HER2 Testing Research Articles

Impact of Primary Subtype on Pattern of Metastases and Survival in Young Women with Metastatic Breast Cancer

Metastatic breast cancer is a heterogeneous disease that varies in biology and organ involvement, and can have a more aggressive course in young patients. This study aimed to describe the pattern of distant metastases of young women with stage 4, or metastatic, breast cancer (MBC). A retrospective study was performed examining women diagnosed with MBC at age 50 years or younger and managed at our institution between 2001 and 2018. Data including primary breast cancer receptor status (ER, PR, and HER2) and distant metastases at MBC diagnosis were recorded. Patients were classified into groups based on receptor status. Metastatic pattern of various MBC groups was assessed using Chi-Square test. Kaplan-Meier method was used to analyze overall survival. 485 patients were eligible for this study. Median age at MBC diagnosis was 42 years (range 25 to 50). Median follow-up time for all patients was 7.1 years. Median survival for all patients is shown in table 1. Primary breast cancer in 230 patients (47%) was ER+ &/or PR+ & HER2- (HR+HER2-), in 78 patients (16%) was ER- & PR- & HER2- (triple-negative), in 59 patients (12%) was ER+ & /or PR+ & HER2+ (HR+HER2+), in 51 patients (11%) was HER2+ & ER- & PR- (HER2+HR-), and in 67 patients (14%) was unclassified. Almost all unclassified patients had unknown HER2 status (65 patients) and received their breast cancer diagnosis prior to 2001 when HER2 testing was not universal, but the majority (45 patients) were known to be hormone receptor positive. 102 patients (21%) had MBC at first diagnosis of breast cancer. The incidence of bone metastasis, brain metastasis, and single metastasis are shown in the table 1. HR+HER2-, HR+HER2+, and unclassified patients had a significantly higher incidence of bone metastasis at MBC diagnosis compared with triple negative and HER2+HR- patients (p<0.05). Conversely, triple negative and HER2+HR- patients had a significantly higher incidence of brain metastasis at MBC diagnosis compared with HR+HER2-, HR+HER2+, and unclassified patients (p<0.05). Incidence of single metastasis at MBC diagnosis was similar across all groups (p = 0.861). Of the 75 patients with single metastasis, 50 had a bone metastasis, 9 had a brain metastasis, and 16 had a metastasis to another site. At MBC diagnosis in young women, the incidence of bone and brain metastasis in HR+HER2+ patients may be closer to that of HR+HER2- patients rather than HER2+HR- patients. In our cohort, the incidence of single metastasis did not appear to vary across primary subtypes.Abstract 2095; TableAll Patients (n = 485)HR+HER2- (n = 230)Triple-negative (n = 78)HR+HER2+ (n = 59)HER2+HR- (n = 51)Unclassified (n = 67)P-valueBone metastasis at MBC diagnosis322 (66%)175 (76%)37 (47%)44 (75%)19 (37%)47 (70%)p<0.05Brain metastasis at MBC diagnosis53 (11%)15 (7%)16 (21%)7 (12%)10 (20%)5 (7%)p<0.05Single metastasis at MBC diagnosis75 (15%)35 (15%)15 (19%)9 (15%)6 (12%)10 (15%)P = 0.861Median Survival (years)3.43.51.45.33.93.1p<0.05 Open table in a new tab

Next-generation assessment of human epidermal growth factor receptor 2 gene (ERBB2) amplification status in invasive breast carcinoma: a focus on Group 4 by use of the 2018 American Society of Clinical Oncology/College of American Pathologists HER2 testing guideline.

The American Society of Clinical Oncology/College of American Pathologists (ASCO/CAP) updated the testing guideline in 2018 to address issues arising from uncommon human epidermal growth factor receptor 2 (HER2) fluorescence in-situ hybridisation (FISH) results according to the 2013 guideline. Next-generation sequencing (NGS) may be used to better classify patients. The aim of this study was to assess the ERBB2 amplification status of invasive breast carcinoma with equivocal HER2 immunohistochemistry (IHC) results by using NGS, focusing on Group 4 (HER2/CEP17 ratio of <2.0; average HER2 signals/cell of ≥4.0 and <6.0). We retrospectively reviewed HER2 FISH and NGS data of HER2 IHC-equivocal breast carcinomas at our centre between January 2009 and September 2019, wherein all three assays were performed on the same tissue block, and compared HER2 FISH results, according to the 2018 ASCO/CAP guideline, and the ERBB2 amplification status determined with NGS. A total of 52 HER2 FISH and NGS results from 51 patients with HER2 IHC-equivocal breast carcinomas were reviewed. The cohort included eight cases classified as 2018 ASCO/CAP in-situ hybridisation Group 1, three classified as Group 2, three classified as Group 3, 14 classified as Group 4, and 24 classified as Group 5. Thirteen of 14 (92.9%) Group 4 (HER2-negative) cases were classified as ERBB2-non-amplified by the use of NGS; the discordant case was later classified as Group 1 with alternative sample FISH testing. NGS revealed no significant difference in somatic mutations or copy number alterations between Groups 4 and 5. Our NGS findings support the reclassification of HER2 FISH-equivocal cases as HER2-negative under the 2018 ASCO/CAP guideline.

Adherence to and determinants of guideline-recommended biomarker testing and targeted therapy in patients with gastroesophageal adenocarcinoma: Insights from routine practice.

12 Background: Precision oncology has transformed care for patients with advanced HER2+ gastroesophageal adenocarcinoma (GEA), where the addition of anti-HER2 therapy with trastuzumab improves overall survival and is now incorporated into national guidelines. However, little is known about adherence to and determinants of timely HER2 testing and trastuzumab initiation in routine care. Methods: We performed a retrospective study of advanced GEA diagnosed between 1/2011 and 6/2019 in the nationwide Flatiron Health EHR-derived deidentified database. We calculated annual prevalence and identified determinants of timely HER2 testing (defined within 21 days after advanced diagnosis) and timely first-line trastuzumab initiation (defined within 14 days after a HER2+ result). Log binomial regressions estimated adjusted prevalence ratios (PR) comparing timely HER2 testing and trastuzumab initiation by patient and tumor characteristics. Multiple imputation was conducted for missing data. Results: We included 6,479 patients with advanced GEA; 973 were HER2+ of whom 585 (60.1%) initiated trastuzumab. Prevalence of timely HER2 testing increased from 22.4% in 2011 to 44.5% in 2019; timely trastuzumab initiation remained stable at 18.0% over the same period. No appreciable differences in timely testing or trastuzumab initiation by age, sex, race or payer category were noted. Patients with early-stage GEA who subsequently developed metastatic disease were less likely to undergo timely HER2 testing and trastuzumab initiation than those with metastatic disease at diagnosis (PR 0.69, 95% CI 0.64-0.75 for testing; PR 0.44, 95% CI 0.28-0.69 for therapy). Similar findings were seen for patients who did not receive surgery (PR 0.56, 95% CI 0.47-0.65 for testing; PR 0.57, 95% CI 0.22-1.46 for therapy). Patients who started chemotherapy before learning their HER2 status (PR 1.42, 95% CI 1.05-1.90) and those with concordant HER2 immunohistochemistry and FISH results (PR 1.76, 95% CI 0.99-3.13) were more likely to initiate timely trastuzumab. Conclusions: Among patients with advanced GEA, guideline-recommended HER2 testing and anti-HER2 therapy initiation remain underutilized. Misclassification due to missing testing or treatment performed outside the Flatiron Network is a study limitation. Uptake of precision oncology may improve with implementation of universal HER2 testing of GEA patients regardless of stage and reflex multidisciplinary review of discordant HER2 test results.

ISH-basierte HER2-Diagnostik

Aprerequisite for all HER2 directed therapies is the demonstration of HER2 receptor protein overexpression and/or gene amplification by in situ hybridization (ISH). ASCO and CAP have published several HER2 test guidelines over the past 15years for both breast and gastric cancer. The latest version for breast cancer (2018) focuses on special issues of ISH related to the definitions of special diagnostic groups (1-5). The guidelines for gastroesophageal adenocarcinoma (2017), essentially based on ToGA trial data, are now also being used for other tumors such as pancreas, gallbladder, and non-small-cell lung cancer. For colorectal cancer, amodified testing procedure has been proposed. Recently, besides overexpression and amplification, athird type of HER gene alteration, namely mutation, has gained much interest. Next-generation sequencing (NGS) allows detection of both amplification and mutation of the HER2 gene providing new options of therapy especially in the case of activating mutations.

Biomarker heterogeneity between primary breast cancer and synchronous axillary lymph node metastases.

Whether the expression status of estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER-2) receptor and Ki-67 show concordance between the primary tumors and the synchronous axillary lymph node (ALN) metastases has been discussed in numerous studies. However, to date, the results of these studies remain controversial. Therefore, the present study aimed to investigate whether the expression of ER, PR, HER-2 and Ki-67 was in concordance between the primary tumors and synchronous ALN metastases in patients with operable breast cancer (BC). A total of 60 tissue samples were collected from patients with primary operable BC diagnosed with primary tumors and synchronous ALN metastases. The expression levels of the four biomarkers, ER, PR, HER-2 and Ki-67, were assessed in primary lesions and synchronous ALN metastases samples using immunohistochemistry. The cut-off values were set to 10% for ER and PR, while the labeling index of Ki-67 was set to 14%. The immunostaining intensity of ER and PR was scored as negative (−), 1+, 2+ and 3+. The criteria for HER-2 testing in BC were implemented according to the American Society of Clinical Oncology (ASCO) and the College of American Pathologists (CAP) guidelines. The concordance rates for ER, PR and HER-2 were 96.7 (58/60), 96.7 (58/60) and 90% (54/60), respectively. In addition, the kappa values of consistency in the primary lesions and the synchronous ALN metastases were 0.773 for ER, 0.654 for PR and 0.785 for HER-2. Furthermore, the P-values of ER, PR and Ki-67 numerical variables between the two groups were 0.393, 0.400 and 0.331, respectively, as demonstrated using a non-parametric Wilcoxon signed rank test. The findings of the present study demonstrated a high degree of concordance between the expression of ER, PR, HER-2 and Ki-67 in the primary tumors and that in the synchronous ALN metastases, suggesting that the BC primary tumor biomarkers may be used for the prognosis of synchronous ALN metastases in patients with operable BC.

Details of human epidermal growth factor receptor 2 status in 454 cases of biliary tract cancer

Human epidermal growth factor receptor 2 (HER2)-targeted therapy has improved clinical outcomes in patients with HER2-positive breast and gastric cancers, although ineffective or recurrent cases are present. One reason for this is the heterogeneity of HER2 expression in cancer cells. The aim of this study was to investigate the clinicopathological characteristics and HER2 status of patients with biliary tract cancers (BTCs). We examined HER2 protein expression by immunohistochemistry, HER2 gene amplification by fluorescence in situ hybridization, and both HER2 protein and gene levels simultaneously by gene-protein assay. Samples were collected from 454 patients who underwent surgical resection for BTCs (110 intrahepatic cholangiocarcinomas [ICC], 67 perihilar extrahepatic cholangiocarcinomas [ECC-Bp], 119 distal extrahepatic cholangiocarcinomas [ECC-Bd], 80 gallbladder carcinomas [GBC], and 79 ampullary carcinomas [AVC]). HER2 status was assessed according to the guidelines for HER2 testing in gastroesophageal adenocarcinoma. HER2-positive status was detected in 14.5% of BTCs (3.7% of ICC, 3.0% of ECC-Bp, 18.5% of ECC-Bd, 31.3% of GBC, and 16.4% of AVC). Furthermore, HER2-positivity tended to correlate with low histological grade, tumor histology, and macroscopic features in certain tumors. HER2 heterogeneity was common and highly frequent (83%) in BTC cases. Reduced HER2 protein expression in the deeper invasive areas with simultaneous dedifferentiation was frequently observed in HER2-positive cancer cells. The findings of this study suggest that a large subgroup of HER2-positive BTC cases can be considered for HER2-targeted therapy. Moreover, the HER2 status in BTCs should be determined carefully using a sensitive approach toward larger cancer tissues.

133P HER2 transcription status of diverse solid tumours with clinical implications

Utility of HER2 mRNA expression as a potential therapeutic biomarker is unclear. We previously reported the discordant HER2 status between copy number variation (CNV), mRNA and immunohistochemistry (IHC) from a large multi-institute dataset. Herein, we investigated comprehensive HER2 status by cancer (CA) type with clinical outcome among HER2 positive patients (pts) at University of California San Diego (UCSD). By Paradigm Diagnostics (CLIA-certified laboratory), HER2 status was evaluated using IHC, qRT-PCR and NGS between 2015-2019. We evaluated pts who had all three HER2 testing available by tumor type, with UCSD outcomes included. Of 5,305 pts, 723 had CNV, mRNA and IHC evaluated (Table). Among diverse solid tumors, HER2 positivity in at least one test were seen in 76.6% (49/64) of NSCLC and 58.8% (20/34) of pancreas CA. Sole mRNA positivity (CNV-/IHC-) were found ≥35% in NSCLC, pancreatic and colorectal cancers. Among 53 UCSD pts who had all three HER2 testing, 22 pts had ≥ 1 positive HER2 test (including 24.5% [13/53] pts with mRNA+ only). We identified 7 pts that were given Anti-HER2 therapy (Tx). One pt with CNV+/mRNA+/IHC+ metastatic esophageal CA achieved complete response with chemotherapy plus trastuzumab followed by trastuzumab maintenance (progression-free survival [PFS]: 48 + months). One pt with metastatic cholangiocarcinoma mRNA+ (CNV and IHC unable to perform) achieved durable partial response with dual-anti-Her2 Tx (trastuzumab and lapatinib) (PFS: 24+ months). In contrast, 5 pts had poor outcomes despite anti-HER2 Tx, given after 3-5 previous Tx lines.Table 133PHER2 status per cancer typeCancer Type (N>10)TotalCNV + mRNA + IHC +CNV+ mRNA + IHC -CNV+ mRNA- IHC+CNV- mRNA+ IHC+CNV+ mRNA- IHC-CNV- mRNA+ IHC-CNV- mRNA- IHC+Any +NN%N%N%N%N%N%N%N%Multi-institutionBreast2603513.5––124.6–4416.983.19938.1Colon8844.5––22.3–3236.422.34045.5NSCLC6411.6––1625–3046.923.14976.6Pancreas34–––38.8–1647.112.92058.8Ovary3013.3––26.7–62013.31033.3Sarcoma27–CUP2528––141414416936Prostate21–––––14.814.829.5Hepatobiliary20210––––1515420Bladder19421.1––421.1–15.315.31052.6Gastroesophageal16212.5–16.3––16.316.3531.3H&N14–Rectal14214.317.1–––535.7–857.1Appendix11–UCSD5323.8––47.5–1324.535.72241.5 Open table in a new tab We found diverse HER2 expression patterns among solid tumors, including difficult to treat cancer of unknown primary, pancreatic CA, and hepatobiliary CA. Although it is a small cohort, patients with HER2 mRNA overexpression could respond to anti-HER2 regimen. Further clinical investigation is warranted.

Impact of 2018 ASCO/CAP guidelines on HER-2 reporting categories of IHC and reflex FISH in breast cancer.

Human epidermal growth factor receptor 2 (HER-2) is an established prognostic and predictive biomarker for breast cancer. To ensure accuracy and uniformity for HER-2 testing, ASCO/CAP published guidelines in 2007 which were updated in 2013 and recently in 2018. In this first study from Indian Oncology center, we evaluated the impact of 2018 ASCO/CAP guidelines. We found a substantial decrease in equivocal IHC cases (P-value<.00001). On reclassification, a total of 5.6% cases from equivocal and positive categories (2013 guidelines) shifted to the negative FISH result category (P-value<.0001), with adoption of 2018 guidelines and eliminated the double equivocal cases.

Abstract 5128: Quantitative mass spectrometry of HER2 protein levels reveals high variability within HER2 IHC grades

Abstract About 15% of breast cancers are HER2 over-expressing (HER2 IHC 3+ or IHC 2+/ISH+)), but another 45% have low levels of HER2 (HER2 IHC 2+/ISH- or IHC 1+), and these patients are not currently approved for treatment with trastuzumab. Recently, a new HER2 ADC, DS-8201 showed anti-tumor activity, not only in patients with HER2 over-expressing breast cancer but also in HER2 low expressing tumors in whom to date, there are no effective anti-HER2 therapies indicated. FDA-approved HER2 in vitro diagnostic tests have recognized several limitations including effects of pre-analytical variable (fixation affects antibody sensitivity), limited dynamic range of chromogen-based IHC, and subjectivity in interpretation of the HER2 score. Additionally, the cut-off values (percentage of cells to be positive) defining HER2 positive have been changing over time. Therefore, more accurate, sensitive, precise and objective assays to better identify patients who may benefit from anti-Her2 treatment therapies (e.g DS-8201) are needed. To address this gap, we evaluated upcoming technologies targeted MS and QRT-PCR and aim to compare expression with current diagnostic HER2 tests in FFPE samples Using selected reaction monitoring mass spectrometry (SRM-MS), we quantified proteins from formalin-fixed, paraffin-embedded tissue samples that had been classified as HER2 0, 1+, 2+ or 3+ by IHC (n=107). HER2 protein concentration measured by SRM-MS was compared between patients in different HER2 IHC classifications using an ANOVA, adjusting for multiple comparisons. HER2 concentration (measured by SRM-MS) was progressively increased according to HER2 IHC grouping (i.e. lowest concentration in HER2 0 samples, highest in HER2 3+ samples). HER2 levels were significantly elevated in 2+ vs. 0 (2.2-fold increase, p &amp;lt; 0.05) samples, and trended higher in 2+ vs. 1+ (1.6-fold increase, p = 0.07) and in 1+ vs. 0 (1.4-fold increase, p = 0.17) samples. About 73% of samples scored as IHC0 had detectable Her2 by SRM-MS (from 168 to 623 amol/µg). Among HER2 IHC 0 samples, ~15% (7/47) had HER2 concentrations above the median levels for the 1+ group. Similarly, 19% (3/16) 1+ samples had HER2 levels above the median for the 2+ group. About 20% of samples co-expressed either ERBB1 and/or ERBB3. Simultaneously from FFPE sections we quantified protein level of payload response and resistance markers (MDR, MRP1, Topo1 and SLFN11). We used an objective multiplex non-antibody-based method to quantify multiple targets from FFPE tissue. SRM-MS revealed a range of HER2 expression over 100 orders of magnitude and identify markers of payload response or resistance in the same assay. The differences seen in payload markers expression could affect therapeutic efficacy and may suggest differing responses to Her2-targeted ADC, depending on tumor biology. Multiplexed quantitative MS could be used to accurately predict which patients will derive the most benefit from Her2-ADC therapy based on the specific biology of their tumor. These studies are ongoing. Citation Format: Fabiola Cecchi, Mark Gustavson, Danielle Carroll, Sriram Sridhar, Steven Coats, Anuja Bhalkikar, Sheeno Thyparambil, Wei-Li Liao, Todd Hembrough. Quantitative mass spectrometry of HER2 protein levels reveals high variability within HER2 IHC grades [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5128.

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

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

Breast Cancer With a HER2 IHC2+ and FISH HER2/CEP17 Ratio ≥2.0 and an Average HER2 Gene Copy Number

Purpose: For breast cancer, accurately illustrating HER2 characteristics is a critical precondition for evaluating the prognosis and predicting the efficacy of anti-HER2 therapy. Our purpose was to expose HER2 mRNA expression through an in situ hybridization assay (RNAscope), to aid the identification of HER2 status in breast cancers with a previously controversial classification for patients suffering from a HER2 IHC2+ and HER2/CEP17 ≥2.0 and a <4.0 mean HER2 gene copy number/cell (entitled FISH group 2 by update 2018 HER2 testing guideline).Methods: A total of 8,983 cases of breast cancer with a known HER2 status detected by initial IHC, and a necessary reflex FISH assay for those with IHC2+, were retrospectively analyzed. Then, 41 cases of HER2 IHC2+ in the FISH group 2 were collected and a RNAscope was performed.Results: The incidence of breast cancers with IHC2+ and in the FISH group 2 was 0.46% (41/8,983) in our single-institutional study cohort. In most of the cases (27/41, 65.9%), low levels of HER2 mRNA expression (score 1 and 2 by RNAscope) were demonstrated. Only one case (1/41, 2.4%) of high-level HER2 mRNA expression (score 4 by RNAscope), harboring a FISH HER2/CEP17 ratio of 2.06 and an average HER2 copy number of 3.70, was revealed. One case with the highest FISH HER2/CEP17 ratio of 3.90, showed the lowest level of HER2 mRNA expression (score 1 by RNAscope). Two cases with the same highest average HER2 signals/cell (3.95) by FISH possessed score 3 and score 2 with RNAscope, respectively. No cases with a score of 0 by RNAscope occurred in our sample. In the majority of cases (35/41, 85.4%), hypodisomy of chromosome 17 (average CEP17 signals/cell ≤1.75) was observed. There was no significant relationship between the mRNA expression and FISH results (average HER2 signals/cell, average CEP17 copy number, or HER2/CEP17 ratio) and clinicopathological features (ER and PR statuses, Ki 67 index, tumor size, and lymph node metastasis) in our population.Conclusion: HER2 mRNA overexpression was not a feature in our group of patients. Based on our data, breast cancers with HER2 IHC2+ and in FISH group 2 support a categorization of HER2 negative.

Actionable alterations (AA) in gastrointestinal (GI) cancers: Rate of detection and receipt of matched therapies (MT).

e15677 Background: Next generation sequencing (NGS) is widely used in pts with advanced cancer to personalize care. Current NCCN guidelines endorse Her2, PD-L1 and MSI testing in esophagogastric cancer (EGC), RAS, BRAF, Her2 and MSI testing in colorectal cancer (CRC) and germline, somatic and MSI testing in pancreas cancer (PC). The proportion of GI cancer pts who receive MT based on NGS is unclear. Methods: We identified pts with advanced EGC (2016-18), PC (2017) and CRC (2016) who underwent NGS with MSK-IMPACT. We assessed the proportion of pts with ≥1 AA as defined by OncoKB (at the time of analysis, levels 1/2a were accepted practice and levels 2b/3/4 were investigational; Chakravaty, JCO PO 2017), those who received MT on trial or off label and 3 and 6 months (mos) progression-free survival (PFS). Results: We identified 260 EGC, 357 PC and 438 CRC pts. After excluding pts who had ongoing benefit from standard therapy (tx), were treated elsewhere or had no active stage IV disease, potential level 2/3/4 AAs occurred in 37% (n = 97) of EGC pts, 32.5% (n = 116) of PC pts and 26.7% (n = 117) of CRC pts (Table). 10, 1 and 17 pts with EGC, PC and CRC respectively were MSI. 1 pt in each subtype had an NTRK fusion (OncoKB level 1). In EGC, 6 pts (6.2% of those with AAs) received MT: 2 pts with MET amplification (a) and 1 each with BRCA2 mutation (m), TSC2m, ERBB2m and EGFRa. The pts with METa treated with crizotinib achieved 3 but not 6 mos PFS. In PC, 11 pts (9.5%) got MT: 10 pts for BRCAm and 1 for NTRK3 fusion. 9 pts with BRCAm treated with PARP inhibitors (i) achieved ≥3 mos PFS and 5 pts reached ≥6 mos PFS. The pt treated with NTRKi progressed rapidly. In CRC, 5 pts with ERBB2a and 9 pts with BRAFm received MT (12%). 3 pts and 2 pts treated with anti-Her2 tx achieved ≥3 and ≥6 mos PFS respectively. Of 6 pts treated with BRAF/MEKi plus irinotecan or anti-EGFR tx, all achieved ≥3 mos PFS; 3 reached ≥6 mos PFS. 3 pts received novel BRAF and ERK1/2i; none reached 3 mos PFS. Conclusions: NGS frequently identified OncoKB level 2 AAs. Few pts received MT, and of those, some achieved ≥6 mos PFS. Pts with CRC and PC received MT which subsequently became standard NCCN recommendations; therefore, a more current analysis may show increased MT use. Still, MT for level 3 and 4 alterations were rare, suggesting expectations of NGS must be managed appropriately. [Table: see text]

Safety and efficacy of pyrotinib in patients with NSCLC and other advanced solid tumors with activating HER2 alterations: A phase I basket trial.

3510 Background: Pyrotinib is a potent, irreversible tyrosine kinase inhibitor (TKI) that blocks signal transduction through the erythroblastic leukemia viral oncogene homolog (erbB) receptors, which has previously demonstrated promising antitumor activity in patients (pts) with breast cancers. We studied the safety and efficacy of pyrotinib in pts with non-small cell lung cancer (NSCLC) and other solid tumors with activating human epidermal growth factor receptor 2 ( HER2, ERBB2) alterations. Methods: This is an open-label, multicenter phase 1 dose expansion basket trial of pyrotinib given 400mg oral daily at 28-day cycles. Expansion cohorts included pts with HER2-mutant NSCLC and advanced solid tumors with HER2 mutation or amplification. HER2 testing was conducted using next generation sequencing or fluorescence in situ hybridization. Primary endpoints included toxicities as evaluated by NCI CTCAE v5.0, and overall response rate (ORR) as evaluated by RECIST v1.1. Secondary objectives included progression-free survival (PFS). Results: A total of 62 pts were enrolled. The median age was 67 (range 40 – 86 years), 61% were female and the median lines of prior systemic therapy was 3 (range 1-11). There were no treatment related deaths. The most common adverse events were diarrhea (96.8%), nausea (82.3%) and vomiting (41.9%). The only ≥ grade 3 treatment related toxicity was diarrhea (24.2%). Prophylactic anti-diarrhea treatment was introduced to facilitate continuation of pyrotinib. At the Jan 13, 2020 cut-off, 24 pts with HER2-mutant NSCLC (20, i.e. 65% of which were the A775_G776insYVMA mutation) and 18 pts with HER2-mutant or amplified solid tumors completed end of Cycle 2 imaging scan and were evaluable for tumor responses. The ORR was 19% (8/42, 95% CI 7-31%); confirmed responses include a complete response (CR) and 3 partial responses (PRs) in HER2-mutant NSCLC, and 4 PRs in HER2-amplified cholangiocarcinoma, ovarian, endometrial and salivary gland carcinomas. There were 7 stable disease ≥ 6 months. Median progression-free survival was 5.4 months (95% CI 4.4-7.3). Conclusions: Pyrotinib demonstrated a manageable safety profile and encouraging efficacy in pts with heavily pre-treated HER2-mutant NSCLC. Furthermore, it is the first TKI to produce durable responses in pts with HER2-amplified biliary tract, ovarian, endometrial and salivary gland cancers. These results warrant further clinical development of pyrotinib. Clinical trial information: NCT02500199 .

Comprehensive analysis of HER2 status through genomic, transcription, and translation among 5,305 patients with diverse malignancies.

3632 Background: HER2 alterations is a predictive biomarker for anti-HER2 regimen. Success in breast and gastric cancers with anti-HER2 therapies translated to explore their efficacy in other tumors. Currently, measurement of HER2 can be done by assessing the expression of protein [e.g. immunohistochemistry (IHC)] or gene amplification of copy number variation (CNV) [e.g. fluorescence in situ hybridization or next-generation sequencing (NGS)]. But little is known about the transcription level (mRNA) of HER2. Herein, we investigated HER2 mRNA expression and its association with gene and protein expressions among diverse cancer types. Methods: Between 2015-2019, HER2 status was evaluated using IHC, qRT-PCR and NGS by Paradigm Diagnostics (CLIA-certified laboratory). All tumors in the database were included for analysis. Correlations between all 3 tests were done. An illustrative patient who was treated with anti-HER2 therapy base on the mRNA testing is presented. Results: HER2 testing was performed on 5305 patients (pts) with diverse cancers including NSCLC (n=1175), breast (n=1040) and colon (n=566); 4.1% (161/3926) had amplification through NGS, 33.3% (615/1848) had mRNA overexpression and 9.3% (236/2533) had overexpression by IHC. Of 723 pts who had all three tests performed, we found 7.5% (54/723) of pts with all three HER2 markers being positive (CNV [+]/mRNA [+]/ IHC [+]). Meanwhile, variety of amplification/ expression patterns were seen (see Table). CNV positivity translated to protein expression in 95% of cases. While only 4% of pts were IHC positive when CNV and mRNA were negative. 20% (144/723) of pts had mRNA overexpression alone among diverse cancer types. Representative case of 70yo female with metastatic cholangiocarcinoma harboring mRNA overexpression (but negative for CNV, IHC unclear due to sample insufficiency) who had near complete response to anti-HER2 therapy with progression-free survival of 24+ months is presented. Conclusions: HER2 status can be discordant with different assays but NGS positivity has excellent correlation with mRNA and protein expression. Of importance, HER2 mRNA can be overexpressed in 20% of pts even when gene amplification and protein expression are negative. Further studies are warranted to determine the clinical utility of mRNA as a biomarker for HER2 and potential use for anti-HER2 targeted therapies. [Table: see text]

Breast cancer receptor subtypes in East Africa: A systematic review and meta-analysis.

e19074 Background: Estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) are determinants of both treatment and mortality for patients with breast cancer (BC). With a mortality to incidence ratio of 0.51, mortality rates for breast cancer in East Africa are among the world’s highest. Building on a meta-analysis of BC receptor subtypes in sub-Saharan Africa (2014) and including all recently available PubMed literature, our meta-analysis aims to investigate the differences among receptor subtype distribution in five East African countries. Methods: Receptor distributions for female samples &gt;30 in all BC articles for Ethiopia, Kenya, Rwanda, Tanzania and Uganda were obtained from PubMed, January 1, 1998 - June 30, 2019. Outcomes were proportions of ER+, PR+, and HER2+ and/or molecular subtypes. Data included study characteristics and mean/median patient age (weighted by observations). Using metaprop for STATA 16, we estimated pooled proportions (ES) with 95% confidence intervals (CI) and assessed I2 statistics for heterogeneity across studies. Results: Among 35 BC studies with receptor data, 21 met criteria. Overall, the weighted mean age was 47.5 years (SD 3.2) and median, 48. For ER+ (18 studies/n=2,875), the overall ES was 0.55 (95% CI 0.47, 0.62), I2 of 94.4%, p&lt;0.001, ranging from 0.62 in Ethiopia (95% CI 0.52, 0.72) to 0.42 in Uganda (95%0.36, 0.49). For HER2+ (18 studies/n=2,689), the overall ES was 0.23 (95% CI 0.20, 0.26), I2 of 70.2%, p&lt;0.001, and ranged from 0.27 in Ethiopia (95% CI 0.18, 0.36) to 0.21 in both Tanzania and Uganda (95% CI 0.13/0.14, 0.28). For triple negative (TN, 15 studies/n=2,510), the overall ES was 0.28 (95% CI 0.23, 0.32), I2 of 84.0%, p&lt;0.001, varying from 0.21 in Ethiopia (95% CI 0.15, 0.27) to 0.46 in Tanzania (95% CI 0.32, 0.60). Conclusions: We found differences between countries, e.g. lower distribution of TN in Ethiopia (21%) compared to Uganda (35%) and Tanzania (46%), and high between-study heterogeneity. Consistent with former studies, ER+, 55%, was the dominant subtype, particularly in Ethiopia, Kenya and Rwanda, indicating the need to prioritize hormonal treatment. Overall rates of HER2 subtype, 23%, approached overall rates of TN, 28%, yet HER2 testing is often not performed due to the cost and unavailability of anti-HER2 therapy, hence this subtype often goes untreated. These findings highlight the benefit of testing and reporting of receptor subtypes at the country level to promote delivery of more effective treatment to reduce the mortality disparity.

Adding precision to 2018 ASCO/CAP HER2 testing guidelines in breast cancer with genomic profiling.

3570 Background: Biological heterogeneity of HER2 positive breast cancers has been suggested by a modest benefit of HER2-targeted therapies reported in the APHINITY and ExteNET trials. This highlights the need for improved biomarkers that more precisely identify patients who benefit from HER2-directed agents. The 80-gene molecular subtyping signature (80GS) classifies breast tumors into Luminal, HER2 or Basal type based on the gene expression of downstream signaling pathways. Previous work showed a substantial proportion of tumors identified as HER2 equivocal or HER2 positive by 2013 ASCO/CAP guidelines may be reclassified as non-HER2 type by 80GS. In 2018, ASCO/CAP HER2 IHC/ISH classification guidelines were revised to reduce the frequency of HER2 equivocal cases, for which treatment recommendations have been ambiguous. Here we evaluated concordance between HER2 status by 2018 ASCO/CAP guideline classification and 80GS molecular subtyping. Methods: Pathology reports are provided by physicians for samples that are tested with the 70-gene risk of distant recurrence signature (70GS) and 80GS as part of routine diagnostic care. This analysis includes data sent to Agendia (Irvine, CA) from January 2019 to January 2020. HER2 IHC/ISH results based on ASCO/CAP 2018 guidelines were available for 1453 samples. Results: Of 1453 samples, 1336 (92%) were HER2 negative, 99 (7%) were HER2 positive, and 18 (1.2%) were HER2 equivocal under 2018 guidelines. 80GS reclassified 57 of 99 (58%) HER2 positive tumors as Luminal and 11 of 99 (11%) as Basal; the remaining 31% were confirmed HER2. Furthermore, 55 of 99 (55%) HER2 positive tumors were also ER and PR positive by IHC, with 48 (87%) of these reclassified as Luminal type. Of HER2 negative tumors, 80GS classified 94 of 1336 (7%) as Basal and 2 of 1336 (0.15%) as HER2. Of HER2 equivocal tumors, 16 of 18 (89%) reclassified as Luminal and 2 of 18 (11%) as Basal. Conclusions: In this real-world diagnostic dataset, 2018 ASCO/CAP guidelines resulted in few HER2 equivocal tumors overall, confirming the positive impact of the revised guidelines. However, 80GS reclassified 69% of HER2 positive tumors to non-HER2 molecular subtypes, suggesting these tumors may have suboptimal responses to HER2-directed therapy compared to HER2 enriched. All HER2 equivocal tumors reclassified to non-HER2 subtypes. Molecular classification by 80GS adds further precision in classifying HER2 positive patients and potential to predict responsiveness to HER2-targeted therapies. Further studies are warranted to validate the utility of HER2 status based on 80GS.

Primary results of NRG Oncology / NSABP B-43: Phase III trial comparing concurrent trastuzumab (T) and radiation therapy (RT) with RT alone for women with HER2-positive ductal carcinoma in situ (DCIS) after lumpectomy.

508 Background: Preclinical studies report that T can boost RT effectiveness. The primary aim of this trial assessed the efficacy of concurrent T + RT vs RT alone in preventing recurrence of ipsilateral breast cancer, ipsilateral skin cancer, or ipsilateral DCIS (IBTR) in women with DCIS. Methods: Eligibility: Women ≥18 yrs, ECOG performance status 0 or 1, DCIS resected by lumpectomy, and clear margins. Whole-breast RT after randomization was with 25+ fractions or accelerated with 16-17 fractions. RT boost was allowed. Centralized HER2 testing and ER and/or PR were required before entry. Stratification was by menopausal status, adjuvant endocrine therapy plan, and nuclear grade. T was given at 8 mg/kg IV within 1 wk before and 5 days after RT began (Dose 1) and at 6 mg/kg IV 3 wks after Dose 1 (Dose 2). Definitive intent-to-treat primary analysis was to be conducted when either 163 IBTR events were recorded or when all accrued pts were on study for ≥5 yrs. Results: 2014 pts were randomized (11/9/08 to 12/8/14);1998 (99.2%) had follow-up information. Median follow-up time on 12/31/19 was 79.2 mos. 2001 pts had RT information, 1965 (98.2%) completed RT: 988 (98.3%) in the RT arm and 977 (98.1%) in the RT+T arm. 996 pts had T compliance information in the RT+T arm, 939 (94.3%) completed two doses of T, 25 (2.5%) had one dose of T, and 32 (3.2%) did not receive T. At primary definitive analysis, 114 IBTR events occurred: 63 in the RT arm and 51 in the RT+T arm (HR = 0.81 [95% CI: 0.56-1.17], p-value = 0.26). 38 were invasive: 18 in the RT arm and 20 in the RT+T arm (HR = 1.11 [95% CI: 0.59-2.10], p-value = 0.74). 76 were DCIS: 45 in the RT arm and 31 in the RT+T arm (HR = 0.68 [95% CI: 0.43-1.08], p-value = 0.10). Annual IBTR event rates were 0.99%/yr in the RT group and 0.80%/yr in the RT+T group. There were 288 events of any kind [iDFS-DCIS] (DFS): 155 in the RT arm and 133 in the RT+T arm (HR = 0.84 [95% CI: 0.66-1.05], p-value = 0.13) and 48 deaths: 26 in the RT arm and 22 in the RT+T arm (OS HR = 0.85 [95% CI: 0.48-1.51], p = 0.59). The study did not reach the 163 protocol-specified events, so the definitive analysis was triggered by all pts having been on study for ≥5 years. Conclusions: The addition of T to RT did not achieve the protocol objective of 36% reduction in the IBTR rate but did achieve a modest, statistically non-significant reduction of 19%. Support: U10-180868, -180822, UG1-189867; Genentech. The authors thank Elaina Harper and Marlon Jones for data management. Clinical trial information: NCT00769379 .

Real-world treatment patterns among metastatic breast cancer patients by BRCA status and phenotype.

e13046 Background: BRCA1/2 mutations account for 5-10% of breast cancer (BC) patients. BRCA1 mutations are associated with aggressive tumors, often triple negative (TNBC) (HER2, estrogen-receptor [ER], progesterone-receptor [PR] negative) and are mainly treated with combination chemotherapy or platinum agents. This study aimed to compare real world treatment patterns in breast cancer patients who were BRCA-mutation positive (BRCA+) and those who were BRCA-mutation negative (BRCA-), focusing on hormone-receptor positive (HR+) and TNBC subgroups. Methods: Optum’s Humedica electronic health records (EHR) database was used to identify a cohort of women with 1) confirmed BC ICD-9/ICD-10 diagnosis between 2008 and 2018, 2) at least 12 months of history in EHR, 3) at least one distant metastasis diagnosis (per ICD-9/ICD-10 or physician notes), and 4) no previous diagnosis of other primary cancers in previous 12 months. HR and HER2 test results were used to create relevant subgroups. Distinct lines of therapy (LOT) in the post-metastatic period were established using business rules. Treatment patterns and duration of treatment were analyzed. Results: Among 65,934 metastatic BC patients identified, 7941 were BRCA+, of whom 5366 (68%) were HR+ and 1323 (17%) had TNBC phenotype, while 4027 were BRCA-, of whom 2859 (71%) were HR+ and 561 (14%) had TNBC phenotype. Seventy-one percent of BRCA+ women underwent drug treatment, compared to 65% of BRCA- women. Drug treatments included hormonal treatment (+/-other agents), targeted therapy, and chemotherapy only, and varied across the different cohorts (Table). Mean (median) duration of treatment in LOT1 in the BRCA+ and BRCA- cohorts was 117 (73) and 117 (72) days, respectively. Conclusions: Real-world evidence on treatment patterns in BC patients revealed substantial differences across important biomarker and hormonal status BC phenotypes. As a preponderance of women with TNBC are still being treated exclusively with chemotherapy, these data highlight how the need for development of effective treatment options for TNBC remains high. [Table: see text]

A basket trial of trastuzumab deruxtecan, a HER2-targeted antibody-drug conjugate, for HER2-amplified solid tumors identified by circulating tumor DNA analysis (HERALD trial).

TPS3650 Background: Trastuzumab deruxtecan, a new HER2-targeting antibody-drug conjugate, has been approved for unresectable or metastatic HER2-positive breast cancer by the Food and Drug Administration. In a phase I/II trial, trastuzumab deruxtecan showed a manageable safety profile and antitumor activity in HER2-positive various cancer types. In addition, a tissue-based HER2 test occasionally cannot identify accurate HER2 status due to spatial and temporal intratumoral heterogeneity, leading to potentially missing an opportunity for responders to receive benefit from anti-HER2-targeted therapy. Circulating tumor DNA (ctDNA) analysis can detect comprehensive somatic genome alterations by assessment of spatial and temporal intratumoral heterogeneity with minimal invasiveness. Methods: We designed an investigator-initiated multicenter phase II basket trial to evaluate efficacy and safety of trastuzumab deruxtecan in advanced solid tumor malignancies with HER2 amplification identified by Guardant360, a 74-gene sequencing ctDNA panel, as a part of the Nationwide Cancer Genome Screening Project (GOZILA study, UMIN000029315). The key eligibility criteria are as follows: 1) Histopathologically confirmed advanced solid tumor malignancy; 2) Identified HER2 amplification by Guardant360; 3) Failed prior standard therapy. The participants will receive intravenously 5.4 mg/kg of trastuzumab deruxtecan every 3 weeks. The primary endpoint is objective response rate (ORR). The planned sample size is 55-65. A Bayesian model considering the potential heterogeneity across cancer types will be applied to detect ORR of 5% versus 25% to a certain level while maintaining the false-positive error rate in each cancer type at 10%. Furthermore, tumor tissue, ctDNA and circulating tumor cells are serially collected and analyzed to investigate the predictive biomarkers and resistance mechanisms. The trial was activated in late 2019. At the time of the abstract submission, 2 patients have been enrolled. This trial is granted by AMED under Grant Number JP18lk0201084. Clinical trial information: JapicCTI-194707 .

HER2 testing across practices-Have we come to a consensus on the ideal method of testing? A systematic literature review.

Human epidermal growth factor (HER2) is an oncogene that codes for HER2 protein. The gene is amplified, and protein is overexpressed in 20% of patients and carries a poor prognosis. HER2-positive tumors tend to be more aggressive and highly proliferative. In 2006, trastuzumab, a monoclonal antibody targeting HER2, was approved for use with chemotherapy as an adjuvant treatment for women with HER2-positive breast cancer. The drug has shown improved survival rates for women with HER2-positive breast cancer. As promised for survival in HER2-positive patients continues with new research, and as novel drug approvals emerge, HER2 assessment plays a significant role in adjuvant and in metastatic setting. HER2 assays approved by the US Food and Drug administration include immunohistochemistry (IHC), fluorescence in situ hybridization (FISH), and bright field dual in situ hybridization (DISH). Accuracy of HER2 testing across laboratories has an impact on treatment as false-negative testing can deprive the patients of trastuzumab therapy. The current review will focus on the variability of HER2 testing across laboratories and the potential impact on treatment.