Identification of tyrosine kinase inhibitors from Panax bipinnatifidus and Panax pseudoginseng for RTK-HER2 and VEGFR2 receptors, by in silico approach.

Breast and stomach cancer is reported as a leading cause for human mortality across the world. The overexpression of receptor tyrosine kinase (RTK) proteins, namely the human epidermal growth factor receptor2 (HER2) and the vascular endothelial growth factor receptor2 (VEGFR2), is reported to be responsible for development and metastasis of breast and stomach cancer. Although several synthetic tyrosine kinase inhibitors (TKIs) as drug candidates targeting RTK-HER2 and VEGFR2 are currently available in the market, these are expensive with the reported side effects. This confers an opportunity for development of alternative novel tyrosine kinase inhibitors (TKIs) for RTK-HER2 and VEGFR2 receptors from the botanical sources. In the present study, we characterized 47 bioactive phytocompounds from the methanol extracts of the rhizomes of Asiatic traditional medicinal herbs-Panax bipinnatifidus and Panax pseudoginseng, of Indian Himalayan landraces using HPLC, GC-MS and high-sensitivity LC-MS tools. We performed molecular docking and molecular dynamics simulation analysis using Schrödinger suite 2020-3 to confirm the TKI phytocompounds showing the best binding affinity towards RTK-HER2 and VEGFR2 receptors. The results of molecular docking studies confirmed that the phytocompound (ligand) luteolin 7-O-glucoside (IHP15) showed the highest binding affinity towards receptor HER2 (PDB ID: 3PP0) with docking score and Glide g score (G-Score) of - 13.272, while chlorogenic acid (IHP12) showed the highest binding affinity towards receptor VEGFR2 (PDB ID: 4AGC) with docking score and Glide g score (G-Score) of - 10.673. Molecular dynamics (MD) simulation analysis carried out for 100 ns has confirmed strong binding interaction between the ligand and receptor complex [luteolin 7-O-glucoside (IHP15) and HER2 (PDB ID: 3PP0)] and is found to be stabilized within 40 to 100 ns of MD simulation, whereas ligand-receptor complex [chlorogenic acid (IPH12) and VEGFR2 (PDB ID: 4AGC)] also showed strong binding interaction and is found to be stabilized within 18-30 ns but slightly deviated during 100 ns of MD simulation. In silico ADME-Tox study using SwissADME revealed that the ligands luteolin 7-O-glucoside (IHP15) and chlorogenic acid (IHP12) have passed majority parameters of the common drug discovery rules. The present study has confirmed luteolin 7-O-glucoside (IHP15) and chlorogenic acid (IHP12) as potential tyrosine kinase inhibitors (TKIs) which were found to inhibit RTKs-HER2 and VEGFR2 receptor proteins, and thus paving the way for development of alternative potential TKIs (drug molecules) for treatment of HER2- and VEGFR2-positive breast and stomach cancer.

Combination of Near-Infrared Photoimmunotherapy Using Trastuzumab and Small Protein Mimetic for HER2-Positive Breast Cancer.

Near-infrared photoimmunotherapy (NIR-PIT) is a promising cancer therapy based on a monoclonal antibody conjugated to a photosensitizer (IR700Dye) that is activated by near-infrared light irradiation. We previously reported on the use of NIR-PIT with a small protein mimetic, the Affibody molecule (6-7 kDa), instead of a monoclonal antibody. In this study, we investigated a combination of NIR-PIT for HER2-positive breast cancer cells (SK-BR3, MDA-MB361, and JIMT1) with HER2 Affibody-IR700Dye conjugate and trastuzumab-IR700Dye conjugate. HER2 Affibody and trastuzumab target different epitopes of the HER2 protein and do not compete. In vitro, the combination of NIR-PIT using both HER2 Affibody-IR700Dye conjugate and trastuzumab-IR700Dye conjugate induced necrotic cell death of HER2-positive breast cancer cells without damage to HER2-negative breast cancer cells (MCF7). It was more efficient than NIR-PIT using either the HER2 Affibody-IR700Dye conjugate alone or the trastuzumab-IR700Dye conjugate alone. Additionally, this combination of NIR-PIT was significantly effective against HER2 low-expressing cancer cells, trastuzumab-resistant cells (JIMT1), and brain metastatic cells of breast cancer (MDA-MB361). Furthermore, in vivo imaging exhibited the strong fluorescence intensity of both HER2 Affibody-IR700Dye conjugates and trastuzumab-Alexa488 conjugates in HER2-positive tumor, indicating that both HER2 Affibody and trastuzumab specifically bind to HER2-positive tumors without competing with each other. In conclusion, the combination of NIR-PIT using both HER2 Affibody and trastuzumab expands the targeting scope of NIR-PIT for HER2-positive breast cancer.

Trans-(-)-Kusunokinin: A Potential Anticancer Lignan Compound against HER2 in Breast Cancer Cell Lines?

Trans-(-)-kusunokinin, an anticancer compound, binds CSF1R with low affinity in breast cancer cells. Therefore, finding an additional possible target of trans-(-)-kusunokinin remains of importance for further development. Here, a computational study was completed followed by indirect proof of specific target proteins using small interfering RNA (siRNA). Ten proteins in breast cancer were selected for molecular docking and molecular dynamics simulation. A preferred active form in racemic trans-(±)-kusunokinin was trans-(-)-kusunokinin, which had stronger binding energy on HER2 trans-(+)-kusunokinin; however, it was weaker than the designed HER inhibitors (03Q and neratinib). Predictively, trans-(-)-kusunokinin bound HER2 similarly to a reversible HER2 inhibitor. We then verified the action of (±)-kusunokinin compared with neratinibon breast cancer cells (MCF-7). (±)-Kusunokinin exhibited less cytotoxicity on normal L-929 and MCF-7 than neratinib. (±)-Kusunokinin and neratinib had stronger inhibited cell proliferation than siRNA-HER2. Moreover, (±)-kusunokinin decreased Ras, ERK, CyclinB1, CyclinD and CDK1. Meanwhile, neratinib downregulated HER, MEK1, ERK, c-Myc, CyclinB1, CyclinD and CDK1. Knocking down HER2 downregulated only HER2. siRNA-HER2 combination with (±)-kusunokinin suppressed HER2, c-Myc, CyclinB1, CyclinD and CDK1. On the other hand, siRNA-HER2 combination with neratinib increased HER2, MEK1, ERK, c-Myc, CyclinB1, CyclinD and CDK1 to normal levels. We conclude that trans-(±)-kusunokinin may bind HER2 with low affinity and had a different action from neratinib.

Immuno-hyperthermia effected by antibody-conjugated nanoparticles selectively targets and eradicates individual cancer cells.

Hyperthermia has been used for cancer therapy for a long period of time, but has shown limited clinical efficacy. Induction-heating hyperthermia using the combination of magnetic nanoparticles (MNPs) and an alternating magnetic field (AMF), termed magnetic hyperthermia (MHT), has previously shown efficacy in an orthotopic mouse model of disseminated gastric cancer. In the present study, superparamagnetic iron oxide nanoparticles (SPIONs), a type of MNP, were conjugated with an anti-HER2 antibody, trastuzumab and termed anti-HER2-antibody-linked SPION nanoparticles (anti-HER2 SPIONs). Anti-HER2 SPIONs selectively targeted HER2-expressing cancer cells co-cultured along with normal fibroblasts and HER2-negative cancer cells and caused apoptosis only in the HER2-expressing individual cancer cells. The results of the present study show proof-of-concept of a novel hyperthermia technology, immuno-MHT for selective cancer therapy, that targets individual cancer cells.Abbreviations: AMF: alternating magnetic field; DDW: double distilled water; DMEM: Dulbecco's Modified Eagle's; Medium; f: frequency; FBS: fetal bovine serum; FITC: Fluorescein isothiocyanate; GFP: green fluorescent protein; H: amplitude; Hsp: heat shock protein; MHT: magnetic hyperthermia; MNPs: magnetic nanoparticles; PI: propidium iodide; RFP: red fluorescent protein; SPION: superparamagnetic iron oxide (Fe3O4) nanoparticle.

Performance of capecitabine in novel combination therapies in colorectal cancer.

Colorectal cancer is one of the most common cancers throughout the world, and no definitive cure has ever been found. Perhaps a new insight into the effectiveness of chemotherapy drugs could help better treat patients. Targeted therapies have significantly improved the median overall survival of colorectal cancer patients. One of the standard chemotherapy regimens used for colorectal cancer is capecitabine, which is important in monotherapy and combination therapies. Capecitabine, with other chemotherapeutic agents (irinotecan, oxaliplatin, perifosine, 17-allylamino-17-demethoxygeldanamycin, aspirin, celecoxib, statins, quinacrine, inositol hexaphosphate and inositol, cystine/theanine, curcumin, and isorhamnetin), and biological ones (antibodies) plays an important role in the inhibition of some signaling pathways, increasing survival, reducing tumor growth and side effects of capecitabine. However, some drugs, such as proton pump inhibitors, are negatively related to capecitabine; therefore, the purpose of this work is to review and discuss the performance of capecitabine combination therapies in colorectal cancer.

Systematic insight into the active constituents and mechanism of Guiqi Baizhu for the treatment of gastric cancer.

Traditional Chinese medicine treatment of diseases has been recognized, but the material basis and mechanisms are not clear. In this study, target prediction of the antigastric cancer (GC) effect of Guiqi Baizhu (GQBZP) and the analysis of potential key compounds, key targets, and key pathways for the therapeutic effects against GC were carried out based on the method of network analysis and Kyoto Encyclopedia of Genes and Genomes enrichment. There were 33 proteins shared between GQBZP and GC, and 131 compounds of GQBZP had a high correlation with these proteins, indicating that the PI3K-AKT signaling pathway might play a key role in GC. From these studies, we selected human epidermal growth factor receptor 2 (HER2) and programmed cell death 1-ligand 1 (PD-L1) for docking; the results showed that 385 and 189 compounds had high docking scores with HER2 and PD-L1, respectively. Six compounds were selected for microscale thermophoresis (MST). Daidzein/quercetin and isorhamnetin/formononetin had the highest binding affinity for HER2 and PD-L1, with Kd values of 3.7 µmol/L and 490, 667, and 355 nmol/L, respectively. Molecular dynamics simulation studies based on the docking complex structures as the initial conformation yielded the binding free energy between daidzein/quercetin with HER2 and isorhamnetin/formononetin with PD-L1, calculated by molecular mechanics Poisson-Boltzmann surface area, of -26.55, -14.18, -19.41, and -11.86 kcal/mol, respectively, and were consistent with the MST results. In vitro experiments showed that quercetin, daidzein, and isorhamnetin had potential antiproliferative effects in MKN-45 cells. Enzyme activity assays showed that quercetin could inhibit the activity of HER2 with an IC50 of 570.07 nmol/L. Our study provides a systematic investigation to explain the material basis and molecular mechanism of traditional Chinese medicine in treating diseases.

HER2 Testing in Endometrial Serous Carcinoma: Time for Standardized Pathology Practice to Meet the Clinical Demand.

CONTEXT.­: Endometrial serous carcinoma is an aggressive subtype of endometrial cancer with the highest rate of recurrence and mortality among all histotypes. A recent clinical trial showed prolonged progression-free survival in advanced-stage and recurrent human epidermal growth factor receptor 2 (HER2)-positive endometrial serous carcinoma when trastuzumab was added to the standard chemotherapy regimen. This targeted therapeutic approach was recently endorsed by the National Comprehensive Cancer Network clinical guidelines. There is a growing interest among clinicians to obtain HER2 testing in endometrial serous carcinoma, and pathologists need to be prepared to recognize the unique characteristics of HER2 protein expression and gene amplification in these tumors and apply specific HER2 scoring criteria. OBJECTIVE.­: To provide a historical overview of targeted HER2 therapy in endometrial serous carcinoma and to summarize key findings from recent studies on the specific features of HER2 protein expression and gene amplification relative to other tumor types. Endometrial carcinoma-specific HER2 testing criteria are proposed based on evidence in the existing literature. DATA SOURCES.­: Sources comprise review of the literature and personal experience of the author. CONCLUSIONS.­: HER2 protein overexpression and/or gene amplification is present in approximately 25% to 30% of endometrial serous carcinomas, providing an opportunity for targeted therapy. Pathologists play a key role in tumor HER2 testing and scoring to ensure appropriate patient selection and successful clinical outcome.

Elaeagnus angustifolia Plant Extract Inhibits Epithelial-Mesenchymal Transition and Induces Apoptosis via HER2 Inactivation and JNK Pathway in HER2-Positive Breast Cancer Cells.

Elaeagnus angustifolia (EA) is a medicinal plant used for treating several human diseases in the Middle East. Meanwhile, the outcome of EA extract on HER2-positive breast cancer remains nascent. Thus, we herein investigated the effects of the aqueous EA extract obtained from the flowers of EA on two HER2-positive breast cancer cell lines, SKBR3 and ZR75-1. Our data revealed that EA extract inhibits cell proliferation and deregulates cell-cycle progression of these two cancer cell lines. EA extract also prevents the progression of epithelial-mesenchymal transition (EMT), an important event for cancer invasion and metastasis; this is accompanied by upregulations of E-cadherin and ß-catenin, in addition to downregulations of vimentin and fascin, which are major markers of EMT. Thus, EA extract causes a drastic decrease in cell invasion ability of SKBR3 and ZR75-1 cancer cells. Additionally, we found that EA extract inhibits colony formation of both cell lines in comparison with their matched control. The molecular pathway analysis of HER2 and JNK1/2/3 of EA extract exposed cells revealed that it can block HER2 and JNK1/2/3 activities, which could be the major molecular pathway behind these events. Our findings implicate that EA extract may possess chemo-preventive effects against HER2-positive breast cancer via HER2 inactivation and specifically JNK1/2/3 signaling pathways.

Targeting Her2-insYVMA with Covalent Inhibitors-A Focused Compound Screening and Structure-Based Design Approach.

Mutated or amplified Her2 serves as a driver of non-small cell lung cancer or mediates resistance toward the inhibition of its family member epidermal growth factor receptor with small-molecule inhibitors. To date, small-molecule inhibitors targeting Her2 which can be used in clinical routine are lacking, and therefore, the development of novel inhibitors was undertaken. In this study, the well-established pyrrolopyrimidine scaffold was modified with structural motifs identified from a screening campaign with more than 1600 compounds, which were applied against wild-type Her2 and its mutant variant Her2-A775_G776insYVMA. The resulting inhibitors were designed to covalently target a reactive cysteine in the binding site of Her2 and were further optimized by means of structure-based drug design utilizing a set of obtained complex crystal structures. In addition, the analysis of binding kinetics and absorption, distribution, metabolism, and excretion parameters as well as mass spectrometry experiments and western blot analysis substantiated our approach.

Extramammary Paget's disease patient-derived xenografts harboring ERBB2 S310F mutation show sensitivity to HER2-targeted therapies.

Although the prognosis of advanced extramammary Paget's disease (EMPD) is poor, there have been no preclinical research models for the development of novel therapeutics. This study aims to establish a preclinical research model for EMPD. We transplanted EMPD tissue into immunodeficient NOD/Scid mice. Histopathological and genetic analyses using a comprehensive cancer panel were performed. For in vivo preclinical treatments, trastuzumab, lapatinib, docetaxel, or eribulin were administered to patient-derived xenograft (PDX) models. Tissue transplanted from the EMPD patient was enlarged in NOD/Scid mice and was transplanted into further generations. Both the transplantation of PDX into nu/nu mice and the reanimation of the cryopreserved xenografted tumors in NOD/Scid mice were successful. We also established an EMPD-PDX-derived primary cell culture. Histopathologically, the xenografted tumors were positive for CK7, which was consistent with the patient's tumors. Genetically, the pathogenic mutation ERBB2 S310F was detected in the patient's tumors (primary intraepidermal lesion, metastatic lymph node) and was observed in the xenografted tumors even after continued passages. The xenografted tumors responded well to trastuzumab and lapatinib therapy. Also, cytotoxic agents (docetaxel and eribulin) were effective against the xenografted tumors. This PDX model (EMPD-PDX-H1) could be a powerful tool for the research and development of EMPD treatments.

Pyrotinib in the treatment of human epidermal growth factor receptor 2-positive metastatic breast cancer: A case report.

RATIONALE: Pyrotinib is a novel dual pan-ErbB receptor tyrosine kinase inhibitor, approved for the treatment of human epidermal growth factor receptor 2 (HER2)-positive metastatic breast cancer (MBC). However, there was still limited information regarding specific effect of pyrotinib on HER2-positive MBC patients with phosphoinositol-3 kinase mutation. PATIENT CONCERNS: A 63-year-old woman accidentally discovered a left breast lesion. The breast cancer was diagnosed by biopsy of breast lesion and postoperative pathological examination in March, 2017. The patient was presented with HER2-positive (3+), invasive carcinoma of the left breast with lymph nodes and lung nodules metastasis, and the clinical stage was T4N2M1. However, the lesion continued to aggressive disease progression with the treatment of trastuzumab plus multiple chemotherapy regimens and traditional Chinese medicine. DIAGNOSES: The woman was diagnosed with invasive carcinoma of the left breast and lymph nodes and lung nodules metastasis. INTERVENTIONS: The patient received 6 cycles of pyrotinib in combination with capecitabine regularly. OUTCOMES: Progression free survival was more than 6 months, and the patient's efficacy evaluation was partial remission. LESSONS: Our clinical observations demonstrated that pyrotinib may be an effective treatment for patients with HER2-positive MBC.

Bio-engineered cell membrane nanovesicles as precision theranostics for perihilar cholangiocarcinoma.

Perihilar cholangiocarcinoma (PHCC) presents a formidable challenge due to its occult anatomic location, aggressive growth, insensitivity to conventional chemotherapy, and poor prognosis. Herein, we engineered a human epidermal growth factor receptor 2 (HER2) affibody to the surface of cell membrane nanovesicles (A-NVs) in a ligand-oriented manner and loaded them with indocyanine green (ICG) as precision theranostics for PHCC treatment. The A-NVs@ICG were prepared and exhibited satisfactory targeting effects in HER2-overexpressing PHCC cells. In vivo fluorescence and photoacoustic imaging demonstrated that A-NVs@ICG promoted the accumulation of ICG in PHCC tissue, leading to enhanced tumor regression and improved anti-cancer effects when combined with photoirradiation. Therefore, bio-engineered A-NVs@ICG represent a promising nanotheranostic agent for PHCC with potential for clinical translation.

A novel anti-HER2 antibody GB235 reverses Trastuzumab resistance in HER2-expressing tumor cells in vitro and in vivo.

HER2 overexpression is frequently associated with tumor metastasis and poor prognosis of breast cancer. More evidence indicates that HER3 is involved in HER2-resistant therapies. Combination treatments with two or more different monoclonal antibodies are a promising strategy to overcome resistance to HER2 therapies. We presented a novel fully human HER2-targeted monoclonal antibody, GB235, screened from a phage-display library against the HER2 antigen. GB235 in combination with Trastuzumab overcomes resistance in HER2-positive tumors and results in more sustained inhibition of tumor growth over time. The competition binding assay showed that the epitopes of GB235 do not overlap with those of Pertuzumab and Trastuzumab on HER2. Further HER2 mutagenesis results revealed that the binding epitopes of GB235 were located in the domain III of HER2. The mechanism of action of GB235 in blocking HER2-driven tumors is different from the mechanisms of Trastuzumab or Pertuzumab. GB235 does not affect the heterodimerization of HER2 and HER3, whereas the GB235 combined treatment with Trastuzumab significantly inhibited heregulin-induced HER3 phosphorylation and downstream signaling. Moreover, GB235 in combination with Trastuzumab reversed the resistance to heregulin-induced proliferation in HER2-overexpressing cancer cell lines. GB235 combined with Trastuzumab treatment in xenograft models resulted in improved antitumor activity. Complete tumor suppression was observed in the HER2-positive NCI-N87 xenograft model treated with the combination treatment with GB235 and Trastuzumab. In a Trastuzumab-resistant patient-derived tumor xenograft model GA0060, GB235 plus Trastuzumab reversed the resistance to Trastuzumab monotherapy. Because GB235 showed a different working mechanism with Pertuzumab and Trastuzumab, these agents can be considered complementary therapy against HER2 overexpression tumors.

HO-1 drives autophagy as a mechanism of resistance against HER2-targeted therapies.

PURPOSE: Targeted therapies have resulted in major advances in the treatment of HER2-positive breast cancers. Despite this, up to 70% of patients will develop resistance to treatment within 2 years and new strategies for targeting resistant disease are needed. METHODS: To identify potential resistance mechanisms, we used the mouse MMTV-NIC-PTEN+/- spontaneous model of HER2-positive breast cancer and the pan-HER family kinase inhibitor sapatinib. Vehicle and sapatinib-treated tumors were evaluated by immunohistochemistry and proteomic analysis. In vitro studies were carried out to define the role of heme oxygenase 1 (HO-1) and autophagy in resistance to sapatinib and lapatinib, another pan-HER family kinase inhibitor. RESULTS: Treatment of tumor-bearing MMTV-NIC-PTEN+/- mice with sapatinib resulted in delayed tumor progression and increased survival. However, tumors eventually progressed on treatment. Proteomic analysis identified proteins associated with cellular iron homeostasis as being upregulated in the sapatinib-treated tumors. This included HO-1 whose overexpression was confirmed by immunohistochemistry. Overexpression of HO-1 in HER2-expressing SKBR3 breast cancer cells resulted in reduced sensitivity to both pan-HER family kinase inhibitors sapatinib and lapatinib. This was associated with increased autophagy in the HO-1 over-expressing cells. Furthermore, increased autophagy was also seen in the sapatinib-treated tumors. Treatment with autophagy inhibitors was able to increase the sensitivity of the HO-1 over-expressing cells to both lapatinib and sapatinib. CONCLUSION: Together these data indicate a role for HO-1-induced autophagy in resistance to pan-HER family kinase inhibitors.

Antibody-drug conjugate T-DM1 treatment for HER2+ breast cancer induces ROR1 and confers resistance through activation of Hippo transcriptional coactivator YAP1.

BACKGROUND: A newly developed drug trastuzumab emtansine (T-DM1) has improved the survival of breast cancer (BC) patients. Despite an impressive initial clinical response, a subgroup of patient develop resistance and present therapeutic challenges. The underlying resistance mechanisms are not fully investigated. We report that T-DM1 treatment modulates the expression of ROR1 (type 1 receptor tyrosine kinase-like orphan receptor) and induces self-renewal of cancer stem cells (CSCs) leading to therapeutic resistance. METHODS: Using BC patient tumor samples, and BC cell lines we gained insight into the T-DM1 treatment induced ROR1 overexpression and resistance. In vitro sphere forming assays and in vivo extreme dilution assays were employed to analyze the stemness and self-renewal capacity of the cells. A series of molecular expression and protein assays including qRT-PCR, FACS-sorting, ELISA, immunostaining, Western blotting were used to provide evidence. FINDINGS: Exposure of cells to T-DM1 shifted ROR1 expression from low to high, enriched within the CSC subpopulation, coincident with increased Bmi1 and stemness factors. T-DM1 induced ROR1 cells showed high spheroid and tumor forming efficiency in vitro and in an animal model exhibiting shorter tumor-free time. Mechanistically, the overexpression of ROR1 is partly induced by the activation of YAP1 and its target genes. Silencing of ROR1 and YAP1 by pharmacologic inhibitors and/or sh/siRNA inhibited spheroid formation, the initiation of tumors and the capacity for self-renewal and ROR1 overexpression. INTERPRETATIONS: The results presented here indicate that simultaneous targeting of ROR1 and YAP1 may suppress CSC self-renewal efficacy and inhibit tumor progression in BC. In this manner such treatments may overcome the T-DM1 mediated therapeutic resistance and improve clinical outcome. FUND: This study was supported by Neurogen Technologies for interdisciplinary research.

TAS0728, A Covalent-binding, HER2-selective Kinase Inhibitor Shows Potent Antitumor Activity in Preclinical Models.

Activated HER2 is a promising therapeutic target for various cancers. Although several reports have described HER2 inhibitors in development, no covalent-binding inhibitor selective for HER2 has been reported. Here, we report a novel compound TAS0728 that covalently binds to HER2 at C805 and selectively inhibits its kinase activity. Once TAS0728 bound to HER2 kinase, the inhibitory activity was not affected by a high ATP concentration. A kinome-wide biochemical panel and cellular assays established that TAS0728 possesses high specificity for HER2 over wild-type EGFR. Cellular pharmacodynamics assays using MCF10A cells engineered to express various mutated HER2 genes revealed that TAS0728 potently inhibited the phosphorylation of mutated HER2 and wild-type HER2. Furthermore, TAS0728 exhibited robust and sustained inhibition of the phosphorylation of HER2, HER3, and downstream effectors, thereby inducing apoptosis of HER2-amplified breast cancer cells and in tumor tissues of a xenograft model. TAS0728 induced tumor regression in mouse xenograft models bearing HER2 signal-dependent tumors and exhibited a survival benefit without any evident toxicity in a peritoneal dissemination mouse model bearing HER2-driven cancer cells. Taken together, our results demonstrated that TAS0728 may offer a promising therapeutic option with improved efficacy as compared with current HER2 inhibitors for HER2-activated cancers. Assessment of TAS0728 in ongoing clinical trials is awaited (NCT03410927).

Targeting neratinib-induced diarrhea with budesonide and colesevelam in a rat model.

PURPOSE: Neratinib is an irreversible pan-ErbB tyrosine kinase inhibitor used for the extended adjuvant treatment of early-stage HER2-positive breast cancer. Its use is associated with the development of severe diarrhea in up to 40% of patients in the absence of proactive management. We previously developed a rat model of neratinib-induced diarrhea and found inflammation and anatomical disruption in the ileum and colon. Here we tested whether anti-diarrheal interventions, budesonide and colesevelam, can reduce neratinib-induced diarrhea and intestinal pathology. METHODS: Rats were treated with 50 mg/kg neratinib via oral gavage for 14 or 28 days (total n = 64). Body weight and diarrhea severity were recorded daily. Apoptosis was measured using immunohistochemistry for caspase-3. Inflammation was measured via a multiplex cytokine/chemokine assay. ErbB levels were measured using PCR and Western Blot. RESULTS: Budesonide co-treatment caused rats to gain significantly less weight than neratinib alone from day 4 of treatment (P = 0.0418). Budesonide (P = 0.027) and colesevelam (P = 0.033) each reduced the amount of days with moderate diarrhea compared to neratinib alone. In the proximal colon, rats treated with neratinib had higher levels of apoptosis compared to controls (P = 0.0035). Budesonide reduced histopathological injury in the proximal (P = 0.0401) and distal colon (P = 0.027) and increased anti-inflammatory IL-4 tissue concentration (ileum; P = 0.0026, colon; P = 0.031) compared to rats treated with neratinib alone. In the distal ileum, while budesonide decreased ErbB1 mRNA expression compared to controls (P = 0.018) (PCR), an increase in total ErbB1 protein was detected (P = 0.0021) (Western Blot). CONCLUSION: Both budesonide and colesevelam show potential as effective interventions against neratinib-induced diarrhea.

Role of MEL-18 Amplification in Anti-HER2 Therapy of Breast Cancer.

BACKGROUND: Resistance to HER2-targeted therapy with trastuzumab still remains a major challenge in HER2-amplified tumors. Here we investigated the potential role of MEL-18, a polycomb group gene, as a novel prognostic marker for trastuzumab resistance in HER2-positive (HER2+) breast cancer. METHODS: The genetic alteration of MEL-18 and its clinical relevance were examined in multiple breast cancer cohorts including METABRIC (n = 1,980), TCGA (n = 825), and our clinical specimens (n = 213, trastuzumab-treated HER2+ cases). MEL-18 amplification was validated by fluorescence in situ hybridization (FISH) analysis. The MEL-18 effect on trastuzumab response was confirmed by in vitro cell viability assays and an in vivo xenograft experiment (n = 7 per group). Gene expression microarray and receptor tyrosine kinase array were performed to identify the trastuzumab resistance mechanism by MEL-18 loss. All statistical tests were two-sided. RESULTS: MEL-18 was exclusively amplified in approximately 30-50% of HER2+ breast tumors and was associated with a favorable clinical outcome (disease-free survival: P = .02 in HER2+ cases, METABRIC; P = .04 in patients receiving trastuzumab). In MEL-18-amplified HER2+ breast cancer, MEL-18 depletion induced trastuzumab resistance by increasing ADAM sheddase-mediated ErbB ligand production and receptor heterodimerization. MEL-18 epigenetically silenced ADAM10/17 expression in cooperation with polycomb-repressive complex (PRC) 1 and PRC2. Combination treatment with an ADAM10/17 inhibitor and trastuzumab could overcome MEL-18 loss-mediated trastuzumab resistance in vivo (BT474/shMEL-18 xenograft: trastuzumab, mean [SD] tumor volume = 406.1 [50.1] mm3, vs trastuzumab + GW280264 30 mg/kg, mean [SD] tumor volume = 68.4 [15.6] mm3, P < .001). Consistently, trastuzumab-treated patients harboring concomitant MEL-18 amplification and low ADAM17 expression showed prolonged relapse-free survival (P = .02 in our cohort, n = 213). CONCLUSION: MEL-18 serves to prevent ligand-dependent ErbB heterodimerization and trastuzumab resistance, suggesting MEL-18 amplification as a novel biomarker for HER2+ breast cancer.

Discovery of Nonpeptide, Reversible HER1/HER2 Dual-Targeting Small-Molecule Inhibitors as Near-Infrared Fluorescent Probes for Efficient Tumor Detection, Diagnostic Imaging, and Drug Screening.

The abnormal expression of epidermal growth factor receptors HER1(EGFR) and HER2 is strongly associated with cancer invasion, metastasis, and angiogenesis. Their molecular detection is mainly executed using genetically encoded or antibody-based diagnostic tracers, but no dual-targeting small-molecule bioprobe has been achieved. Here, we report the novel small-molecule fluorescent probes Cy3-AFTN and Cy5-AFTN as potent dual-targeting inhibitors for efficient detection of HER1/HER2 expression in cancer cells and in vivo tumor diagnostic imaging. Unlike the irreversible HER1/HER2 inhibitors, Cy3-AFTN and Cy5-AFTN were designed as reversible/noncovalent probes based on the clinical drug afatinib, by making the molecule structurally impossible for receptor-mediated Michael additions. The synthesized probes were validated with live cell fluorescence imaging, flow cytometry and confocal-mediated competitive binding inhibition, molecular docking study, and in vivo xenograft tumor detection. The probes are competitively replaceable by other HER1/HER2 inhibitors; thus, they are potentially useful in fluorometric high-throughput screening for drug discovery.

Milestones over the development of SB3, a trastuzumab biosimilar.

The development of a biosimilar requires large extensive preclinical and clinical comparability exercises to demonstrate equivalence to the reference medical product. The holistic results from this large assessment should be taken into account to appreciate the validity of the development and the interpretations. SB3 is the first trastuzumab biosimilar approved for routine use in Europe. The present manuscript reviews the development and the results of SB3, including clinical assessment and the clinical Phase I, as well as the large randomized Phase III comparing efficacy between SB3 versus Herceptin® containing regimen in neoadjuvant setting. Key points of the design and interpretations of the findings are extensively discussed in this review of SB3 development.