TEST:

HercepTest

The advent of novel therapeutic molecules that require fewer membrane epitopes to be effective has prompted a reevaluation of the current immunohistochemistry testing protocols, with special emphasis on the detection limit. Here, we have used Boston Cell Standards technology to determine the sensitivity of 2 commercially available HER2 immunohistochemistry assays, including a lower limit of detection.

Our cohort included 102 cases (64% male, 36% female); 71% were GC and 30,29% GEJ cancers. Of these, 94 patients had both determinations, while 98 only had dMMR and another 98 only HER2. Global HER2 positivity was 13,4% (13/98), while dMMR was 11,22% (11/98).

According to our study, in Southwest China, one-third of UTUC patients overexpressed HER2. Tumors with high grade or luminal phenotype tended to be HER2 positive. HER2 may represent a promising target for therapy in UTUC.

Initial BC diagnosis (2019): Localized infiltrating ductal carcinoma (pT2N1M0) with hormone receptor expression (ER 95%, PR 60%) and a negative HercepTest, treated with surgery, chemotherapy (docetaxel and anthracyclines), radiotherapy, and tamoxifen. The simultaneous management of APL and RBC necessitates a multidisciplinary approach. Despite the complexity and cardiotoxicity from prior anthracycline treatment, the patient achieved complete remission.

Recently, a new type of antibody-drug conjugate, trastuzumab-deruxtecan (T-DXd), has been approved for the treatment of metastatic breast cancer with low level of human epidermal growth factor receptor 2 (HER2) gene expression...The ability of pathologists to achieve acceptable diagnostic accuracy in identifying patients with HER2-low breast cancer could be enhanced by short-term training. Potential routes to improve the quality of HER2-low scoring in clinical practice have been identified.

As assays were validated for detecting HER2-amplified tumours, not all assays and antibodies proved suitable for HER2-low detection. Some tests showed distinct expression in the negative cell line. Dynamic range cell line controls and quantitative analysis using calibrators demonstrated more interlaboratory variability than commonly appreciated. Revalidation of HER2 tests by laboratories is needed to ensure clinical applicable HER2-low assays.

In March 2019, AstraZeneca entered into a global development and commercialization collaboration agreement with Daiichi Sankyo for trastuzumab deruxtecan (T-DXd; DS-8201)... Data show the gastric and breast algorithms are comparable in HER2 IHC 3+ identification; lower concordance was observed for IHC 2+ and 1+, and between the gastric and endometrial algorithms. Findings indicate greater awareness of best scoring practice is needed to ensure consistent assessment of HER2 IHC status in solid tumors. Table: 177P *Salivary gland tumors (n=18) and DP-02 other cohort; †PPA was 100% as no cases identified by gastric/breast algorithm

Local vs central HER2 agreement by standardized IHC techniques decreased for HER2-low compared to HER2 status. Monoclonal HercepTest proved higher agreement with gold standard than polyclonal HercepTest in terms of HER2 status and HER2-low BC identification, especially for 1+ and 2+ IHC cases. This highlights the need for standardized HER2 testing review, particularly when evaluating potential benefits of novel therapies for HER2-low BC patients.

Across the six European labs, there was a minor increase in the percentage of 2+ cases identified in the 1800 cases after implementation of HercepTest™ mAb pharmDx compared to 1800 cases tested with the labs’ previous HER2 assay. However, the overall percentage of IHC 2+ cases remain closely aligned with published data for IHC 2+ positivity range. Overall, there is no change (0.19%) in the number of patients being detected ISH-positive, i.e., no change in patients eligible for targeted treatment.

The HER2 “ultra-low� category is not currently recognized as a target for new HER2 anti-ADCs. However, this could change soon depending on the results of ongoing clinical trials. If this becomes the case, HER2 assays must provide supporting evidence on their capability to accurately differentiate HER2 null from HER2 “ultra-low�.

HER2 scores vary markedly between different clinically approved HER2 antibodies, both in primary tumours and their corresponding metastasis, and across different antibodies. Especially DG44 deviated from the other antibodies. Discrepancies in testing methods could impact patient selection for Trastuzumab-deruxtecan treatment.

With the advent of novel ADCs where the amount of HER2 receptors required for a clinical response is lower, the significance of HER2 epitope detection levels at the low end of the expression range is increasingly relevant. This study demonstrates a significant difference in the lower limit of HER2 detection between the two tested assays, which holds potential relevance when considering low levels of HER2. D0113716.

The results showed that the amount of mRNA in Oncotype DX is not useful as an auxiliary diagnostic result for distinguishing between HER2 low (score 1) and HER2 null (score 0) using Dako Herceptest II. In the future, we would like to conduct similar studies with HER2 (4B5), a companion diagnostic for HER2 low

In recent years, trastuzumab deruxtecan has been applied to breast cancer with low HER2 expression, and the evaluation criteria for HER2 protein expression have also changed...In addition, some cases with a HER2 score of 0 were scored 2 or higher by RNA-scope, and a significant positive correlation was observed between ERBB2 expression and NRG1 expression when HER2 score 0 cases were included. In the future, we plan to use more cases to clarify the HER2 dependency of ERBB2 expression-positive cases in RNA-scope and the significance of the expression of HER2-related factors in HER2-low-expressing breast cancer

Background With the advent of trastuzumab deruxtecan (T-DXd), a new concept of HER2-low and HER2-zero has been proposed for HER2-negative breast cancer... Thirty patients (46.2%) were in the HER2-low group and 35 patients (53.8%) were in the HER2-zero group. In terms of clinicopathological factors, the HER2-zero group had significantly more bone metastases than the HER2-low group (p=0.041). Ki-67 was also significantly higher in the HER2-low group than in the HER2-zero group (p=0.002).

P2; Trial completion date: Apr 2025 --> Apr 2028 | Trial primary completion date: Apr 2024 --> Apr 2026

By using USC-specific IHC scoring criteria, both HercepTest and 4B5 assays showed high specificities (100%) for HER2 gene amplification in IHC 3+ cases, high IHC/FISH concordance, and comparable sensitivity for detecting HER2 gene amplification. The notable false negative rates using IHC 2+ as a cut-off for reflexing FISH analysis may warrant consideration for performing FISH in IHC 1+ cases until more data become available.

However, only the ADC tisotumab vedotin (TV) is approved by the Food and Drug Administration (FDA) for use in cervical cancer... TROP2, TF and NECTIN4 are highly expressed in cervical cancer. Clinical trials evaluating the safety and efficacy of ADCs are highly relevant in cervical cancer.

We demonstrated a concordant analytical performance between the tested CLDN18/CLDN18.2 IHC assays in GC. Their implementation in the clinical setting might help identify patient candidates that could benefit from treatment with CLDN18.2 targeted therapies. Additional research is needed to confirm similar analytical performance between the three IHC assays in other indications.

Our findings support the adverse prognostic impact on HER2 in PCa. We propose the use of a modified scoring system to evaluate HER2 expression in PCa. The observed high prevalence of HER2 expression supports the consideration of novel HER2-targeted therapies addressing even low levels of HER2 expression in future PCa trials.

In the BAT, no basophil activation has been triggered by trastuzumab and pertuzumab to-date. Conclusion HER2-targeting IgE antibodies may offer a promising avenue for treatment of HER2-positive tumours, including those with low expression levels.

P2; Unknown status --> Completed

Background: DESTINY-Breast04 (DB-04) showed that trastuzumab deruxtecan improves survival vs chemotherapy in patients with human epidermal growth factor receptor 2 (HER2)-low (immunohistochemistry [IHC] 1+ or IHC 2+ with in situ hybridization negative) metastatic breast cancer (BC)... The degree of concordance between the CTA and non-CTAs varied among assay types and laboratory locations. A low NPA for some non-CTAs suggests the need for further assay optimization for HER2 IHC 0 evaluation.

P=N/A; Enrolling by invitation --> Completed

Background: Anti-HER2 antibody-drug conjugate trastuzumab-deruxtecan has shown efficacy in patients with HER2-low metastatic breast cancer (BC)... These results are the evidence for the need of newly developed algorithms that are capable of distinguishing HER2-low from HER2-negative accurately.

HER2 cytoplasmic granular staining was present in 29.7% of pure ACs using PATHWAY 4B5 platform. The rate of HER2 cytoplasmic granular staining on HercepTest platform was significantly lower than that of PATHWAY 4B5 and was only appeared in 2+ and 3+ cases, suggesting that the HercepTest platform is more suitable for the HER2 low expression cases. The IHC interpretation of HER2 3+ were highly consistent with FISH status.

There is a considerable overlap between mRNA values when correlated with HER2 IHC values regardless of the IHC scoring system used. Only a small percentage of cases fell in H-score category different from ASCO-CAP category, contributing to minimal difference seen in terms of correlation with mRNA scores suggesting that H-score has no advantage over ASCO-CAP scoring system.

Background: The DESTINY-Breast04(DB-04) trial has shown efficacy of breast cancer (BC) patients with HER2-low in the metastatic setting to Anti-HER2 antibody-drug conjugate trastuzumab-deruxtecan... We conclude that a significant number of patients will receive incorrect Anti-HER2-low therapy by simply taking the reported score on a historic sample. We recommend repeating the assay following the FDA-approved protocol on a large sample size such as excisional biopsy.

The current IHC assays are suboptimal in evaluating HER2-low breast cancers. RNAscope is a simple assay that can be objectively quantified. RNAscope is better than the current IHC assays in evaluating HER2 expression in HER2-low breast cancers and may identify patients who would benefit from T-DXd treatment.

ER and PR staining show a high level of concordance (97.3% and 87.1%, respectively) between IBC and DCIS. HER2 showed a slight trend toward increased staining in DCIS, however, only 4 of the 147 cases (2.7%) were 3+ with an associated HER2 negative IBC. Our findings suggest that ER, PR, and HER2 show concordance between IBC and DCIS, with only a slight increase in HER2 staining in DCIS.

Background: The DESTINY-Breast04(DB-04) trial has shown efficacy of breast cancer (BC) patients with HER2-low in the metastatic setting to Anti-HER2 antibody-drug conjugate trastuzumab-deruxtecan... We conclude that a significant number of patients will receive incorrect Anti-HER2-low therapy by simply taking the reported score on a historic sample. We recommend repeating the assay following the FDA-approved protocol on a large sample size such as excisional biopsy.

P1/2; Trial completion date: Dec 2025 --> Dec 2026 | Trial primary completion date: Jul 2025 --> Oct 2026

The RNA levels were significantly higher (p=0.030) in responders (6.4±8.2 dots/cell, n=12) than in non-responders (2.6±2.2, n=20) to T-DXd. RNAscope is a simple assay that can be objectively quantified and is a promising alternative to current IHC assays in evaluating HER2 expression in breast cancers, especially HER2-low cases, and may identify patients who would benefit from T-DXd.

P1; Not yet recruiting --> Recruiting

This assay is linked to the use of the monoclonal antibody trastuzumab in the treatment of HER2 positive breast cancer...Currently, more than 60 drugs or drug combinations, primarily in hematology and oncology, have been approved by the FDA, with CDx assays linked to their use. The current article briefly discusses the subject of CDx and provides an overview of its evolution over the past 25 years, with particular emphasis on the United States.

In this large single institutional analysis of dual HER-2 testing, only 12% of cases demonstrate HER-2 amplification, and overall in our institutional pathology database, from 2001 – March 2023, 11.2% of cases are HER-2+ by either IHC or ISH. In our dual tested cohort, only 13.6% of IHC 2+ cases demonstrate amplification, and 13.8% are non-amplified, but have excess HER-2 copy number. We demonstrate a numerical increase in the rate of amplification, and copy number in non-amplified cases, associated with IHC score.

T-DXd has been approved by the FDA to treat patients with metastatic HER2-low and -positive breast cancer...There were significant differences of dots/cell comparing 1+, 2+ and 3+ cases. Note: ns not significant (p>0.05), * p<0.05, *** p<0.001, **** p<0.0001

We have reason to believe that the new antibody will reduce the diagnostic time and avoid unnecessary costs. Due to the small study group, further investigation is needed.

The rate of identifiable tumors with HER2 low or HER2 ultra-low expression varies depending on assay parameters that can easily be modified. Irrespective of assay sensitivity, our data identify various tumor entities with significant fractions of "HER2-low" cases that might benefit from therapy with new antibody drug conjugates.

Furthermore, C-erbB-2 and SP3 were used with different antibody concentrations showing inconsistent HER2-expression. Overall, HER2 antibodies need to be re-validated and the use of a standardized reference material (e.g. cell lines) with AI is a promising method to determine HER2-low for clinical use.

P2; N=55 --> 0 | Trial completion date: Oct 2009 --> Jul 2023 | Completed --> Withdrawn | Trial primary completion date: Oct 2009 --> Jul 2023

Conclusions In this study, there was no prognostic difference based on HER2 expression status in HER2-negative early-stage BC, but there was a frequency difference in determining HER2-low and HER2-0 depending on the immunostaining kits. While there are many assays for HER2 evaluation approved in the practice, an appropriate companion diagnosis for HER2-low seems to be critical to select patients who may benefit from newer treatment such as Trastuzumab Deruxtecan.