HER2 Gene Research Articles

Folic acid-functionalized PEGylated niosomes co-encapsulated cisplatin and doxoribicin exhibit enhanced anticancer efficacy

The medical field is faced with the difficult task of developing a new approach to curing cancer, which is prevalent in organs such as the breast and ovaries and has a high mortality rate. Since chemotherapy is the conventional method of treatment, efforts are being made to improve it to help patients function better. Fortunately, with the use of nanocarriers and their remarkable ability to manage and direct drug delivery, progress is being made in cancer treatment. In addition, folic acid-coated nanocarriers offer several advantages in drug delivery, including improved stability, bioavailability, targeted delivery and drug solubility. These properties make them promising tools for improving cancer treatment efficacy. This research focused on investigating the stability of a specific niosomal formulation (consisting of Span 60 and cholesterol) under different temperature conditions (4 and 25 ℃) for 2 months. In addition, the drug release rate of the formulation was evaluated. The results showed that the size and polydispersity index increased significantly in the stability studies, but the entrapment efficiency% decreased dramatically over time. In addition, encapsulation of drugs in niosomal formulations resulted in stable and slow drug release. The cytotoxicity evaluation results of formulations containing doxorubicin and cisplatin show their significant inhibitory effect on both breast and ovarian cancer cell lines (IC50 for DOX–CIS–Nio@PEG–FA formulation was 6.11 and 17.87 µg/mL for A2780 and MCF-7, respectively). Niosomes loaded with a combination of two drugs were found to affect gene expression in the cancer cell lines tested. They decreased the expression of BCl2, VEGF, CCND1, and HER2 genes while increasing the expression of BAX gene. Flow cytometry results indicated that niosomes loaded with doxorubicin and cisplatin increased the rate of apoptosis in both cell lines compared to a drug mixture. ROS and cell cycle arrest, confirm the significant inhibition of cancer cells and their destruction in the presence of the synthesized noisome formulation in comparison to free drugs and the combination of two drugs. The potential of this novel approach for delivering drugs to cancer cells lies in the ability to combine treatments and target multiple cancers simultaneously. Such formulations allow co-delivery of drugs to different cancer cells, thereby improving the efficacy of chemotherapy through synergistic effects between drugs.Graphical

Prognostic impact of HER2 biomarker levels in trastuzumab-treated early HER2-positive breast cancer

BackgroundOverexpression of human epidermal growth factor receptor 2 (HER2) caused by HER2 gene amplification is a driver in breast cancer tumorigenesis. We aimed to investigate the prognostic significance of manual scoring and digital image analysis (DIA) algorithm assessment of HER2 copy numbers and HER2/CEP17 ratios, along with ERBB2 mRNA levels among early-stage HER2-positive breast cancer patients treated with trastuzumab.MethodsThis retrospective study comprised 371 early HER2-positive breast cancer patients treated with adjuvant trastuzumab, with HER2 re-testing performed on whole tumor sections. Digitized tumor tissue slides were manually scored and assessed with uPath HER2 Dual ISH image analysis, breast algorithm. Targeted ERBB2 mRNA levels were assessed by the Xpert® Breast Cancer STRAT4 Assay. HER2 copy number and HER2/CEP17 ratio from in situ hybridization assessment, along with ERBB2 mRNA levels, were explored in relation to recurrence-free survival (RFS).ResultsThe analysis showed that patients with tumors with the highest and lowest manually counted HER2 copy number levels had worse RFS than those with intermediate levels (HR = 2.7, CI 1.4–5.3, p = 0.003 and HR = 2.1, CI 1.1–3.9, p = 0.03, respectively). A similar trend was observed for HER2/CEP17 ratio, and the DIA algorithm confirmed the results. Moreover, patients with tumors with the highest and the lowest values of ERBB2 mRNA had a significantly worse prognosis (HR = 2.7, CI 1.4–5.1, p = 0.003 and HR = 2.8, CI 1.4–5.5, p = 0.004, respectively) compared to those with intermediate levels.ConclusionsOur findings suggest that the association between any of the three HER2 biomarkers and RFS was nonlinear. Patients with tumors with the highest levels of HER2 gene amplification or ERBB2 mRNA were associated with a worse prognosis than those with intermediate levels, which is of importance to investigate in future clinical trials studying HER2-targeted therapy.

Abstract A005: Ultra-rapid and precise measurement of HER2 copy number alteration by next-generation digital PCR capable of real-time analysis in patients with breast cancer: A multicenter retrospective study

Abstract The HER2 diagnostics is necessary for selection of patients harboring HER2 gene amplification or protein overexpression who will benefit from anti-HER2 therapies in breast cancer. However, HER2 testing is still challenging due to the subjective natures of immunohistochemistry (IHC) and in situ hybridization (ISH), standard methods for determining HER2 status. Thus, a new method is needed to accurately quantify HER2 levels. Here, we developed a clinically reliable HER2 testing method enabling ultra-fast detection of HER2 gene amplification with high accuracy by using the digital real-time PCR (drPCR) system, a potential new diagnostic platform with improved performance by integrating both real-time and digital PCR technologies. For drPCR-based HER2 copy number (CN) measurement, primer-probe sets specific to HER2 gene and a genomic region adjacent to chromosome 17 centromere (CEP17) were designed, and the optimal drPCR condition was determined in clinical breast tumor specimens. To test the clinical validity and standardize procedures of drPCR-based HER2 status evaluation, three independent breast cancer cohorts from different institutions were enrolled, which assigned as a training (SCHU hospital, n = 103) and two validation sets (SNU hospital, n = 170; CNUH hospital, n = 45), and the drPCR assay was compared with current standard HER2 testing methods. In the training cohort, the HER2/CEP17 ratio values from FISH and drPCR tests were highly correlated (r2 = 0.81; P < 0.001), and the drPCR results displayed 98.1% concordance to HER2 status defined by IHC and/or FISH with 92.6% sensitivity and 100% specificity. Eight samples further verified by targeted NGS showed 100% concordance of dPCR to NGS. Consistently, two validation cohorts also showed high concordance of drPCR to IHC and/or ISH results (accuracy = 97.1% and 97.8% in SNU and CNUH cohorts, respectively). The optimal cutoff for HER2 positivity in the drPCR assay was set as a HER2/CEP17 ratio ≥ 1.9 with AUC of 0.963 based on the results from training cohort, and the same cut-off for drPCR was applicable to two independent validation cohorts, supporting the clinical validity of our drPCR-based HER2 assessment. In some discordant cases, low tumor purity (≤ 25%) was observed and microdissection partly improved the drPCR results. The discordance between drPCR and ISH results was also found in marginal HER2+ cases with HER2/CEP17 ratio 2-3, but these cases showed inter-observer variability when re-evaluating the ISH/IHC data due to intratumoral HER2 heterogeneity. Of note, in HER2 IHC3+ cases with negative drPCR results, re-evaluation of IHC using an artificial intelligence (AI)-based HER2 scoring system revised the HER2 IHC 3+ score to 2+, and ISH assessment also confirmed that these cases are indeed HER2-negative, proving the high accuracy of HER2 CN drPCR assay. In conclusion, given the advantages of drPCR-based HER2 assessment with high accuracy, sensitivity, and simplicity, the drPCR assay could be a complementary or alternative method to IHC and ISH to greatly improve current HER2 testing. Citation Format: Jin hyuk Chang, YoonSik Kim, Hee-Joo Choi, Soo Young Park, Ji-Hye Park, Hee-Young Won, Min Ji Song, Da Sol Kim, Hayeon Kim, Sohyeon Yang, Nam Hun Heo, Minsik Song, Seung-Shick Shin, Do Young Lee, Han Suk Ryu, Si-Hyong Jang, Jeong-Yeon Lee. Ultra-rapid and precise measurement of HER2 copy number alteration by next-generation digital PCR capable of real-time analysis in patients with breast cancer: A multicenter retrospective study [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr A005.

Abstract IA21: Mechanisms of Resistance to HER2-Targeted Therapies in HER2-positive Breast Cancer

Abstract Recent advancements in HER2-targeted therapies for HER2+ BC have revolutionized patient outcome. Yet, intrinsic and acquired resistance, especially in the advanced setting remains a clinical challenge. In the era of precision oncology, better understanding of these mechanisms of resistance is key to identifying new strategies to overcome resistance and for developing personalized treatment approaches. Our recent studies suggest that a subset of HER2+ BCs, associated with high and homogeneous levels of HER2 gene amplification, protein, and activity, are addicted to HER2 and may therefore benefit from chemotherapy-sparing HER2-targeted regimens. However, even in such tumors, constitutive activation of the PI3K/AKT pathway due to deregulations in the downstream PI3K pathway can lead to resistance. In all HER2+ BCs, failure to achieve a comprehensive blockade of the HER receptor layer is one of the major mechanisms of resistance due to the functional redundancy of the HER receptors and compensatory signaling within the pathway. But, in some cases, effective inhibition of HER receptors might be challenged by molecular masking of the receptors (e.g., MUC4) or by epi/genetic or post-translational alterations in the receptors itself (e.g., HER2 L755S, p95HER2). Indeed, we recently reported that acquisition of HER2 L755S, either alone or together with PIK3CA mutation, and hyperactive EGFR signaling due to acquired EGFR amplification or excess HER ligands are associated with resistance to HER2-targeted therapy, especially tyrosine kinase inhibitors. Likewise, clinically, HER mutations have been reported to be further enriched in the metastatic vs. primary HER2+ tumors. When HER2 does remain sustainably inhibited by HER2-targeted agents, resistance may arise due to the emergence of alternative signaling pathways transmitting compensatory proliferative and survival signals, such as ER, receptor tyrosine kinases, or downstream/intracellular signaling. Further, the emergence of metabolic pathways as alternative pathways of resistance have also been documented. We recently reported the activation of the cholesterol biosynthetic mevalonate (MVA) pathway as an escape mechanism that provides alternative signals through the YAP/TAZ-mTORC1-survivin axis to evade HER2 blockade. This resistance could be overcome using MVA pathway inhibitors (e.g., statins). Additionally, activation of the cell cycle regulatory complex cyclin D1/CDK4 has been shown to be associated with HER2-targeted therapy resistance, which could be exploited as a potential therapeutic vulnerability by using CDK4/6 inhibitors. Finally, response to HER2-targeted therapy could potentially be also modulated by the tumor microenvironment itself, including components of the host immune system and extracellular matrix. Together, a better understanding of the underlying mechanisms of resistance in the context of tumor and host biology, and treatment setup is fundamental to develop new treatment strategies and to guide patient stratification for personalized treatments and improved patient outcomes. Citation Format: Jamunarani Veeraraghavan, Fu-Tien Liao, Lanfang Qin, Tia Gordon, Alekya Raghavan, Caroline Sabotta, Rachel Kaplan, Carolina Gutierrez, C. Kent Osborne, Mothaffar F. Rimawi, Rachel Schiff. Mechanisms of Resistance to HER2-Targeted Therapies in HER2-positive Breast Cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr IA21.

A modified triplet combination of docetaxel, oxaliplatin, and fluorouracil for gastric cancer (GC) with peritoneal carcinomatosis (PC) and inoperable malignant bowel obstruction (MBO): A multi-center, non-randomized, three-cohort, phase II trial (Zhen Jing).

TPS426 Background: For GC, PC is ranked as the most common in relapse and the second common in metastasis. MBO is a frequent and preterminal complication of PC, which has been reported to have a poor overall survival (OS) of only 3 months without effective treatment. Medical therapy of gastroenteric drainage and symptomatic drugs is the cornerstone management for MBO, just alleviating symptoms. Interventional therapy and surgery are effective but indicated in limited patients (pts) with strict selection. Chemotherapy is the mainstay treatment for metastatic GC but its role in MBO is poorly defined. Only a few retrospective studies are available, showing conflicting results on obstruction clearance (OC) and OS. Methods: We are conducting the Zhen Jing trial in five Chinese centers to investigate the tolerability and efficacy of a dose-sense regimen of docetaxel, oxaliplatin and fluorouracil in MBO from GC. Cohort A is for pts in first-line setting. Cohort B is for second-line setting. Cohort C is for pts who have failed to two or more regimens. Eligible pts will receive the same experimental regimen every 28 days: docetaxel 25 mg/m2 on days 1, 8, 15; oxaliplatin 85 mg/m2 on days 1, 15; fluorouracil 1200 mg/m2 24-hour infusion on days 1, 8, 15. A Simon's two-stage optimal design is applied (table below). The primary endpoint is 60-day OC rate. Secondary endpoints are 30-day OC rate, time to OC, OC duration, safety, overall survival and quality of life. Till July 2023, a total of 34 pts has been enrolled (16 in cohort A, 9 in cohort B, 9 in cohort C). Inclusion Criteria: 18-75 years old; Eastern Cooperative Oncology Group performance status ≤ 3; adenocarcinoma of stomach or gastroesophageal junction; peritoneal carcinomatosis established by imaging data or pathological evidence; MBO below the Treitz ligament based on clinical grounds or radiological findings; inappropriate for operation judged independently by two surgeons; written informed consent. Exclusion Criteria: Treated by a combination regimen containing all the study drugs; allergy to any of the study drugs; HER-2 gene overexpression; high microsatellite instability or mismatch repair deficiency; strangulated obstruction; active gastrointestinal bleeding; uncontrolled active infection; unstable heart diseases; severe lung diseases; central nervous system diseases; concomitant cerebral or meningeal metastasis; surgery or stent are required. Clinical trial information: NCT04840264 . [Table: see text]

Abstract A065: Lineage tracing oncogenic isoforms of HER2 in a mouse model of pancreatic cancer

Abstract Among the numerous clinical challenges of pancreatic ductal adenocarcinoma (PDAC) is the eventual development of therapeutic resistance. Although previous studies report that only about 3% of PDAC cases are HER2 gene amplified, as many as 82% of PDAC tumors may have HER2 overexpressed. Anti-HER2 therapy with trastuzumab has yielded tremendous therapeutic success in breast cancer and multiple gastrointestinal malignancies, however minimal benefit is found in PDAC. Experimental mouse models have demonstrated initiation and acceleration of pancreatic tumorigenesis through overexpression of activated mutant HER2, yet the role of HER2 alone as a driver of PDAC and treatment resistance is still unknown. Multiple HER2 isoforms exist that are generated either via alternative splicing and loss of exon 16 (d16 HER2) or through N-terminal truncations (p95 HER2). These isoforms are implicated in differential responses to anti-HER2 therapy in the breast. Previously we described a HER2+ cancer rainbow mouse model in which human wild-type HER2 (WT HER2), d16 HER2, and p95 HER2 were expressed in the same mouse and lineage traced through co-expression of fluorescent protein reporters. Studies in the mammary gland revealed that each isoform drives distinct tumor phenotypes. Herein, we hypothesize that these isoforms drive HER2 positive PDAC. HER2 Crainbow mice were crossed with mice expressing PDX1-Cre recombinase to initiate recombination and expression of each fluorescently labelled HER2 isoform during pancreas development. PDX1-Cre/HER2 Crainbow (PHBOW) mice were followed for up to one year and sacrificed to lineage trace each isoform and register HER2 lineages with histopathology. Within our pilot cohort, early-stage pre-malignant pancreatic intraepithelial neoplasia (PanIN-1) was observed in 75% of PDAC HER2bow mice as early as five weeks. By one year, 50% demonstrated high grade PanIN (i.e. PanIN-3) and features of early invasion. Although WT HER2 was expressed abundantly throughout the pancreas, lineage tracing revealed WT HER2 was not found in pre-malignant lesions. Conversely, the pre-malignant PanIN lesions and early invasive histopathologic features traced to d16 HER2 expressing cells, while p95 HER2 traced with areas of acinar-to-ductal metaplasia, lesions known to be early initiators of pancreatic dysplasia. Our data show how the oncogenic HER2 isoforms, and not WT HER2, are able to drive PanIN lesions and, at lower frequency, invasive PDAC after one year. Future work seeks to complete HER2 Crainbow lineage tracing in PHBOW mice, test HER2 targeted therapy in PHBOW mice, and quantify HER2 isoforms in HER2+ human PDAC cases. Citation Format: Elishama N. Kanu, Joelle Sills, Rebecca Shelley, Joshua D. Ginzel, Joseph Fernandes, Ashley A. Fletcher, Peter J. Allen, Daniel P. Nussbaum, Joshua C. Snyder. Lineage tracing oncogenic isoforms of HER2 in a mouse model of pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr A065.

Association of mutation profiles with metastasis in patients with non-small cell lung cancer.

This study focused on the analysis of the correlation between common gene mutation types and metastatic sites in NSCLC patients. We retrospectively studied 1586 NSCLC patients and used fluorescence Polymerase chain reaction (PCR) to detect EGFR, ALK, ROS1, RET, MET, BRAF, HER2, KRAS, NRAS, and PIK3CA gene mutations, and also investigated sex, smoking status, age at diagnosis, histological type and TNM stage. In addition, we analyzed the site of metastasis in patients with stage IV NSCLC. The EGFR-mutation group more frequently metastasized to lung (18.9%, P = 0.004), brain (18.9%, P = 0.001) and bone (27.1%, P = 0.004) than wild-type patients. ALK-mutation group (71.0%, P < 0.001), BRAF-mutation group (82.4%, P = 0.005) and NRAS-mutation group (100%, P = 0.025) were more likely to metastasize than the wild-type group. In the ALK mutation, lung metastasis (24.2%, P = 0.013), brain (24.2%, P = 0.007), bone metastasis (32.3%, P = 0.024), liver metastasis (19.4%, P = 0.001), and pleural metastasis (29.0%, P = 0.021) were common. In the KRAS-mutation group, lung metastasis (21.7%, P = 0.012) and brain metastasis (23.3%, P = 0.001) were more common. Less metastasis occurred in the HER2-mutation group (28.3%, P = 0.014). There was no difference in the RET, MET and PIK3CA mutations. Patients with ALK mutant, BRAF mutant or NRAS mutant were more prone to metastasis, while the HER 2 mutation group was less metastatic. Patients with EGFR mutant NSCLC are more likely to develop bone, lung, or brain metastasis.

CCQM-K176 breast cancer biomarker HER2 copy number variation (CNV) measurement

Main textBreast cancer is the most common malignant tumor in women all over the world. Breast cancers with amplification of the human epidermal growth factor receptor 2 (HER2) gene (referred to as HER2-positive breast cancer) accounts for about 20% ~ 30% of invasive breast cancer, and this type of tumor is characterized by high invasiveness, high risk of recurrence, rapid progression and poor prognosis. The determination of HER2 gene expression or copy number variant (CNV) (HER2 copy number normalized to cell number or a reference gene) in tumor cells of breast cancer patients is beneficial to the choice of treatment, through targeted therapies such as trastuzumab, and prognosis. Testing of the core competencies of laboratories to deliver services of CNV measurements for different genetic analytes has not been covered previously. Agreement was received in the October 2019 meeting of the NAWG in Torino to conduct the HER2 CNV measurement as a key comparison. Evidence of successful participation in formal, relevant international comparisons is needed to support calibration and measurement capability claims (CMCs) made by national metrology institutes (NMIs) and designated institutes (DIs).Ten National Metrology Institutions participated in the Track A Key Comparison: CCQM-K176 "Breast cancer biomarker HER2 copy number variation (CNV) measurement". Participants were requested to evaluate the copy number concentration (expressed in [μL-1]) and copy number ratio of defined genomic sequences in eukaryotic intact genomic DNA at an indicative concentration of 90~130 ng/μL. Participation in K176 required assay selection/design and quantitative detection of the analyte in the buffer. The objectives of the key comparison were to measure 1) HER2 copy number concentration; 2) the copy number concentration of the single copy reference gene ribonuclease P RNA component H1 (RPPH1); 3) the HER2/ RPPH1 copy number ratio, using two defined sequences representative of the two genes (HER2 and RPPH1) in mixed genomic DNA samples, and 4) provide evidence for CMC claims by participating laboratories when measuring target sequence content in purified genomic DNA. All participants performed measurements by digital PCR (dPCR), with alternative assays and pre-treatment of samples (for example, restriction enzyme digestion). Interlaboratory reproducibility (%CV) was less than 13% and 5% for copy number concentration and HER2/RPPH1 copy number ratio, respectively, demonstrating very high interlaboratory reproducibility in CNV measurements.Successful participation in CCQM-K176 demonstrates the following measurement capabilities in determining copy number ratio (of a defined target sequence expressed relative to a reference gene sequence) in the range from 1.0 to 40.0 at a background of relatively high concentration of intact (non-fragmented) genomic DNA. The study will also support participants' claim for measurement of defined DNA sequence/gene copy number concentration in the range ~104-106 μL-1 (taking into account both reported reference gene and HER2 copy number concentration values).To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/.The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

IN-SILICO CHARACTERIZATION OF HUMAN HER2 GENE TO PREDICT THE BREAST CANCER ASSOCIATED BIOMARKERS

HER2 gene hyperactivation is responsible for the incidence of invasive breast cancer (BC). It leads to over expression of HER2 receptors in breast cells resulting in abnormal receptor dimerization and signal transduction cascade initiation. It ultimately causes uncontrolled proliferation of breast cells. Therefore, this is the most susceptible target gene for BC treatment. HER2 gene transcript with accession ID ENST00000269571.10 was selected for association analysis in present study. A total of ten single nucleotide polymorphisms (SNPs) of gene were analyzed using in-silico tools like CELLO, PROTPARAM, SOPMA, and SWISSMODEL. These tools helped in determining the effect of variants on sub-cellular localization, physiochemical properties, and secondary (2D) and tertiary (3D) structures of mutated proteins. Variants rs1249755832, rs769703068, rs1567913219, and rs1213602748 altered the localization of mutated proteins. SNPs rs1213602748, rs769703068, and rs1249755832 considerably changed the isoelectric point (pI), extinction coefficient, instability index, aliphatic index, and GRAVY. Variant rs769703068 caused a change in all four properties of the 2D structure, i.e., alpha helix, extended strand, beta-turn, and random coil. Polymorphisms that caused a significant deviation in 3D configuration were rs769703068, rs144434331, and rs146177313. These variants can be recommended as biomarkers for HER2+ BC cell diagnosis.

Advanced breast cancer diagnosis: Multiplex RT-qPCR for precise typing and angiogenesis profiling

Breast cancer (BC) remains the foremost cause of cancer-related mortality, with an estimated 2.3 million new cases anticipated globally. The timely diagnosis of BC is pivotal for effective treatment. Currently, BC diagnosis predominantly relies on Immunohistochemistry (IHC), a method known for its sluggishness, expense, and dependence on proficient pathologists for confident cancer typing. In this study, we introduce a novel approach to enhance the accuracy, speed, and cost-effectiveness of BC diagnosis. We employ multiplex Reverse Transcription quantitative Polymerase Chain Reaction (RT-qPCR) with touch-down methods, which consistently yield significantly lower Cycle Threshold (CT) values. The study evaluates gene expression profiles of HER2, PGR, ESR, and Ki67 genes across 61 samples representing four BC subtypes, using RPL13A as the endogenous control gene. The results demonstrate that our method offers remarkable precision, nearly equivalent to IHC, in detecting gene expressions vital for BC diagnosis and subtyping. Moreover, we explore the gene expression of Hif1A, ANG, and VEGFR genes involved in angiogenesis, shedding light on the metastatic potential of the tested BC tumours. Notably, numerous samples exhibit elevated levels of Hif1A and VEGFR, indicating their potential as valuable biomarkers for assessing metastatic status. Collectively, our RT-qPCR methodology emerges as a powerful diagnostic tool for swiftly identifying BC subtypes and can be complemented with other essential tumorigenic biomarker assessments, such as angiogenesis, to further refine cancer characterisation and inform personalised therapeutic strategies for BC patients. This innovation holds the promise of revolutionising BC diagnosis and treatment, offering expedited and reliable insights for improved patient care.

HER2 amplification level by in situ hybridization predicts survival outcome in advanced HER2-positive breast cancer treated with pertuzumab, trastuzumab, and docetaxel regardless of HER2 IHC results

BackgroundThe role of HER2 amplification level in predicting the effectiveness of HER2-directed therapies has been established. However, its association with survival outcomes in advanced HER2-positive breast cancer treated with dual HER2-blockade remains unexplored.MethodsThis is a single-center retrospective study of patients with advanced HER2-positive breast cancer treated with first-line pertuzumab, trastuzumab, and docetaxel. The primary objective was to ascertain the relationship between treatment outcomes and the level of HER2 amplification by in situ hybridization (ISH).ResultsA total of 152 patients were included with a median follow-up duration of 50.0 months. Among the 78 patients who received ISH, a higher HER2/CEP17 ratio correlated significantly with longer PFS (HR 0.50, p = 0.022) and OS (HR 0.28, p = 0.014) when dichotomized by the median. A higher HER2 copy number also correlated significantly with better PFS (HR 0.35, p < 0.001) and OS (HR 0.27, p = 0.009). In multivariate analysis, the HER2/CEP17 ratio was an independent predictive factor for PFS (HR 0.66, p = 0.004) and potentially for OS (HR 0.64, p = 0.054), along with HER2 copy number (PFS HR 0.85, p = 0.004; OS HR 0.84, p = 0.049). Furthermore, the correlation between HER2 amplification level by ISH with PFS and OS was consistent across the HER2 IHC 1+/2+ and 3+ categories.ConclusionsThis is the first study to report that a higher level of HER2 amplification by ISH is associated with improved PFS and OS in advanced HER2-positive breast cancer treated with dual HER2-blockade. Notably, HER2 amplification level had a predictive role regardless of IHC results. Even in patients with HER2 protein expression of 3+, treatment outcome to HER2-directed therapy was dependent on the level of HER2 gene amplification.

Design, synthesis and bioactivity evaluation of novel fusion peptides and their CPT conjugates inducing effective anti-tumor responses on HER2 positive tumors

Human epidermal growth factor receptor 2 (HER2) represents an ideal target for antibody drug development, abnormal expression of the HER2 gene is associated with multiple tumor types. Pertuzumab, as the first monoclonal antibody inhibitor of HER2 dimerization, has been FDA-approved for HER2-positive patients. In order to enhance the activity of HER2-targeted peptide-drug conjugates (PDCs) developed based on pertuzumab, a novel class of conjugates 1–9 was designed and synthesized by fusing the N-terminal peptide sequence of the second mitochondria-derived activator of caspases (SMAC) with P1, followed by conjugation with CPT molecules. Compound 4 exhibited excellent in vitro anti-tumor activity across the three HER2-positive cell lines, comparable to the activity of CPT. Apoptosis induction assays indicated that the synergistic effect of the SMAC sequence enhanced the pro-apoptotic activity of the conjugate. Western Blot analysis and Caspase activity studies validated the mechanism through which SMAC peptides, in synergy with CPT, enhance the activity of PDCs. In vivo studies demonstrated that compound 4 possesses superior anti-tumor activity compared to CPT and can effectively mitigate potential renal toxicity associated with free SMAC peptides. In conclusion, conjugate 4 exhibited excellent anti-tumor activity both in vitro and in vivo, offering potential for further development as a novel peptide-conjugated drug.

Multimodal MRI for Estimating Her-2 Gene Expression in Endometrial Cancer.

To assess the value of multimodal MRI, including amide proton transfer-weighted imaging (APT), diffusion kurtosis imaging (DKI), and T2 mapping sequences for estimating human epidermal growth factor receptor-2 (Her-2) expression in patients with endometrial cancer (EC). A total of 54 patients with EC who underwent multimodal pelvic MRI followed by biopsy were retrospectively selected and divided into the Her-2 positive (n = 24) and Her-2 negative (n = 30) groups. Her-2 expression was confirmed by immunohistochemistry (IHC). Two observers measured APT, mean kurtosis (MK), mean diffusivity (MD), and T2 values for EC lesions. The Her-2 (+) group showed higher APT values and lower MD and T2 values than the Her-2 (-) group (all p < 0.05); there was no significant difference in MK values (p > 0.05). The area under the receiver operating characteristic curve (AUC) of APT, MD, T2, APT + T2, APT + MD, T2 + MD, and APT + MD + T2 models to identify the two groups of cases were 0.824, 0.695, 0.721, 0.824, 0.858, 0.782, and 0.860, respectively, and the diagnostic efficacy after combined APT + MD + T2 value was significantly higher than those of MD and T2 values individually (p = 0.018, 0.028); the diagnostic efficacy of the combination of APT + T2 values was significantly higher than that of T2 values separately (p = 0.028). Weak negative correlations were observed between APT and T2 values (r = -0.365, p = 0.007), moderate negative correlations between APT and MD values (r = -0.560, p < 0.001), and weak positive correlations between MD and T2 values (r = 0.336, p = 0.013). The APT values were independent predictors for assessing Her-2 expression in EC patients. The APT, DKI, and T2 mapping sequences can be used to preoperatively assess the Her-2 expression in EC, which can contribute to more precise treatment for clinical preoperative.

Hybrid Membrane Camouflaged Chemodrug-Gene Nanoparticles for Enhanced Combination Therapy of Ovarian Cancer.

Recently, cell membrane camouflaged nanoparticles (NPs) endowed with natural cellular functions have been extensively studied in various biomedical fields. However, there are few reports about such biomimetic NPs used to codeliver chemodrug and genes for synergistic cancer treatment up to now. Herein, we first prepare chemodrug-gene nanoparticles (Mito-Her2 NPs) by the electrostatic interaction coself-assembly of mitoxantrone hydrochloride (Mito) and human epidermal growth factor receptor-2 antisense oligonucleotide (Her2 ASO). Then, Mito-Her2 NPs are coated by a hybrid membrane (RSHM), consisting of the red blood cell membrane (RBCM) and the SKOV3 ovarian cancer cell membrane (SCM), to produce biomimetic chemodrug-gene nanoparticles (Mito-Her2@RSHM NPs) for combination therapy of ovarian cancer. Mito-Her2@RSHM NPs integrate the advantages of RBCM (e.g., good immune evasion capability and long circulation lifetime in the blood) and SCM (e.g., highly specific cognate recognition) together and improve the anticancer efficacy of Mito-Her2 NPs. The results show that Mito-Her2@RSHM NPs can be devoured by SKOV3 ovarian cancer cells and effectively degraded to release Her2 ASOs and Mito simultaneously. Her2 ASOs can inhibit the expression of endogenous Her2 genes and recover cancer cells' sensitivity to Mito, which ultimately led to a high apoptosis rate of 75.7% in vitro. Mito-Her2@RSHM NPs also show a high tumor suppression rate of 83.33 ± 4.16% in vivo without significant damage to normal tissues. In summary, Mito-Her2@RSHM NPs would be expected as a versatile and safe nanodrug delivery platform with high efficiency for chemo-gene combined cancer treatment.

Abstract C035: Chemotherapy plus HER2-targeted therapy versus endocrine therapy plus HER2-targeted therapy as first-line treatment in patients with HR+/HER2+advanced breast cancer: A real-world trial

Abstract Purpose:This study is a real-world research to compare the efficacy of using chemotherapy plus anti-HER2 therapy versus endocrine therapy plus anti-HER2 therapy as first-line treatment for HR+/HER+ advanced breast cancer patients, aiming at providing a better treatment option for patients. Patients and methods: This study enrolled patients with advanced breast cancer who had complete medical records and were treated in the breast oncology department of Peking University Cancer Hospital from October 2010 to October 2022. These patients were pathologically confirmed with HR+/HER2+ and underwent first-line treatment with anti-HER2 therapy + endocrine therapy (HE group) or anti-HER2 therapy + chemotherapy (with/without endocrine maintenance therapy) (HC group). HER2 positive was defined as HER2+++ or HER2++ on immunohistochemical staining of tumor tissue and fluorescence in situ hybridization (FISH) testing showing HER2 gene amplification in patients. HR+ was considered when more than 1% of tumor cells expressed estrogen receptor (ER) and/or progesterone receptor (PR). The primary endpoints being investigated are progression-free survival (PFS) and overall survival (OS). The second endpoint is objective response rate (ORR) and clinical benefit rate (CBR). Results:From October 2010 to October 2022, a total of 181 patients who met the inclusion criteria were enrolled in the study. Among them, 18 patients received endocrine therapy + anti-HER2 therapy in the first-line treatment stage, while 163 patients received chemotherapy + anti-HER2 therapy (70 patients underwent endocrine maintenance therapy). The median follow-up time was 67 months. The median PFS (mPFS) of the patients in HC group and the HE group were 14.32 months (95% confidence interval (CI) 12.83-15.81) and 9.78 months (95%CI 4.48-15.8), respectively (p=0.299), and no significant statistical difference was observed. Similarly, no significant statistical difference was achieved in the median OS (mOS) of the HC group and the HE group patients, with 126.87 months (95%CI 102.08-151.658) versus 96.00 months (95%CI 78.41-113.59) (p=0.076). In the HC group, the mPFS of patients who received endocrine maintenance therapy and those who did not receive endocrine therapy were 16.24 months (95%CI 12.11-20.37) and 12.36 months (95%CI 10.26-14.46), respectively. The ORR of the two groups of patients were 46.6% and 16.7% (p=0.0015), respectively. Conclusion: The analysis demonstrated that the combination of anti-HER2 therapy with endocrine therapy is not inferior to the combination of anti-HER2 therapy with chemotherapy as first-line treatment for HR+/HER2+ advanced breast cancer. It provides compelling evidence to support the selection of endocrine therapy plus HER2-targeted therapy as a first-line treatment strategy that that enhances the quality of life for patients without shortening their survival time. Citation Format: Yuehua Liang, Hui Ping Li, Zehui Yun. Chemotherapy plus HER2-targeted therapy versus endocrine therapy plus HER2-targeted therapy as first-line treatment in patients with HR+/HER2+advanced breast cancer: A real-world trial [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C035.

Decreased HER2 expression in endometrial cancer following anti-HER2 therapy.

Trastuzumab has demonstrated clinical efficacy in the treatment of HER2-positive serous endometrial cancer (EC), which led to its incorporation into standard-of-care management of this aggressive disease. Acquired resistance remains an important challenge, however, and its underlying mechanisms in EC are unknown. To define the molecular changes that occur in response to anti-HER2 therapy in EC, targeted next-generation sequencing (NGS), HER2 immunohistochemistry (IHC), and fluorescence in situ hybridization (FISH) were performed on pre- and post-treatment tumour samples from 14 patients with EC treated with trastuzumab or trastuzumab emtansine. Recurrent tumours after anti-HER2 therapy acquired additional genetic alterations compared with matched pre-treatment ECs and frequently showed decreased HER2 protein expression by IHC (7/14, 50%). Complete/near-complete absence of HER2 protein expression (score 0/1+) observed post-treatment (4/14, 29%) was associated with retained HER2 gene amplification (n = 3) or copy number neutral status (n = 1). Whole-exome sequencing performed on primary and recurrent tumours from the latter case, which exhibited genetic heterogeneity of HER2 amplification in the primary tumour, revealed selection of an early HER2-non-amplified clone following therapy. Our findings demonstrate that loss of target expression, by selection of HER2-non-amplified clones or, more commonly, by downregulation of expression, may constitute a mechanism of resistance to anti-HER2 therapy in HER2-positive EC. © 2023 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Stemness signature and targeted therapeutic drugs identification for Triple Negative Breast Cancer

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer and carries the worst prognosis, characterized by the lack of progesterone, estrogen, and HER2 gene expression. This study aimed to analyze cancer stemness-related gene signature to determine patients’ risk stratification and prognosis feature with TNBC. Here one-class logistic regression (OCLR) algorithm was applied to compute the stemness index of TNBC patients. Cox and LASSO regression analysis was performed on stemness-index related genes to establish 16 genes-based prognostic signature, and their predictive performance was verified in TCGA and METABERIC merged data cohort. We diagnosed the expression level of prognostic genes signature in the tumor immune microenvironment, analyzed the TNBC scRNA-seq GSE176078 dataset, and further validated the expression level of prognostic genes using the HPA database. Finally, the small molecular compounds targeted at the anti-tumor effect of predictive genes were screened by molecular docking; this novel stemness-based prognostic genes signature study could facilitate the prognosis of patients with TNBC and thus provide a feasible therapeutic target for TNBC.

Functional analysis of a putative HER2-associated expressed enhancer, Her2-Enhancer1, in breast cancer cells

HER-2/neu (HER2) is a member of the epidermal growth factor receptors family, encoding a protein with tyrosine kinase activity. Following the gene amplification or increased HER2 transcription, carcinogenesis has been observed in some cancers. Genetic and epigenetic changes occurring in enhancer sequences can deeply affect the expression and transcriptional regulation of downstream genes, which can cause some physiological and pathological changes, including tumor progression. A therapeutic approach that directly targets the genomic sequence alterations is of high importance, with low side effects on healthy cells. Here, we employed the CRISPR/Cas9 method to genetically knockout an expressed putative enhancer (GH17J039694; we coined it as Her2-Enhancer1) located within the HER2 gene, 17q12: 39,694,339–39,697,219 (UCSC-hg38). We then investigated the potential regulatory effect of Her2-Enhancer1 on HER2 and HER2-interacting genes. To evaluate the cis and trans effects of Her2-Enhancer1, genetic manipulation of this region was performed in HER2-positive and -negative breast cancer cells. Our bioinformatics and real-time PCR data revealed that this putative enhancer region is indeed expressed, and acts as an expressed enhancer. Further functional analysis on edited and unedited cells revealed a significant alteration in the expression of HER2 variants, as well as some other target genes of HER2. Moreover, the apoptosis rate was considerably elevated within the edited cells. As we expected, Western blot analysis confirmed a reduction in protein levels of HER2, GRB7, the gene interacting with HER2, and P-AKT in the PI3K/AKT pathway. Altogether, our findings revealed an enhancer regulatory role for Her2-Enhancer1 on HER2 and HER2-interacting genes; and that this region has a potential for targeted therapy of HER2-positive cancers.

Correlation on HER2 gene amplification by fluorescence in situ hybridization and membrane integrity expression pattern by immunohistochemistry in breast cancer

Correlation on HER2 gene amplification by fluorescence in situ hybridization and membrane integrity expression pattern by immunohistochemistry in breast cancer

Liquid biopsy based HER2 amplification status in gastric cancer patients indicates clinical response

Gastric carcinomas are among the most common cancers in Germany, with approximately 18,000 new cases per year.About 10 years ago, based on results of the Trastuzumab for gastric cancer (ToGA) trial, the addition of the monoclonal antibody trastuzumab to a platinum-fluoropyrimidine chemotherapy backbone became the standard-of-care 1st-line therapy for human epidermal growth factor receptor 2 (HER2)-positive gastric cancers. Only patients with primary HER2 gene amplification benefit from this therapy. Thus, accurate HER2 gene amplification detection is predictive and critical for therapy selection. As a gold standard the HER2 status is currently determined in tumor tissue specimens using immune histochemistry and fluorescent in situ hybridisation. However, HER2 amplification is detectable in only about 20 % of gastric carcinomas. The recent approval of an antibody-drug conjugate Trastuzumab deruxtecan (T-DXd) and the establishment of a new subgroup of HER2-low tumors due to the bystander effect associated with T-DXd increases the relevance of precise HER2 diagnostics.Aim of this analysis was to determine the HER2 amplification status from circulating DNA fragments in blood using a HER2 Copy Number Variation assay to establish a minimal invasive approach.For the present study, a digital droplet PCR-based method was validated relative to established tissue-based methods. Furthermore and most importantly, the changes of HER2 status during therapy were investigated in seven patients indicating that the changes of HER2 status and number of HER2 copies detected in blood can reflect on therapy efficiency and uncover treatment resistance.