying a spherical shape with a neutral surface charge of −1.2 mV. Pal encapsulation reached ∼86%, and the release pattern followed a two-phase pH-dependent mechanism. MTT assessments demonstrated enhanced apoptosis induction, particularly in HER2-positive cells, by Trz-Pal-pHSNs. Fluorescence imaging further validated the internalization of particles into cells. In conclusion, Trz-Pal-pHSNs emerge as a promising platform for personalized medicine in the treatment of HER2-positive breast cancer.

EB1089 Increases the Antiproliferative Response of Lapatinib in Combination with Antiestrogens in HER2-Positive Breast Cancer Cells.

HER2-positive breast cancer is associated with aggressive behavior and reduced survival rates. Calcitriol restores the antiproliferative activity of antiestrogens in estrogen receptor (ER)-negative breast cancer cells by re-expressing ERα. Furthermore, calcitriol and its analog, EB1089, enhance responses to standard anti-cancer drugs. Therefore, we aimed to investigate EB1089 effects when added to the combined treatment of lapatinib and antiestrogens on the proliferation of HER2-positive breast cancer cells. BT-474 (ER-positive/HER2-positive) and SK-BR-3 (ER-negative/HER2-positive) cells were pre-treated with EB1089 to modulate ER expression. Then, cells were treated with EB1089 in the presence of lapatinib with or without the antiestrogens, and proliferation, phosphorylation array assays, and Western blot analysis were performed. The results showed that EB1089 restored the antiproliferative response to antiestrogens in SK-BR-3 cells and improved the inhibitory effects of the combination of lapatinib with antiestrogens in the two cell lines. Moreover, EB1089, alone or combined, modulated ERα protein expression and reduced Akt phosphorylation in HER2-positive cells. EB1089 significantly enhanced the cell growth inhibitory effect of lapatinib combined with antiestrogens in HER2-positive breast cancer cells by modulating ERα expression and Akt phosphorylation suppression. These results highlight the potential of this therapeutic approach as a promising strategy for managing HER2-positive breast cancer.

Artemisia argyi extracts overcome lapatinib resistance via enhancing TMPRSS2 activation in HER2-positive breast cancer.

Breast cancer stands as the predominant malignancy and primary cause of cancer-related mortality among females globally. Approximately 25% of breast cancers exhibit HER2 overexpression, imparting a more aggressive tumor phenotype and correlating with poor prognoses. Patients with metastatic breast cancer receiving HER2 tyrosine kinase inhibitors (HER2 TKIs), such as Lapatinib, develop acquired resistance within a year, posing a critical challenge in managing this disease. Here, we explore the potential of Artemisia argyi, a Chinese herbal medicine known for its anti-cancer properties, in mitigating HER2 TKI resistance in breast cancer. Analysis of the Cancer Genome Atlas (TCGA) revealed diminished expression of transmembrane serine protease 2 (TMPRSS2), a subfamily of membrane proteolytic enzymes, in breast cancer patients, correlating with unfavorable outcomes. Intriguingly, lapatinib-responsive patients exhibited higher TMPRSS2 expression. Our study unveiled that the compounds from Artemisia argyi, eriodictyol, and umbelliferone could inhibit the growth of lapatinib-resistant HER2-positive breast cancer cells. Mechanistically, they suppressed HER2 kinase activation by enhancing TMPRSS2 activity. Our findings propose TMPRSS2 as a critical determinant in lapatinib sensitivity, and Artemisia argyi emerges as a potential agent to overcome lapatinib via activating TMPRSS2 in HER2-positive breast cancer. This study not only unravels the molecular mechanisms driving cell death in HER2-positive breast cancer cells induced by Artemisia argyi but also lays the groundwork for developing novel inhibitors to enhance therapy outcomes.

Targeting HER2-positive breast cancer cells by a combination of dasatinib and BMS-202: Insight into the molecular pathways

BackgroundRecent investigations have reported the benefits of using a tyrosine kinase inhibitor, dasatinib (DA), as well as programmed death-ligand 1 (PD-L1) inhibitors in the management of several solid tumors, including breast cancer. Nevertheless, the outcome of the combination of these inhibitors on HER2-positive breast cancer is not explored yet.MethodsHerein, we investigated the impact of DA and PD-L1 inhibitor (BMS-202) combination on HER2-positive breast cancer cell lines, SKBR3 and ZR75.ResultsOur data reveal that the combination significantly inhibits cell viability of both cancer cell lines as compared to monotreatment. Moreover, the combination inhibits epithelial-mesenchymal transition (EMT) progression and reduces cancer cell invasion by restoring E-cadherin and β-catenin expressions and loss of vimentin, major biomarkers of EMT. Additionally, the combination reduces the colony formation of both cell lines in comparison with their matched control. Also, the combination considerably inhibits the angiogenesis of the chorioallantoic membrane model compared with monotreatment. Molecular pathway analysis of treated cells shows that this combination blocks HER2, AKT, β-catenin, and JNK1/2/3 activities.ConclusionOur findings implicate that a combination of DA and BMS-202 could have a significant impact on the management of HER2-positive breast cancer.

Encapsulation and release of calcein from herceptin-conjugated eLiposomes

Achieving an optimal therapeutic level is crucial in effectively eradicating cancer cells during treatment. However, conventional chemotherapy-associated systemic administration of anticancer agents leads to many side effects. To achieve the desired control over the target site, active targeting of HER2-positive breast cancer cells can be achieved by conjugating liposomal vesicles with Human Epidermal growth factor Receptor 2 (HER2) and inducing release of the encapsulated drug using ultrasound. To further enhance the delivery efficiency, nanoemulsion droplets exhibiting responsiveness to low-frequency ultrasound are encapsulated within these lipid vesicles. In this study, we prepared four different liposomal formulations, namely pegylated liposomes, emulsion liposomes (eLiposomes), HER-conjugated liposomes, and HER-conjugated eLiposomes, each loaded with calcein and subjected to a thorough characterization process. Their sizes, phospholipid concentration, and amount of antibody conjugation were compared and analyzed. Cryogenic transmission electron microscopy was used to confirm the encapsulation of nanoemulsion droplets within the liposomes. The drug-releasing performance of Herceptin-conjugated eLiposomes was found to surpass that of other liposomal formulations with a notably higher calcein release and established it as a highly effective nanocarrier. The study showcases the efficacy of calcein-loaded and Herceptin-conjugated eLiposomes, which demonstrate rapid and efficient drug release among other liposomal formulations when subjected to ultrasound. This discovery paves the way for a more targeted, efficient, and humane approach to cancer therapy.

Comparison of trastuzumab emtansine, trastuzumab deruxtecan, and disitamab vedotin in a multiresistant HER2-positive breast cancer lung metastasis model

Human epidermal growth factor 2 (HER2)-positive breast cancer with lung metastases resistant to targeted agents is a common therapeutic challenge. Absence of preclinical lung metastasis models that are resistant to multiple anti-HER2 targeted drugs hampers the development of novel therapies. We established a novel HER2-positive breast cancer cell line (L-JIMT-1) with a high propensity to form lung metastases from the parenteral JIMT-1 cell line by injecting JIMT-1 cells into immunodeficient SCID mice. Lung metastases developed in all mice injected with L-JIMT-1 cells, and more rapidly and in greater numbers compared with the parental JIMT-1 cells. L-JIMT-1 cells expressed more epidermal growth factor receptor and HER2 than JIMT-1 cells. L-JIMT-1 cells were resistant to all five tyrosine kinase inhibitors tested in vitro (afatinib, erlotinib, lapatinib, sapitinib, and tucatinib). When we compared JIMT-1 and L-JIMT-1 sensitivity to three HER2-targeting antibody-drug conjugates (ADCs) trastuzumab emtansine (T-DM1), trastuzumab deruxtecan (T-DXd), and disitamab vedotin (DV) in vitro, JIMT-1 cells were resistant T-DXd, partially sensitive to T-DM1, and sensitive to DV, while L-JIMT-1 cells were resistant to both T-DM1 and T-DXd, but moderately sensitive to DV. In a mouse model, all three ADCs inhibited the growth of L-JIMT-1 lung metastases compared to a vehicle, but DV and T-DXd more strongly than T-DM1, and DV treatment led to the smallest tumor burden. The L-JIMT breast cancer lung metastasis model developed may be useful in the evaluation of anti-cancer agents for multiresistant HER2-positive advanced breast cancer.