anti-HER2 Antibody-drug Conjugate Research Articles

A multicenter, phase II trial of RC48-ADC combined with radiotherapy, PD-1/PD-L1 inhibitor, GM-CSF, and sequential IL-2 (PRaG3.0 regimen) for salvage therapy in patients with HER2-expressing advanced solid tumors.

e14614 Background: Cancer immunotherapy has emerged as one of the most promising approaches in many kinds of advanced tumors. The PRaG therapy is an innovative cancer immunotherapy, when combined with radiotherapy, PD-1/L1 inhibitor and GM-CSF to generate the desired in situ vaccine effect to activate the immune response and modulate the tumor microenvironment. Previous studies have showed encouraging efficacy of the PRaG regimen in the treatment of advanced refractory tumors. RC48-ADC, an anti-HER2 antibody-drug conjugate with MMAE as cytotoxic payload has been recently demonstrated that ADCs are also able to induce immunogenic cell death and widespread release of cancer cell antigens, synergize with immunotherapy by promoting effector T-cell activation. The aim of this study is to explore efficacy and safety of PRaG3.0 regimen for therapy of HER2-expressing advanced solid tumors. Methods: Participants with advanced, confirmed HER2-expressing (IHC3+, 2+ or 1+) solid tumors that had progressed after standard of care, or intolerance were enrolled. In the cycle, those received RC48-ADC (2.0 mg/kg d1, every 3 weeks), then HFRT (2-3 doses of 5-8Gy) was delivered for one metastatic lesion every other day, followed by GM-CSF (200 μg d3-7), sequential IL-2 (2million IU d8-12), and PD-1/L1 inhibitor was dosing within one week after completion of HFRT. After RC48-ADC combined with PD-1/L1 inhibitor sequential cytokines for at least 6 cycles, then maintenance with PD-1/PD-L1 inhibitor was administered until disease progression or unacceptable toxicity. The primary endpoint was objective response rate (ORR). Results: As of Jan 2023, a total of 32 patients (n=6 for gynecological cancer, n=5 for pancreatic cancer, n=21 for other cancers) were enrolled, 26 of them completed at least 1 tumor assessment. The ORR was 38.5%, and the disease control rate was 69.2%. The ORR was 66.7% in gynecological cancer, 25.0% in pancreatic cancer, and 31.3% in other cancers. Notably, patients who were HER2 IHC1+ responded similarly to those who were IHC2+~3+, with ORR of 43.8% and 30.0%, respectively. Median progression-free survival was 7.2 months (95%CI: 4.9, 9.5). The most common treatment-related adverse events (TRAEs) included fatigue (28.1%), fever (28.1%), alopecia (28.1%) and anorexia (18.8%). Grade ≥3 TRAEs occurred in two patients (6.3%). In addition, activated plasmacytoid dendritic cell was significantly higher at baseline in responders (CR+PR) vs. nonresponders (SD+PD). Conclusions: The schedule of RC48-ADC was changed from once every 2 weeks to once every 3 weeks, and was still effective with significantly reduced side effects. These preliminary results suggest that PRaG3.0 regimen has a manageable safety profile and enhancing potential sensitivity in pretreated patients with HER2-expressing cancers. Clinical trial information: NCT05115500 .

Comparing digital and glass slide methods for determining the HER2-low and negative categories in breast cancer.

e13042 Background: The promising clinical use of anti-HER2 antibody–drug conjugates (ADCs) in patients with immunohistochemistry (IHC)-based HER2-Low (1+, and 2+/FISH−) breast cancer has been discovered recently. Thus, pathologists and clinicians are now under increasing pressure to discriminate HER2-low from HER2-0. Methods: This multi-institutional retrospective breast cancer study selected 60 cases of paired digital and glass slides with HER2-Low (IHC 1+, and 2+/FISH−) and HER2-0 according to the original pathology reports, including 20 cases per staining intensity. These cases were re-evaluated by 37 pathologists from three hospitals. After excluding those cases with HER2 ≥ 2+ confirmed by both digital and glass slides with ≥ 85% agreement across 37 pathologists, the remaining cases were reclassified into “HER2-Neg” and “HER2-Low” categories with ≥ 85% agreement. If a slide could not be placed into one of these two categories, it will be labeled “HER2-Ambiguous”. Results: After excluding 13 cases with HER2 ≥ 2+ confirmed by both digital and glass slides with ≥ 85% agreement, the remaining 47 cases were reclassified into “HER2-Neg”, “HER2-Ambiguous” and “HER2-Low” categories. Therefore, the Kappa value for these three categories between the digital and glass slides was 0.597 (p = 2.50e-07). Interestingly, our study demonstrated that a significantly higher proportion of HER2-Low cases were identified on digital slides (48.9%, 23/47) than on glass slides (38.3%, 18/47; BhapkarTest, p = 3.17e-04). Five of the twelve (41.7%) HER2-Ambiguous cases diagnosed using glass slides were reclassified as HER2-Low using digital slides. Conclusions: Our study indicated that, compared to glass slides, digital slides may tend to shift HER2-0 to HER2-low category, potentially increasing breast cancer patients access to anti-HER2 ADCs, although the clinical benefit and biological characteristics have not been fully investigated. Thus, further investigation is warranted.

Disitamab vedotin, a novel humanized anti-HER2 antibody-drug conjugate (ADC), combined with toripalimab in patients with locally advanced or metastatic urothelial carcinoma: An open-label phase 1b/2 study.

4566 Background: Disitamab vedotin has shown promising data across a spectrum of HER2 expression in patients with locally advanced or metastatic urothelial carcinoma (la/mUC) who progressed on chemotherapy. This study was conducted to evaluate the safety and efficacy of disitamab vedotin plus anti-PD-1 antibody in advanced urothelial carcinoma. Methods: This is an open-label, multicenter, phase 1b/2 trial to evaluate the safety and activity of RC48-ADC combined with toripalimab, a humanized immunoglobin G4 monoclonal antibody against PD-1 in mUC. Patients received RC48-ADC at 1.5 or 2 mg/kg, in combination with 3mg/kg toripalimab every two weeks in a dose-escalation and expansion cohort until confirmed disease progression assessed by the investigators, unacceptable toxicity, or voluntary withdrawal. The primary endpoint was safety; secondary endpoints included efficacy and tumor tissue biomarkers. Results: Forty-one la/mUC patients were enrolled (22 males; median age 66 y [42-76]; 61% treatment-naïve). 54% of patients had visceral metastases (mets), including 24% with liver mets. The primary site was in upper tract UC in 54%. HER2 expression IHC 2+ or 3+ was in 59% patients, and PD-L1 positive in 32%. No dose-limiting toxicity was observed and the recommended dose was RC48-ADC 2mg/kg plus toripalimab 3mg/kg every two weeks. By the cutoff date of 18 November 2022, the confirmed ORR was 73.2% (95%CI: 57.1, 85.8), including 9.8% CR. The ORR was 76.0% for treatment-naïve patients. In HER2 IHC 3+/2+, IHC 1+, and IHC 0 subgroups, the ORR was 83.3%, 64.3%, and 33.3%, respectively. The ORR was 61.5% and 78.6% in PD-L1 positive and negative subgroups. DCR was 90.2% (95% CI, 76.9–97.3), with a median progression-free survival (PFS) of 9.2 months (95%CI: 5.7-10.3) and 2-year overall survival (OS) rate of 63.2%. All patients experienced treatment-related adverse events (TRAEs). The most common TRAEs were AST/ALT increase (68.3%), peripheral sensory neuropathy (61.0%), asthenia (61.0%), γ-glutamyl transferase increase (56.1%), hypertriglyceridemia (53.7%), and appetite decrease (51.2%). Grade 3 or greater TRAEs occurred in 43.9% of patients. Twenty-three patients (56.1%) had immune-related AEs (14.6% ≥ G3), including immune-related skin reactions, hyperglycemia, pneumonitis, hepatitis, and myositis. Conclusions: RC48-ADC in combination with toripalimab demonstrated promising efficacy in patients with la/mUC and a manageable safety profile. A phase 3 study is currently ongoing to compare the safety and efficacy of this regimen with standard of care. Clinical trial information: NCT04264936 .

ADC vs HP dual-antibody for HER2-positive advanced breast cancer after failed to TKI.

e13036 Background: The application of tyrosine kinase inhibitor (TKI) is higher than that of antibody-drug conjugate (ADC) in the second-line anti-HER2 targeted therapy of patients with advanced breast cancer in China. The purpose of this study is to explore the efficacy and safety of the new generation anti-HER2 ADC, the previous generation ADC(T-DM1) and HP dual-antibody (trastuzumab + pertuzumab) in patients with HER2 positive advanced breast cancer who failed TKI therapy. Methods: This is a retrospective, nonrandomized, controlled study to compare the regimen of the new generation ADC, HP dual-antibody and T-DM1 for patients who failed TKI treatment. The primary end point was progression-free survival (PFS). The secondary end points included the overall response rate (ORR), the clinical benefit rate (CBR) and the safety. Results: A total of 213 patients were enrolled from January 2013 to June 2022. All patients were from the department of oncology, the fifth medical center of Chinese PLA General Hospital. The median age of all patients was 47y. Among these patients,73 cases were in the new generation ADC group (including DS8201 30 cases, MRG002 24 cases, ARX788 10 cases and RC48 9 cases), 71 cases were in T-DM1 group and 69 cases were in HP group (combination chemotherapy including taxane, vinorelbine, gemcitabine and capecitabine). The median PFS of the patients in the new generation ADC group , HP group and T-DM1 group was 7.0m, 6.6m, 4.0m respectively. The difference between the new generation ADC group and T-DM1 group was significant (HR=0.44,95%CI 0.30-0.66, P<0.0001), which between HP group and T-DM1 group was also significant (HR=0.50,95%CI 0.34-0.74, P<0.0001), meanwhile, which between the new generation ADC group and HP group was not significantly (HR=0.88, 95%CI 0.58-1.32, P=0.527)(Figure 1). The ORR in three groups was 54.8%, 27.5%, 22.5% respectively, and the new generation ADC group was significantly higher than the other two groups (P<0.0001). The CBR in three groups was 65.8%, 62.3%, 47.9% respectively. The difference between the new generation ADC group and HP group was not significantly (P=0.67), and both were significantly higher than T-DM1 group (P=0.03). The most common grade 3 or 4 adverse event in the new generation ADC group was neutropenia (20.5%) and which in T-DM1 group was thrombocytopenia (28.1%). The incidence of interstitial lung disease in the two groups was 5.4% and 1.4% respectively. No grade 5 adverse event occurred. Conclusions: In our study, the new generation ADC significantly improved PFS and ORR compared with that for T-DM1 with manageable toxicity, so it can be considered an optimized subsequent treatment for HER2 positive advanced breast cancer after TKI failure. In addition, there was no significant difference in PFS and CBR between HP and the new generation ADC, thus HP dual-antibody regimen is also an optional treatment for patients who never used pertuzumab.

Safety and efficacy of DB-1303 in patients with advanced/metastatic solid tumors: A multicenter, open-label, first-in-human, phase 1/2a study.

3023 Background: DB-1303 is an antibody-drug conjugate (ADC) consisting of a humanized anti-HER2 IgG1 monoclonal antibody, covalently linked to proprietary DNA topoisomerase I inhibitor (P1003) via a maleimide tetrapeptide-based cleavable linker, with a high drug-to-antibody ratio (~8). Methods: This is a global first-in-human, dose-escalation and -expansion study in patients (pts) with advanced/metastatic solid tumors. Pts (ECOG 0-1) with HER2 – (high or low) expressing or mutant cancers who failed previously systemic therapies were recruited and received DB-1303 intravenously Q3W as monotherapy. The objectives were safety, tolerability, maximum tolerated dose (MTD) or recommended phase 2 dose (PR2D), pharmacokinetics (PK), and preliminary antitumor activity. Here we report the results from dose-escalation. Results: As of Jan 13, 2023, 85 pts received DB-1303 at 6 dose levels (2.2, 4.4, 6.0, 7.0, 8.0, and 10.0 mg/kg) and received a median of 7 (range, 1-27) prior lines of therapy, including previous anti-HER2 ADC therapy in 32.9% of pts. The median duration of treatment was 63.0 (range, 21-211) days, and 68 pts (80.0%) remained on treatment. Treatment-emergent adverse events (TEAEs) and ≥ grade (G) 3 TEAEs occurred in 74 pts (87.1%) and 18 pts (21.2%), respectively; the most common TEAEs were nausea (51.8%, 3.5% ≥ G3), vomiting (43.5%, 1.2% ≥ G3), platelet count decreased (35.3%, 3.5% ≥ G3), and anemia (29.4%, 5.9% ≥ G3). Few pts experienced neutropenia (10 [11.8%]) and alopecia (3 [3.5%]). There was no DLT or TEAEs leading to death. Interstitial lung disease occurred in 2 pts (2.4%, G1), without any ≥ G2. The exposure parameters (Cmax and AUC) of DB-1303 ADC increased with ascending doses from 2.2 to 10.0 mg/kg. The half-life of DB-1303 ADC is approximately 6-7 days for 6.0-8.0 mg/kg dose cohorts. The exposure of serum release payload was magnitudes lower than that of DB-1303 ADC, demonstrating stability of the ADC in systemic circulation. A total of 52 pts had undergone at least one post-baseline tumor scan. Twenty-three pts (44.2%, 23/52) had objective partial tumor response per RECIST 1.1: Thirteen HER2+ breast cancer (BC) (50.0%, 13/26, including 5 pts with brain metastases [55.6%, 5/9]), 5 HER2 low BC (38.5%, 5/13), 2 colorectal cancer (66.7%, 2/3), 1 endometrial carcinoma (33.3%, 1/3), 1 esophageal cancer (50.0%, 1/2), and 1 ovarian cancer (50.0%, 1/2). Among all the pts, the DCR was 88.5% (46/52); for pts with HER2+ BC and HER2-low BC, the DCRs were 96.2% (25/26) and 84.6% (11/13), respectively. Conclusions: DB-1303 was well tolerated with encouraging preliminary antitumor activity in heavily pretreated pts with advanced/metastatic solid tumors, especially in pts with HER2+ BC and brain metastasis as well as in HER2-low BC. Expansion is ongoing in selected tumor pts treated at the RP2D. Clinical trial information: NCT05150691 .

Novel Anti-HER2 Antibody-Drug Conjugates Versus T-DM1 for HER2-Positive Metastatic Breast Cancer After Tyrosine Kinase Inhibitors Treatment.

Antibody-drug conjugates (ADCs) have been the preferred regimens for human epidermal growth factor receptor 2 (HER2)-positive metastatic breast cancer (MBC) after trastuzumab. Unfortunately, there is little data showing which ADCs should be chosen for those patients whose treatment with tyrosine kinase inhibitors (TKIs) failed. This study aims to analyze the efficacy and safety between novel anti-HER2 ADCs and trastuzumab emtansine (T-DM1) for those with TKIs failure. HER2-positive MBC using ADCs from January 2013 to June 2022 were included, and all of them were treated with TKIs. The primary study endpoint was progression-free survival (PFS), and the secondary study endpoints were objective response rate (ORR), clinical benefit rate (CBR), and safety. A total of 144 patients with 73 patients in the novel anti-HER2 ADCs group and 71 patients in the T-DM1 group. In these novel ADCs, 30 patients received trastuzumab deruxtecan (T-Dxd), 43 patients receive other novel ADCs. The median PFS in the novel ADCs group and T-DM1 group were 7.0 months versus 4.0 months, respectively, and ORR was 54.8% versus 22.5%, CBR was 65.8% versus 47.9%, respectively. In subgroups analysis, the PFS were both significantly improved in patients receiving T-Dxd and other novel ADCs compared with T-DM1. The most common grades 3-4 adverse events in the novel anti-HER-2 ADCs group were neutropenia (20.5%) and thrombocytopenia (28.1%) in the T-DM1 group. In patients with HER2-positive MBC previously treated with TKIs, both T-Dxd and other novel anti-HER2 ADCs yielded statistically significant better PFS than T-DM1 did, with tolerable toxicities.

Abstract 3160: A rapid, non-invasive test to determine HER2-low status in triple negative breast cancer (TNBC) patients

Abstract Background: TNBC is defined as a HER2-negative disease, as assessed via immunohistochemistry (IHC). An IHC 3+ score is considered positive and 2+ (with a negative ISH), 1+, and zero scores are considered negative. Recently, HER2-low (1+, 2+) breast cancer has been proposed as a targetable disease distinct from TNBC. Indeed, use of an anti-HER2 antibody drug conjugate (ADC) in HER2-low metastatic patients resulted in improved progression free and overall survival (DESTINY-Breast04), and remarkably generated a response in HER2-zero cases. The potential to expand the current treatment for HER2-low and -zero patients, highlights the limitations of traditional HER2 assessment, including HER2 intra-tumoral heterogeneity, differential IHC pathology staining, and inter-reader variability. We therefore sought to address how to optimally identify HER2-low TNBC patients that may benefit from HER2 ADCs. Methods: We developed a rapid, non-invasive test to determine HER2-low status in TNBC tumors. Our test uses radiological imaging (DCE-MRIs) coupled with the biophysical simulation platform TumorScope and is informed by single cell RNA-sequencing data to connect molecular regulation to the tumor microenvironment in individual patients. We leveraged these relationships to design a rapid, non-invasive test, HER2Scope, that captures the fraction of each TNBC tumor likely to express significant levels of HER2 (a “HER2 score”). We then characterized the HER2 score in a cohort of TNBC patients from our Virtual Tumor Bank, leading us to identify 3 distinct cohorts: HER2-negative, -low, and -moderate tumors. Results: Using our platform, we identified that HER2 expression is associated with less proliferation and lower nutrient use. Interestingly, we found that there was significant overlap in HER2 expression between HER2+ and TNBC tumors. HER2Scope-identified HER2-low and- moderate tumors were associated with a lower likelihood of pCR in response to chemotherapy compared to HER2-zero tumors (25.3% in HER2-moderate vs. 40% in HER2-zero), suggesting a more aggressive and less drug-responsive phenotype in HER2-low and -moderate tumors. We then tested the expected rate of HER2-low and -moderate tumors within TNBC tumors using our Virtual Tumor Bank (n=264) and found a HER2-low and -moderate response rate of 56% (95% CI = 49-62%), which is strikingly similar to the TNBC response rate to trastuzumab deruxtecan (TDXd) reported in the DAISY trial (42%). HER2Scope may therefore be an effective test to identify TNBC patients for HER2 ADC therapies. Conclusion: We present a novel test to identify HER2-nonzero TNBC patients rapidly and non-invasively. HER2Scope relies on radiological imaging, and therefore does not depend on an inherently variable score derived from biopsy-driven pathologic staining. Additionally, by evaluating the entire tumor, it addresses the challenge of HER2 intra-tumoral heterogeneity. Citation Format: Daniel Cook, Snehal Patel, John Whitman, Nicole Hobbs, Dorys A. Lopez-Ramos, John Pfeiffer, Joseph Peterson, Tushar Pandey, Anuja K. Antony, John Cole. A rapid, non-invasive test to determine HER2-low status in triple negative breast cancer (TNBC) patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3160.

Pathological identification of HER2-low breast cancer: Tips, tricks, and troubleshooting for the optimal test.

The introduction of novel anti-HER2 antibody-drug conjugates (ADC) for the treatment of HER2-low breast cancers has transformed the traditional dichotomy of HER2 status to an expanded spectrum. However, the identification of HER2-low (i.e., immunohistochemistry (IHC) score 1 + or IHC score 2+, without gene amplification) tumors is challenged by methodological and analytical variables that might influence the sensitivity and reproducibility of HER2 testing. To open all possible therapeutic opportunities for HER2-low breast cancer patients the implementation of more accurate and reproducible testing strategies is mandatory. Here, we provide an overview of the existing barriers that may trouble HER2-low identification in breast cancer and discuss practical solutions that could enhance HER-low assessment.

Abstract HER2-16: HER2-16 Inter-lesion heterogeneity of HER2-status in metastatic breast cancer: possible implications for treatment with anti-HER2 antibody-drug conjugates

Abstract Background. Trastuzumab deruxtecan (T-DXd) has shown promising activity in patients with HER2-low metastatic breast cancer. As the HER2-status can vary between the primary and its corresponding metastases, treatment decisions should ideally be based on HER2 assessment of a recent biopsy. However, limited data is available on intra-patient inter-metastatic heterogeneity in HER2-status, affecting representability of a single biopsy and potential therapeutic options and outcome. We therefore assessed HER2 status on multiple metastases from patients with primary ER-positive/HER2-non-amplified breast cancer in our prospective post-mortem tissue donation program UPTIDER (NCT04531696). Methods. Ninety-one metastatic samples retrieved during the autopsies of 6 patients (range: 13–16/patient) and their respective primary tumours were immunohistochemically (IHC) stained for HER2 (HercepTestTM, RTU, ISO-15189 accredited) in our institution. Consensus scoring was performed between two pathologists according to ASCO/CAP 2018 guidelines. The observers were blinded for patient ID. Reflex fluorescence in situ hybridization (FISH) testing was performed for samples with IHC score of 2+. HER2 status was categorized as HER2-zero (IHC 0), HER2-low (IHC 1+ or IHC 2+ with negative FISH), or HER2-positive (IHC 3+ or IHC 2+ with positive FISH). To assess stability of the performance of IHC scoring in the post-mortem setting, an additional 13 samples taken from 3 metastases at regular (every 1.5h) time intervals during the autopsy underwent HER2 IHC scoring. Results. Evaluation of HER2-status in the primary tumour showed 2 patients with HER2-zero disease and 4 with HER2-low disease. A discordance between HER2 status of the metastases and their respective primary was seen in all patients. Not a single lesion was found to be HER2-positive. For every patient, at least one HER2-low metastasis was observed, with the percentage being highly variable between patients and ranging between 7 and 100%. No association was observed between HER2 status and organ site: HER2-low as well as HER2-zero lesions were found in all organs evaluated in at least 4 patients (liver, bone, pleura, lymph nodes). For 5 patients, multiple lesions within the liver were evaluated: while HER2-zero versus HER2-low status was concordant in those lesions in 4 patients, a mix of HER2 IHC scores was seen in 3 of them. IHC scores were stable over time for tumour lesions assessed repeatedly. Discussion. Important inter-lesion heterogeneity in terms of HER2-low status was observed in patients with primary ER-positive/HER2-non-amplified breast cancer participating to our post-mortem tissue donation program. This observed heterogeneity is unlikely to be due to post-mortem changes in HER2 expression. HER2-low status was found in at least one distant lesion in all patients, complicating therapeutic decision-making based on a single biopsy. Of note, IHC 1+ and 2+ scores varied between metastases of each patient too, making assessment on a single biopsy less reliable for stratification in clinical trials. Further assessment on samples from UPTIDER-patients with ER-negative disease is currently ongoing and results will be available to be presented. Citation Format: Tatjana Geukens, Maxim De Schepper, François Richard, Marion Maetens, Karen Van Baelen, Amena Mahdami, Ha-Linh Nguyen, Edoardo Isnaldi, Sophia Leduc, Anirudh Pabba, Imane Bachir, Freya Mertens, Sara Vander Borght, Ann Smeets, Ines Nevelsteen, Kevin Punie, Patrick Neven, Hans Wildiers, Wouter Van Den Bogaert, Giuseppe Floris, Christine Desmedt. HER2-16 Inter-lesion heterogeneity of HER2-status in metastatic breast cancer: possible implications for treatment with anti-HER2 antibody-drug conjugates. [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr HER2-16.

Abstract HER2-07: HER2-07 Genomic Characterization of Primary and Metastatic HER2-low Breast Cancers

Abstract Background: Varying levels of HER2 expression without ERBB2 gene amplification can be detected by immunohistochemistry (IHC) in approximately 60% of all invasive breast cancers (BCs). HER2-low-expressing BCs have recently been shown to respond to novel anti-HER2 antibody drug conjugates. Several studies have demonstrated that HER2-low BCs do not seem to constitute a distinct clinical and transcriptomic subtype as compared to HER2-0 BCs. Here we sought to define the clinicopathologic features and repertoire of somatic genetic alterations in HER2-low BCs. Methods: We retrieved clinical, pathological, and genomic data of BCs that were subjected to targeted sequencing using the FDA-cleared MSK-IMPACT assay from April 2014 to December 2021. After removing cases where any available biopsy had HER2 3+ and/or positive ERBB2 fluorescence in situ hybridization (FISH), 3608 samples (primary=1347; post-treatment/metastatic=2261) were included. Tumors were classified as HER2-low if they had an HER2 IHC score of 1+ or 2+ with a non-amplified FISH assay and HER2-0 if they had an HER2 IHC score of 0. Somatic mutations and DNA copy number alterations from MSK-IMPACT were analyzed. Multiple testing correction using the Benjamini-Hochberg method was applied to control for the false discovery rate (q). Q values < 0.1 were considered significant. Results: Among 3608 HER2- BCs, 1460 (40%) and 2148 (60%) were HER2-0 and HER2-low, respectively. Hormone receptor (HR) expression was significantly higher in HER2-low than HER2-0 tumors in both primary (781 [68.3%] vs 362 [31.7%]; p< 0.001) and metastatic (1031 [60.5%] vs 673 [39.5%]; p< 0.001) samples. A higher proportion of HER2-low tumors was found in metastatic than primary samples (59% vs 41%; p< 0.001) in this cohort. No difference in histology subtype, tumor grade, disease stage (among primary tumors), mutational signatures, and tumor mutational burden was found overall and when cases were stratified by HR expression. In HR-positive BCs, HER2-0 BCs harbored higher frequency of TP53 (33% vs 25%; odds ratio [OR] 1.49, 95% confidence interval [CI] 1.25-1.78, q< 0.001) and CDKN1A (1% vs 0%; OR 17.47, 95% CI 2.48-756.37, q=0.02) alterations than HER2-low BCs. Similar findings were observed in metastatic but not in primary HR-positive BCs. No differences were detected in HR-negative BCs stratified into HER2-low and HER2-0. Given the potential misclassification that exists between IHC HER2-0 and HER2-1+, we then conducted an exploratory analysis splitting the HER2-low group into HER2 1+ and 2+. TP53 alterations remained significantly enriched in HER2-0 compared to HER2-1+ HR-positive tumors (33% vs 24%; OR 1.55, 95% CI 1.28-1.87, q< 0.001). In HR-positive BCs, HER2-2+ displayed a higher frequency of genetic alterations in genes encoding for transcription factors, such as MYC (14.2% vs 7.3%; OR 2.09, 95% CI 1.44-3.04, q=0.02) and YAP1 (2% vs 0.3%; OR 6.86, 95% CI 1.7-39.57, q=0.1), and DNA damage response, such as FAM175A (1.6% vs 0%; OR 18.23, 95% CI 2.43-807.73, q=0.03) and BRCA2 (4% vs 1%; OR 3.09, 95% CI 1.49-6.55, q=0.1), than HER2-0 tumors. In HR-negative HER2-2+ tumors, a higher frequency of PIK3CA mutations was observed in comparison to HER2-0 (36.9% vs 19.5%; OR 2.41, 1.4-4.1, q=0.1), overall and in the metastatic setting. Conclusions: HER2-low BCs seem not to represent a distinct pathologic subtype. At the genomic level, however, some differences were identified and these became more conspicuous upon subclassification of HER2 IHC expression into 1+ and 2+. Further investigation into methods that more accurately detect and quantify low levels of HER2 expression in BC samples as well as better characterize the biology behind the HER2-low/ultralow expression is warranted. Citation Format: Antonio Marra, Anton Safonov, Joshua Drago, Emanuela Ferraro, Pier Selenica, Andrea Gazzo, Giuseppe Curigliano, Shanu Modi, Pedram Razavi, Jorge Reis-Filho, Sarat Chandarlapaty. HER2-07 Genomic Characterization of Primary and Metastatic HER2-low Breast Cancers [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr HER2-07.

Abstract HER2-02: HER2-02 HER2-Low Status is Associated with Worse Clinical Outcomes in Hormone Receptor-Positive, HER2-Negative Advanced Breast Cancer Patients Treated With First-Line Cyclin-Dependent Kinase 4/6 Inhibitors Plus Endocrine Therapy

Abstract Introduction: Cyclin-dependent kinase 4/6 inhibitors (CDK4/6i) in combination with endocrine therapy (ET) are the standard first-line treatment for patients with hormone receptor-positive, human epidermal growth factor receptor 2 (HER2)-negative advanced breast cancer (HR+/HER2- aBC). HER2-low BC, which is defined by an IHC score for HER2 of 1+ or 2+ with negative ISH assay, accounts for more than half of all HR+/HER2- aBC cases, and it is associated with remarkable clinical benefit from the novel anti-HER2 antibody drug conjugate (ADC) trastuzumab-deruxtecan. Evidence on the prognostic impact of HER2-low status is controversial in both limited-stage and advanced BC. Here, we sought to investigate the possible prognostic relevance of HER2-low status in a population of aBC patients treated with CDK4/6i plus ET. Methods: We conducted a retrospective-prospective study in six Italian Cancer Centers to investigate the impact of HER2 status (low vs. 0) on the progression-free survival (PFS) and overall survival (OS) of consecutive HR+/HER2- aBC patients treated with CDK4/6i plus ET (aromatase inhibitors or fulvestrant) as a first-line therapy. In the main study analysis, we considered HER2 status in the last tumor assessment (i.e., primary tumor, or, when available, a metastatic lesion). We also performed a subgroup analysis including only patients with HER2 status evaluation in a metastatic lesion collected before CDK4/6i plus ET therapy initiation. The association between HER2 status (low vs. 0) and PFS or OS was evaluated using log-rank test and Cox regression modeling. Results: We evaluated 767 consecutive HR+/HER2- aBC patients treated with CDK4/6i plus ET between January 2017 and January 2022. Of these, 436 patients (56.8%) received CDK4/6i plus ET as a first-line therapy, and they were included in this analysis. Median age was 63 years (range 27-87), and 362 patients (83.0%) were postmenopausal. The majority of patients were treated with palbociclib (68.3%), while 91 (20.9%) and 47 (10.8%) patients received ribociclib and abemaciclib, respectively. Regarding HER2 status, 269 (62.9%) patients had HER2-low tumors, while 159 (37.1%) patients had HER2-0 neoplasms. HER2-low status was associated with significantly lower PFS when compared to HER2-0 status [median PFS (mPFS) 23.6 vs. 32.3 months, respectively; p=0.014]. HER2-low status was also associated with significantly worse OS (mOS 48.7 vs 58.3 months, respectively; p=0.025). These results were confirmed in multivariable models adjusting the impact of HER2 status for clinically-relevant covariates, namely estrogen receptor status, Ki-67, age, number of metastatic sites, presence of liver metastases, disease free interval, ECOG Performance Status. In this analysis, HER2-low status, compared with HER2-0 status, was independently associated with worse PFS [adjusted Hazard Ratio (aHR): 1.62; 95% confidence interval (CI): 1.17-2.24; p< 0.01] and OS (aHR: 1.74; 95% CI: 1.09-2.76; p=0.019). Subgroup analysis conducted in the subset of 256 patients with available metastatic tumor samples collected before CDK4/6i plus ET initiation confirmed that HER2-low status (n=157), when compared to HER2-0 status (n=99), was independently associated with worse PFS (mPFS 24.5 vs 35.2 months, p=0.01; aHR 2.07; 95% CI: 1.28-3.34, p< 0.01) and worse OS (mPFS 48.7 vs 72.3 months, p=0.027; aHR 3.12; 95% CI 1.44-6.77, p< 0.01). Conclusions: This multicenter Italian study revealed that HER2-low status has independent, negative prognostic value in patients with HR+/HER2- aBC treated with CDK4/6i plus ET in the first-line setting. Our results suggest that HER2-low status might be associated with different clinical benefit from standard anticancer therapies in specific clinical settings. The definition of treatment algorithms also taking into account HER2 status is a clinical priority in patients with HR+/HER2- aBC. Citation Format: Emma Zattarin, Caterina Sposetti, Rita Leporati, Luigi Mariani, Alice Menichetti, Chiara Corti, Chiara Benvenuti, Giovanni Fucà, Riccardo Lobefaro, Francesca Ligorio, Daniele Presti, Leonardo Provenzano, Andrea Vingiani, Gaia Griguolo, Marianna Sirico, Ottavia Bernocchi, Antonio Marra, Paola Zagami, Elisa Agostinetto, Flavia Jacobs, Pierluigi Di Mauro, Andrea Esposito, Carlo Alberto Giorgi, Luca Lalli, Laura Boldrini, Pier Paolo Maria Berton Giachetti, Ambra Carnevale Schianca, Valentina Guarneri, Rebecca Pedersini, Agnese Losurdo, Alberto Zambelli, Daniele Giulio Generali, Giuseppe Curigliano, Giancarlo Pruneri, Filippo de Braud, Maria Vittoria Dieci, Claudio Vernieri. HER2-02 HER2-Low Status is Associated with Worse Clinical Outcomes in Hormone Receptor-Positive, HER2-Negative Advanced Breast Cancer Patients Treated With First-Line Cyclin-Dependent Kinase 4/6 Inhibitors Plus Endocrine Therapy [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr HER2-02.

Abstract P4-07-33: Differences in Treatment Outcomes Between Patients with HER2-Low versus HER2-Zero, Hormone Receptor-Positive Advanced-Stage Breast Cancer Treated with CDK4/6 Inhibitors

Abstract Background: The cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors transformed the care of patients with hormone receptor-positive (HR+), human epidermal growth factor receptor-2 (HER2)-negative (HR+/HER2−) advanced-stage breast cancer (aBC). Breast cancers with no expression of HER2 are recently classified into two groups: HER2-Zero subtype which include those with HER2-immunohistochemistry (IHC) score of 0 (IHC-0) and “HER2-low”, defined as HER2-IHC score of 1+ or (2+ with negative in situ hybridization (ISH) assay). There is increasing interest in the HER2-low subtype which is becoming a new distinct entity with promising data from recently reported clinical trials using novel anti-HER2 antibody-drug conjugates (ADC) in this subgroup. This study investigates the differences in treatment outcomes between patients with aBC with HER2-low versus those with HER2-Zero (IHC-0) disease treated with CDK4/6 inhibitors and endocrine therapy (ET). Methods: We retrospectively reviewed patients with (HR+/HER2−) aBC who received CDK4/6 inhibitors with an aromatase inhibitor (AI) or fulvestrant from June 2017 to May 2020 at a single cancer center. Data was extracted from patients’ electronic medical records and from our institutional cancer registry. Progression-free survival (PFS), defined as the time from the initiation of CDK4/6 inhibitors to the date of radiological or clinical progression or death, and was estimated by the Kaplan-Meier method and compared by the log-rank test. Multivariate Cox regression modeling was performed with covariates including progesterone receptor (PR) status, prior chemotherapy, site of metastasis (visceral versus bone-only disease), line of treatment (first-line or beyond), menopausal status, age (less than or ≥ 45) and number of metastatic sites (< or ≥ 3). P-value < 0.05 was considered statistically significant. Results: During the study period, a total of 256 patients with advanced-stage breast cancer who received ET and CDK4/6 inhibitors (ribociclib in all patients) were included, median age was 48 (22-87) years. Majority (n=162, 63.3%) received ribociclib as a first-line therapy while the others had it as a second line and beyond. 136 (53.1%) patients had de novo metastatic disease, and 122 (47.7%) were premenopausal. In total, 114 (44.5%) of the patients where HER2-Zero (IHC-0), while 142 (55.5%) others had HER2-low disease as defined above. The overall response rate (ORR) for the HER2-Zero group was 52% versus 39% for the HER2-low group (P= 0.005).The median PFS was 23.0 (95% confidence interval [CI], 19-40) months for HER2-Zero versus 17.0 (95% CI 14-20,) months for HER2-low; P= 0.0035. In multivariate analysis, HER2-low expression remained significant determinant of inferior PFS after adjusting for other factors including the line of treatment (aHR:2.10, 95% CI 1.03-4.27, P=0.041), age (aHR 2.20, 95% CI 1.29-3.77, P=0.004)), number of metastasis (aHR:1.96, 95% CI 1.32-2.91, P=0.001), and site of metastasis (aHR:1.64, 95%CI 1.058-2.55, P=0.027). Conclusion: In patients with advanced-stage breast cancer treated with CDK4/6 inhibitors and ET, level of HER2 negativity may affect treatment outcomes; patients with HER2-Zero had better PFS compared to those with HER2-low disease. These findings, if confirmed in larger studies, should help oncologists select patients with HER2-low for better treatment options including a combination of anti-HER2 therapy and CDK4/6 inhibitors. Citation Format: Baha’ Sharaf, Hala Abu-Fares, Faris Tamimi, Suhaib Al-Sawajneh, Osama Salama, Rand Daoud, Abdulrahman A. Alhajahjeh, Sawsan Al-lababidi, Hikmat Abdel-Razeq. Differences in Treatment Outcomes Between Patients with HER2-Low versus HER2-Zero, Hormone Receptor-Positive Advanced-Stage Breast Cancer Treated with CDK4/6 Inhibitors [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P4-07-33.

Abstract OT2-08-02: SOLTI-1907 ATREZZO: Targeting hormonal receptor negative (HR-) or PAM50 non-luminal disease with atezolizumab in combination with trastuzumab and vinorelbine in HER2-positive metastatic breast cancer (MBC)

Abstract Background Today, there is no clear therapeutic algorithm for patients with metastatic HER2-positive (HER2+) breast cancer (BC) who have progressed to trastuzumab, pertuzumab, tyrosine kinase inhibitors and antibody-drug conjugates (ADC). Among the emerging strategies, the use of immune checkpoint inhibitors in combination therapy is showing promising clinical benefit in the advanced setting of HER2+ BC by overcoming immune resistance and enhancing antitumor cellular immunity. The intrinsic subtypes Basal-like and HER2-enriched (i.e PAM50 non-luminal tumors) represent approximately the 60% of HER2+ BC and are associated with higher expression of immune-related genes, tumor-infiltrating lymphocytes (TILs) presence and high tumor mutational burden (TMB), compared to luminal subtypes. Additionally, immune infiltration and TMB in HER2+ BC are associated with chemo/antiHER2 responsiveness and with potential benefit from anti-PD-1/PD-L1 inhibitors. We hypothesize that combining atezolizumab with trastuzumab and vinorelbine may improve outcomes in HR- or PAM50 non-luminal/HR-positive (HR+) disease within HER2+ MBC. Methods ATREZZO is an open-label, single-arm, Simon 2-stage, multicenter phase II study. The trial will include 55 pre- or post-menopausal female or male patients with unresectable locally advanced or metastatic HR- or PAM50 non-luminal/HR+ HER2+ BC and progressed to trastuzumab-based chemotherapy and anti-HER2 ADC. Prior pertuzumab is allowed, but not required. Treatment consists of atezolizumab IV 1200 mg every 3 weeks combined with trastuzumab and vinorelbine. Patients with stable, progressing, or untreated brain metastasis not requiring immediate local therapy are eligible. The primary objective is to evaluate the Overall Response Rate (ORR) according to RECIST v 1.1 and secondary endpoints include ORR in patients with PD-L1 positive breast cancer, clinical benefit rate, overall survival and progression-free survival. The final recruited population will contain no more than 60 % of patients with PD-L1 negative tumors. Tumor assessments will be performed every 9 weeks. Incidence, duration and severity of adverse events, and further correlative molecular analyses will be also evaluated. An interim analysis will be conducted when 19 patients are evaluable for ORR and if the number of responses is ≥ 3, 36 additional patients will be included. As of July 15th, 2022, 48 patients were screened and 15 were included in sixteen Spanish sites. This study was funded by Roche Farma SA. Trial identification: NCT04759248 Citation Format: Eva Ciruelos, Antonia Perelló, Santiago González-Santiago, Ana López, Francisco Javier Salvador Bofill, Cinta Albacar, Juan Miguel Cejalvo, Santiago Escrivá-de-Romani, Isabel Blancas, Sonia Pernas, Olga Martínez-Sáez, Josefina Cruz, Jose Ponce, Sonia Servitja, Maria-Eva Perez-Lopez, Juan A Guerra, Esther Sanfeliu, Cesar A Rodríguez, Guillermo Villacampa, Lorea Villanueva, Pablo Tolosa, Tomás Pascual, Aleix Prat. SOLTI-1907 ATREZZO: Targeting hormonal receptor negative (HR-) or PAM50 non-luminal disease with atezolizumab in combination with trastuzumab and vinorelbine in HER2-positive metastatic breast cancer (MBC) [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr OT2-08-02.

Abstract P3-05-23: Pathological complete response rate and disease-free survival after neoadjuvant chemotherapy in patients with HER2-low and HER2-0 breast cancers

Abstract ABSTRACT Background: Half of HER2-negative breast cancers (BC) show HER2-low expression. The strong efficacy of recent anti-HER2 antibody-drug conjugates (ADC) in HER2-low tumors has risen the interest of HER2-low as a proper BC subtype. Chemosensitivity and prognosis of this subtype are not clear when compared to HER2-0 tumors. We investigated the pathological complete response (pCR) and disease-free survival (DFS) rates in BC patients receiving neoadjuvant chemotherapy for HER2-low or HER2-0 tumors. Methods: Data were collected from the Institut Paoli-Calmettes European Comprehensive Cancer Center database. HER2-low tumors were defined by HER2 IHC score of 1+ or 2+ with negative FISH, and HER2-0 by IHC score of 0+. Clinicopathological characteristics, pCR (defined as [ypT0/ypTis] and [pN0sn or ypN0]) and DFS rates were compared between the two cohorts. Results: From Jan/2005 to Jun/2021, 1,111 patients receiving neoadjuvant chemotherapy were evaluable. The incidence of HER2-low was 41%, including 63% of hormone receptor (HR)-positive and 37% of HR-negative tumors (p< 0.001). In the whole population, the pCR rate was lower in HER2-low (23%) versus HER2-0 (30%) tumors (p=0.013), but this association was lost in multivariate analysis. In HR-positive patients, HER2-low negatively impacted pCR rates when compared to HER2-0 (10% vs. 16%, p=0.046), but not in HR-negatives (46% vs. 42%), and this result was maintained in multivariate analysis (OR 0.60 [95CI 0.36-1.00]; p=0.049, and OR 1.15 [95CI 0.77-1.71]; p=0.490, respectively. No correlation existed between DFS and HER2-status. Conclusion: HER2-low is associated with HR positivity. HER2 status did not impact pCR in HR-negative patients, whereas HER2-low was associated with lower pCR rate in HR-positive patients, supporting the development of anti-HER2 ADC in this setting. Citation Format: Alexandre de Nonneville, Gilles HOUVENAEGHEL, Monique Cohen, Laura SABIANI, Marie BANNIER, Frederic VIRET, Anthony Gonçalves, François BERTUCCI. Pathological complete response rate and disease-free survival after neoadjuvant chemotherapy in patients with HER2-low and HER2-0 breast cancers [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P3-05-23.

Abstract HER2-09: HER2-09 Multiomics Profiling Characterizes Distinct HER2-low Breast Cancer Subgroups in the East Asian Population

Abstract Background: The emergence of anti-HER2 antibody–drug conjugates (ADCs) gave rise to the concept of HER2-low breast cancer (BC). HER2-low BC, which refers to a subgroup of HER2-negative BC with relatively higher HER2 expression (defined as 1+ or 2+ by immunohistochemistry (IHC) staining, without ERBB2 amplification), represents a rather large part of all BCs. However, the molecular nature and internal heterogeneity of HER2-low breast cancer remain obscure, and little is known about the ethnic differences of HER2-low BC. These limitations prevent us from a more precise patient selection and better drug combination strategies in the era of ADCs. To provide a comprehensive and intensive landscape of HER2-low BCs, we characterized HER2-low BCs both clinically and molecularly, which may help clinicians to achieve a more precise clinical management of these patients. Patients and methods: We established a HER2-low BC cohort (N=441) in early-stage Chinese patients and included HER2-0 (N=114) and HER2-positive (N=181) tumors as auxiliary cohorts to characterize HER2-low breast cancers both clinically and molecularly. Whole-exome sequencing, copy number variation assays, RNA sequencing and isobaric quantitative proteomics were conducted to obtain multiomics data. We compared the clinicopathological and molecular features between HER2-low tumors and other HER2 status subgroups stratified and not stratified hormone receptor (HR) status to clarify the distinctness of HER2-low BCs. And we analyzed the internal heterogeneity and ethnic difference of HER2-low BCs by characterizing a distinct subgroup of patients with unique driving mechanisms. Results: HER2-low BCs showed different molecular manifestations from HER2-0 BCs in different HR subgroups. In the HR-negative subgroup, HER2-low BCs consisted of more non-basal-like subtypes than HER2-0 tumors (40.0% vs. 9.1%, P = 0.002), which was an East Asian-specific phenomenon absent in Western cohorts. Also, HR-negative HER2-low BCs showed significant internal molecular heterogeneity, of which basal-like tumors closely mimicked HER2-0 BCs, whereas non-basal-like tumors were similar to HER2-positive BCs. These non-basal-like tumors were mostly categorized as HER2-enriched and luminal androgen receptor (LAR) subtypes. These molecularly distinct tumors might be driven by frequent mutation in PIK3CA and overexpression of FGFR4 and PTK6, which may also serve as therapeutic targets. These results have also been proved in a triple negative breast cancer cohort we reported previously. In contrast, in the HR-positive subgroup, HER2-low BCs showed no large-scale molecular difference from HER2-0 BCs or internal heterogeneity. However, HER2-low patients showed significantly better distant metastasis-free survival than HER2-0 patients (P = 0.029), which might be attributed to the lower loss/deletion levels of 17q11.12 and 17q21.31 in HER2-low breast cancers, in which genes including NF1 and BRCA1 are located. Conclusions: We reported the largest single-center multiomics HER2-low BC cohort in East Asian hitherto, and revealed its molecular nature, internal heterogeneity and ethnic difference. Compared with HR-positive diseases, HER2-low BCs in the HR-negative subgroup were more likely to be a molecularly distinct entity from HER2-0 tumors. Furthermore, HR-negative HER2-low BC also accommodates higher internal heterogeneity, which was ethnicity-specific in our East Asian cohort and may infer a different treatment response. Our work emphasized the need of a more precise stratification within HER2-low BCs and across ethnic groups, which has also been inferred by the results in the subgroup analysis of DESTINY-Breast04 trial. Citation Format: Lei-Jie Dai, Ding Ma, Yi Xiao, Xi Jin, Song-Yang Wu, Ya-Xin Zhao, Han Wang, Wen-Tao Yang, Yi-Zhou Jiang, Zhi-Ming Shao. HER2-09 Multiomics Profiling Characterizes Distinct HER2-low Breast Cancer Subgroups in the East Asian Population [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr HER2-09.

Resistance to Antibody-Drug Conjugates Targeting HER2 in Breast Cancer: Molecular Landscape and Future Challenges.

The treatment of HER2-positive metastatic breast cancer (mBC) with Trastuzumab emtansine (T-DM1) and Trastuzumab deruxtecan (T-DXd), two antibody-drug conjugates (ADCs) targeting HER2, is burdened by progression of disease related to the acquisition of mechanisms of resistance. Resistance to T-DM1 is caused by the decrease of HER2 expression, the alteration of intracellular trafficking, the impairment of lysosome functions, the drug expulsion through efflux pumps and the activation of alternative signal pathways. Instead, the decrease of HER2 expression and SLX4 loss of function mutations represent the first evidences of mechanisms of resistance to T-DXd, according to the results of DAISY trial. Several strategies are under evaluation to overcome resistances to anti-HER2 ADCs and improve clinical outcomes in patients progressing on these agents: combinations with tyrosine kinase inhibitors, statins, immune checkpoint inhibitors and synthetic DNA-damaging agents are emerging as promising approaches. Furthermore, novel anti-HER2 ADCs with innovative structures and mechanisms of action are in development, in the attempt to further improve the activity and tolerability of currently available agents.

Comparative analysis of the molecular profile and tumor immune microenvironment (TIME) of human epidermal growth factor receptor 2 (HER2) low (L)- versus high (H)-expressing gastroesophageal cancers (GEC).

287 Background: Addition of immune checkpoint blockade to anti-HER2 therapy has improved outcomes in HER2-positive GEC. Anti-HER2 antibody-drug conjugates have shown activity in some HER2-L tumors in other tumor types. We aimed to compare the molecular profile and TIME of HER2-L and HER-H GEC. Methods: 8678 GEC (gastric, GE junction, and esophageal) adenocarcinoma and squamous cell carcinoma samples were analyzed by next-generation sequencing (NGS) of RNA (whole transcriptome, NovaSeq), DNA (592 genes, NextSeq, or whole exome sequencing, NovaSeq), and immunohistochemistry (IHC, Caris Life Sciences, Phoenix, AZ). Cohorts were stratified by IHC HER2 values of 0 (non-expressors), 1-2+ (HER2-L), or 3+ (HER2-H) and compared using X2 or Fisher-Exact. Statistical significance was determined as P-value adjusted for multiple comparisons (q < 0.05). Microenvironment cell population (MCP) counter was used to quantify immune cell infiltration. Results: Tumor were grouped into HER2 non-expressors (N = 5217), L (N = 2660), and H (N = 801). Mutations of TP53 (72% vs 92%) and amplification of MYC (4% vs 7%), CCNE1 (7% vs 12%), CCND3 (2% vs 4%), CDK6 (3% vs 6%), SMARCE1 (2% vs 25%) and RARA (3% vs 24%) were significantly lower in HER2-L compared to HER2-H (q < 0.05). ARID1A (14% vs 9%), PIK3CA (8% vs 3%), KRAS (9% vs 2%), GNAS (2% vs 0.3%), KMT2D (6% vs 1%), CDH1 (5% vs 1%), and ATM (4% vs 1%) mutations were significantly higher in HER2-L compared to HER2-H (q < 0.05). HER2-L was associated with more TMB-H (9.3% vs. 5%; q<0.05), MSI-H (4.7% vs. 0.6%; q<0.05), and a trend towards higher PD-L1 expression than in HER2-H (80.8% vs 76.1%, p < 0.05 but q > 0.05). Immuno-oncology (IO)-related gene expression inversely correlated with HER2 expression with lowest expression of PDCD1LG2, CD274, CTLA4, PDCD1, HAVCR2, CD80, IFNG, LAG3, and CD86 in HER2-H (q < 0.05). HER2-L had significantly higher median immune infiltration of B cells (fold change [FC]: 1.22), T cells (FC: 1.16), CD8+ T cells (FC: 1.56), NK cells (FC: 1.12), neutrophils (FC: 1.10), cytotoxic lymphocytes (FC: 1.36), and myeloid dendritic cells (FC: 1.34), compared to HER2-H (q < 0.05). HER2 non-expressors showed similar immune cell infiltrates compared to HER2-H. HER2-H was associated with lower T-cell inflamed scores and IFN gamma signature when compared to HER2-L and non-expressors (q < 0.05). Conclusions: To our knowledge, this study is the first and largest comparison of molecular profile and TIME in HER2-expressing GEC. We demonstrated distinct molecular and TIME profiles with higher immunogenic profiles in HER-L as compared to HER2-H. IO-related gene expression and TIME cell distribution differences in HER2-H GEC suggest that response to IO and HER2 therapy combinations is likely related to HER2-targeted treatment effect on TIME rather than baseline immunogenicity of the tumor.

Pharmacovigilance Analysis of HeartFailure Associated With Anti-HER2 Monotherapies and Combination Regimens for Cancer.

Trastuzumab improves outcomes in patients with HER2-overexpressing malignancies but is associated with decreases in left ventricular ejection fraction. Heart failure (HF) risks from other anti-HER2 therapies are less clear. Using World Health Organization pharmacovigilance data, the authors compared HF odds across anti-HER2 regimens. In VigiBase, 41,976 patients had adverse drug reactions (ADRs) with anti-HER2 monoclonal antibodies (trastuzumab, n=16,900; pertuzumab, n=1,856), antibody-drug conjugates (trastuzumab emtansine [T-DM1], n=3,983; trastuzumab deruxtecan, n=947), and tyrosine kinase inhibitors (afatinib, n=10,424; lapatinib, n=5,704; neratinib, n=1,507; tucatinib, n=655); additionally, 36,052 patients had ADRs with anti-HER2-based combination regimens. Most patients had breast cancer (monotherapies, n=17,281; combinations, n=24,095). Outcomes included comparison of HF odds with each monotherapy relative to trastuzumab, within each therapeutic class, and among combination regimens. Of 16,900 patients with trastuzumab-associated ADRs, 2,034 (12.04%) had HF reports (median time to onset 5.67months; IQR: 2.85-9.32months) compared with 1% to 2% with antibody-drug conjugates. Trastuzumab had higher odds of HF reporting relative to other anti-HER2 therapies collectively in the overall cohort (reporting OR [ROR]: 17.37; 99% CI: 14.30-21.10) and breast cancer subgroup (ROR: 17.10; 99% CI: 13.12-22.27). Pertuzumab/T-DM1 had 3.4 times higher odds of HF reporting than T-DM1 monotherapy; tucatinib/trastuzumab/capecitabine had similar odds as tucatinib. Among metastatic breast cancer regimens, HF odds were highest with trastuzumab/pertuzumab/docetaxel (ROR: 1.42; 99% CI: 1.17-1.72) and lowest with lapatinib/capecitabine (ROR: 0.09; 99% CI: 0.04-0.23). Trastuzumab and pertuzumab/T-DM1 had higher odds of HF reporting than other anti-HER2 therapies. These data provide large-scale, real-world insight into which HER2-targeted regimens would benefit from left ventricular ejection fraction monitoring.

A single-arm, open-label, phase II study investigating anti-HER2 ADC plus anti-PD-1 antibody in patients with unresectable locally advanced or metastatic BTC with HER2 overexpression.

TPS619 Background: 1. Biliary tract cancer (BTC) accounts for 4% of all digestive tract cancers. In 2016-2020, the incidence of BTC has gradually increased worldwide. In China, gallbladder carcinoma accounted for 1.7% of all tumors and 1.9% of deaths in some regions. 2. The standard second-line treatment regimen is mFOLFOX, with an ORR of 5%, PFS of 2 months and OS of 6.2 months. Methods: This is a single-arm, open-label, phase II study, approximately 31 patients of unresectable locally advanced or metastatic BTC with HER2 overexpression will be enrolled. Each treatment cycle is 14 days. Therapeutic regimen: Disitamab Vedotin 2.5mg/kg IV, D1, Q2W + Zimberelimab 240mg IV, D1, Q2W All patients treated until a loss of clinical benefit or unacceptable toxicity is observed by the investigator. Objective: to evaluate the efficacy and safety of Disitamab Vedotin + Zimberelima in patients with unresectable locally advanced or metastatic BTC with HER2 overexpression Disitamab Vedotin: Vedecitumab is an antibody-drug conjugate (ADC), with a humanized antibody conjugated to four tubulin inhibitor molecules (DAR=4) Zimberelimab: Zimberelimab is a fully human IgG4 monoclonal antibody that specifically binds to PD-1 molecules on the surface of lymphocytes. This drug is approved for r/r cHL and is also effective in the cervical cancer. Clinical trial information: NCT05540483 .

Tetrapeptide-based mimotope affinity monolith for the enrichment and analysis of anti-HER2 antibody and antibody-drug conjugate

Selective enrichment and analysis of therapeutic antibodies in biological fluids are crucial for the development of biopharmaceuticals. Recently, peptide-based affinity chromatography has exhibited fascinating prospects for antibody enrichment due to the high affinity and specificity of small peptides. However, the post-modification approach of peptide ligands on the material surface is complicated and time-consuming. In this study, a methacrylate modified tetrapeptide (m-EDPW) was firstly demonstrated as the affinity ligand of trastuzumab (Kd = 1.91 ± 1.81 μM). Next, the m-EDPW based affinity monolith was prepared using a facile one-step polymerization method, which could overcome the drawbacks of traditional post-modification preparation strategies. Based on the monolith as described above, a simple enrichment approach was developed under the optimal washing and elution conditions. Based on the excellent properties, such as high porosity (53.09%), weak electrostatic interaction and suitable affinity (1.00 ± 2.14 μM for anti-HER2 ADC), this novel monolith exhibited good specificity and recovery for antibodies (91.6% for trastuzumab, 98.37% for anti-HER2 ADC), and low nonspecific adsorption for human serum albumin (DBC10% = 0.5 mg/g polymer). Particularly, this material was successfully applied to enrich trastuzumab and its related antibody-drug conjugate (ADC) from different cell culture medias. The dynamic tracking analysis of ADC in the critical quality attributes (e.g., charge variants, drug to antibody ratio and subunit conjugation ratio) was also achieved by combining the enrichment approach, capillary electrophoresis or reversed phase liquid chromatography. In summary, the exploited peptide-based mimotope affinity materials showed a great potential for the application in biopharmaceutical analysis.