anti-HER2 Therapy In Breast Cancer Research Articles

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

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

N6-methyladenosine regulated FGFR4 attenuates ferroptotic cell death in recalcitrant HER2-positive breast cancer

Intrinsic and acquired anti-HER2 resistance remains a major hurdle for treating HER2-positive breast cancer. Using genome-wide CRISPR/Cas9 screening in vitro and in vivo, we identify FGFR4 as an essential gene following anti-HER2 treatment. FGFR4 inhibition enhances susceptibility to anti-HER2 therapy in resistant breast cancer. Mechanistically, m6A-hypomethylation regulated FGFR4 phosphorylates GSK-3β and activates β-catenin/TCF4 signaling to drive anti-HER2 resistance. Notably, suppression of FGFR4 dramatically diminishes glutathione synthesis and Fe2+ efflux efficiency via the β-catenin/TCF4-SLC7A11/FPN1 axis, resulting in excessive ROS production and labile iron pool accumulation. Ferroptosis, a unique iron-dependent form of oxidative cell death, is triggered after FGFR4 inhibition. Experiments involving patient-derived xenografts and organoids reveals a synergistic effect of anti-FGFR4 with anti-HER2 therapy in breast cancer with either intrinsic or acquired resistance. Together, these results pinpoint a mechanism of anti-HER2 resistance and provide a strategy for overcoming resistance via FGFR4 inhibition in recalcitrant HER2-positive breast cancer.

72P Prognostic value of HER2/CEP17 ratio in HER2-positive early breast cancer by FISH testing

Human Epidermal Growth Factor Receptor 2 (HER2) fluorescence in situ hybridization (FISH)-positivity requires anti-HER2 therapy in breast cancer (BC). This study aimed to determine whether the expression of HER2/centromeric probe for chromosome 17 (CEP17) ratio was associated with prognosis of early HER2-positive BC.

An Audit to Evaluate the Assessment, Monitoring and Management of Cardiac Health in Patients Receiving Anti-HER2 Therapy for Breast Cancer – a Single Institution Experience

Purpose: This audit aimed to assess current standards with regards to the assessment and management of cardiac health in patients receiving anti-HER2 therapy. Therapeutic monoclonal antibodies to the HER2 receptor can also bind to myocytes expressing HER2, resulting in left ventricular dysfunction and clinical heart failure. Therefore, echocardiogram monitoring is essential [1]. Practice was assessed against the UK National Cancer Research Institute recommendations for prevention, monitoring and management of cardiac events in trastuzumab-treated patients [2].

Anti-HER2 therapy for breast cancer in older patients.

Older patients now form between 30 and 40% of breast cancer (BC) patients. Managing older patients with BC is particularly challenging due to the limited availability of high-quality evidence. In this review we discuss the available evidence on the efficacy and safety of anti-HER2 agents in older patients with HER2-positive BC is presented, with a particular look at the latest results of promising new agents such trastuzumab-deruxtecan. The data suggest that older patients can expect similar efficacy when using standard regimens, with higher toxicity, particular cardiac toxicity and diarrhea. Anti-HER2 agents should thus be used in most older patients most as per standard of care as long as adequate follow-up is available to manage toxicities.

P179 - Positive HER3 could be a marker of poor response to neoadjuvant dual anti-HER2 therapy in breast cancer

P179 - Positive HER3 could be a marker of poor response to neoadjuvant dual anti-HER2 therapy in breast cancer

Synthetic lethality between HER2 and transaldolase in intrinsically resistant HER2-positive breast cancers

Intrinsic resistance to anti-HER2 therapy in breast cancer remains an obstacle in the clinic, limiting its efficacy. However, the biological basis for intrinsic resistance is poorly understood. Here we performed a CRISPR/Cas9-mediated loss-of-function genetic profiling and identified TALDO1, which encodes the rate-limiting transaldolase (TA) enzyme in the non-oxidative pentose phosphate pathway, as essential for cellular survival following pharmacological HER2 blockade. Suppression of TA increases cell susceptibility to HER2 inhibition in two intrinsically resistant breast cancer cell lines with HER2 amplification. Mechanistically, TA depletion combined with HER2 inhibition significantly reduces cellular NADPH levels, resulting in excessive ROS production and deficient lipid and nucleotide synthesis. Importantly, higher TA expression correlates with poor response to HER2 inhibition in a breast cancer patient cohort. Together, these results pinpoint TA as a novel metabolic enzyme possessing synthetic lethality with HER2 inhibition that can potentially be exploited as a biomarker or target for combination therapy.

Clinical Significance of Glycoprotein Non-metastatic B and Its Association with EGFR/HER2 in Gastrointestinal Cancer.

Glycoprotein non-metastatic B (GPNMB), a type I transmembrane glycoprotein, is overexpressed in melanoma and breast cancer and promotes cancer-cell invasion and motility. We previously reported cross-talk between GPNMB and human epidermal growth factor receptor 2 (HER2) in breast cancer, suggesting that GPNMB might play an important role in resistance to anti-HER2 therapy in breast cancer. Here, we clarified the association between GPNMB and HER-family proteins in gastrointestinal cancer by examining their relationships using gastric and colorectal cancer cell lines. We found that GPNMB depletion of by small-interfering RNA increased epidermal growth factor receptor (EGFR) expression and phosphorylation through AKT8 virus oncogene cellular homolog (AKT) and mitogen-activated protein kinase (MAPK) pathways. Additionally, treatment with cetuximab (CTX) also increased GPNMB expression, and combination therapy consisting of GPNMB depletion and CTX treatment significantly suppressed cell growth in colorectal cancer cell lines, but not in gastric cancer cell lines. Furthermore, we also evaluated changes in GPNMB expression in vivo, with immunohistochemistry detecting GPNMB overexpression in a colorectal cancer patient following anti-EGFR therapy. These results suggested possible cross-talk between GPNMB and EGFR, and that GPNMB might play an important role in resistance to anti-EGFR therapy in gastrointestinal cancer.

DUSP4 is associated with increased resistance against anti-HER2 therapy in breast cancer.

The majority of patients develop resistance against suppression of HER2-signaling mediated by trastuzumab in HER2 positive breast cancer (BC). HER2 overexpression activates multiple signaling pathways, including the mitogen-activated protein kinase (MAPK) cascade. MAPK phosphatases (MKPs) are essential regulators of MAPKs and participate in many facets of cellular regulation, including proliferation and apoptosis. We aimed to identify whether differential MKPs are associated with resistance to targeted therapy in patients previously treated with trastuzumab. Using gene chip data of 88 HER2-positive, trastuzumab treated BC patients, candidate MKPs were identified by Receiver Operator Characteristics analysis performed in R. Genes were ranked using their achieved area under the curve (AUC) values and were further restricted to markers significantly associated with worse survival. Functional significance of the two strongest predictive markers was evaluated in vitro by gene silencing in HER2 overexpressing, trastuzumab resistant BC cell lines SKTR and JIMT-1. The strongest predictive MKPs were DUSP4/MKP-2 (AUC=0.75, p=0.0096) and DUSP6/MKP-3 (AUC=0.77, p=5.29E-05). Higher expression for these correlated to worse survival (DUSP4: HR=2.05, p=0.009 and DUSP6: HR=2, p=0.0015). Silencing of DUSP4 had significant sensitization effects – viability of DUSP4 siRNA transfected, trastuzumab treated cells decreased significantly compared to scramble-siRNA transfected controls (SKTR: p=0.016; JIMT-1: p=0.016). In contrast, simultaneous treatment with DUSP6 siRNA and trastuzumab did not alter cell proliferation. Our findings suggest that DUSP4 may represent a new potential target to overcome trastuzumab resistance.

Comparison of neoadjuvant anti-HER2 therapy for breast cancer in public versus private health care: Time to shorten the distance.

e18045 Background: In the neoadjuvant setting (NAC) of HER2-positive breast cancer (BC), the addition of pertuzumab (P) to trastuzumab (T) and chemotherapy (chemo) has increased pathological complete response (pCR) from 40-50% to over 70%. Recent data has shown that pCR is a reliable surrogate marker of disease-free (DFS) and overall survival (OS). In Brazil, there is an important gap between cancer drugs approved in the public health care system (PHCS) vs. the private setting. Although P approval for NAC (Aug/2016) and metastatic setting (Sep/2015) in the private health system, it is not approved for any indication in the PHCS. Likewise, T is not approved for use as NAC at the PHCS. We aim to compare private practice experience of T + P + chemo in the NAC vs. HER2-positive advanced BC patients treated at the PHCS. Methods: Retrospective analysis of 95 patients diagnosed with locally advanced HER2-positive BC by immunohistochemistry 3+ or FISH according to ASCO/CAP definition. We evaluated 17 consecutive patients at the COAEM private cancer center in Brazil since Feb/2014 (cohort 1). Neoadjuvant therapy was T and P plus docetaxel-based chemo. A cohort of 78 patients with locally advanced HER2-positive BC treated in a PHCS cancer center was analyzed (cohort 2). They received standard NAC regimen according to PHCS guidance: anthracycline and taxane-based therapy without anti-HER2 therapy. Results: Cohort 1: 17 patients with median age of 50 years (range 34-70); Four patients had clinical stage IIA; 2 patients IIB; 7 patients IIIA; 3 patients IIIB; one patient IIIC. The pCR rate was 70.5% (12 patients). Cohort 2: 78 patients with median age of 48 years (range 26-79). One patient IIB; 33 patients IIIA; 37 patients IIIB; 7 patients IIIC. The pCR rate was 16.7% (13 patients); 29 patients (37.2%) had clinical complete response. Only 26 patients (33.3%) received T in the adjuvant setting. Conclusions: Neoadjuvant P and T combined with chemo has shown pCR rates superior to 70% in clinical trials, which is a surrogate marker for DFS and OS in HER2-positive BC. The absence of anti-HER2 therapy in the PHCS must be urgently reviewed not just in light of double-blockage strategy but also in T monotherapy use.

Hormone receptor status does not alter the effect of trastuzumab in breast cancer

Overexpression of human epidermal growth factor receptor 2 (HER2) predicts response to anti-HER2 therapy in breast cancer. We investigated whether hormone receptor (HR) status influences the treatment benefit of trastuzumab in patients with breast cancer. Data from 8338 patients with primary nonmetastatic breast cancer from the cancer registry of Saxony-Anhalt Germany were analyzed. A total of 5554 patients were eligible for analysis. The median follow-up of the study was 6 years. Of the 5554 patients investigated, 1128 (20.3%) showed HER2 overexpression and 656 (58.2%) of them received adjuvant trastuzumab. The 10-year overall survival (OS) in the study cohort according to HR, HER2 status, and trastuzumab treatment was as follows: 78.4% for HR-/HER2-, 85.0% for HR+/HER2-, 70.4% HR-/HER2+/TRA-, 71.4% for HR+/HER2+/TRA-, 80.9% for HR-/HER2+/TRA+, and 89.2% for HR+/HER2+/TRA+. Trastuzumab treatment improved OS in the HR- patients only in the first 3 years, whereas in the HR+ group the effect of trastuzumab was still apparent 5 years after diagnosis. Notably, the relative improvement in a patient outcome was higher for HR+ patients. Nevertheless, matching for age, histological type, tumor stage, tumor grade, and performance status between patients with HR- and HR+ tumors demonstrated that the survival effect of trastuzumab was not affected by HR status; P=0.890. Trastuzumab treatment improves patients' survival regardless of HR status and should be offered to all HER2+ patients.

Abstract P5-08-22: DUSP4 is associated with increased resistance against anti-HER2 therapy in breast cancer

Abstract Background. The majority of patients develop resistance against suppression of HER2-mediated signaling by trastuzumab in HER2 positive breast cancer (BC). HER2 overexpression activates multiple signaling pathways, including the mitogen-activated protein kinase (MAPK) cascade. MAPK phosphatases (MKPs) are essential regulators of MAPKs and participate in many facets of cellular regulation, including proliferation and apoptosis. We aimed to identify whether differential MKPs are associated with resistance to targeted therapy in patients previously treated with trastuzumab. Methods. Using Affymetrix HGU133plus2 gene chip data of 88 HER2-positive, trastuzumab treated BC patients, candidate MKPs were identified by Receiver Operator Characteristics analysis performed in R. Genes were ranked using their achieved area under the curve (AUC) values and were further constricted to those markers significantly associated to worse survival. Functional significance of the two strongest predictive biomarkers was evaluated in vitro experiments after gene silencing in the HER2 overexpressing, trastuzumab resistant breast cancer cell lines SKBR-3-TR and JIMT-1. Results. Out of 10 investigated MKPs, the strongest predictive genes were DUSP4/MKP2 (AUC=0.75, p=0.0096) and DUSP6/MKP3 (AUC=0.77, p=5.29E-05). Furthermore, higher expression for these correlated to worse survival in 221 HER2 positive BC patients (DUSP4: HR=1.6, p=0.04 and DUSP6: HR=1.8, p=0.0053). Silencing of DUSP4 had significant sensitization effects - viability of DUSP4 siRNA transfected, trastuzumab treated cells decreased significantly compared to scramble-siRNA transfected, trastuzumab treated controls (SKBR-3-TR: p=0.016; JIMT-1: p=0.016). In contrast, simultaneous treatment with DUSP6 siRNA and trastuzumab did not alter cell proliferation. Conclusions. Our findings suggest that DUSP4 is involved in the development of trastuzumab resistance in HER2 positive BC. Citation Format: Györffy B, Munkacsy G, Esteva FJ, Miquel TP, Menyhart O. DUSP4 is associated with increased resistance against anti-HER2 therapy in breast cancer. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P5-08-22.

Mechanisms of resistance and sensitivity to anti-HER2 therapies in HER2+ breast cancer.

Breast Cancer (BC) is a highly prevalent disease. A woman living in the United States has a 12.3% lifetime risk of being diagnosed with breast cancer [1]. It is the most common female cancer and the second most common cause of cancer death in women [2]. Of note, amplification or overexpression of Human Epidermal Receptor 2 (HER2) oncogene is present in approximately 18 to 20% of primary invasive breast cancers, and until personalized therapy became available for this specific BC subtype, the worst rates of Overall Survival (OS) and Recurrence-Free Survival (RFS) were observed in the HER2+ BC cohort, compared to all other types, including triple negative BC (TNBC) [3].HER2 is a member of the epidermal growth factor receptor (EGFR) family. Other family members include EGFR or HER1, HER3 and HER4. HER2 can form heterodimers with any of the other three receptors, and is considered to be the preferred dimerization partner of the other HER or ErbB receptors [4]. Phosphorylation of tyrosine residues within the cytoplasmic domain is the result of receptor dimerization and culminates into initiation of a variety of signalling pathways involved in cellular proliferation, transcription, motility and apoptosis inhibition [5].In addition to being an important prognostic factor in women diagnosed with BC, HER2 overexpression also identifies those patients who benefit from treatment with agents that target HER2, such as trastuzumab, pertuzumab, trastuzumab emtansine (T-DM1) and small molecules tyrosine kinase inhibitors of HER2 [6, 11, 127]. In fact, trastuzumab altered the natural history of patients diagnosed with HER2+ BC, both in early and metastatic disease setting, in a major way [8–10]. Nevertheless, there are many women that will eventually develop metastatic disease, despite being treated with anti-HER2 therapy in the early disease setting. Moreover, advanced tumors may reach a point where no anti-HER2 treatment will achieve disease control, including recently approved drugs, such as T-DM1.This review paper will concentrate on major biological pathways that ultimately lead to resistance to anti-HER2 therapies in BC, summarizing their mechanisms. Strategies to overcome this resistance, and the rationale involved in each tactics to revert this scenario will be presented to the reader.

Non-HER2 signaling pathways activated in resistance to anti-HER2 therapy in breast cancer.

HER2 receptor is overexpressed approximately in 20 % of human breast cancer (BC) and is a poor prognostic factor. Although therapies targeting this receptor have improved the prognosis of this cancer, up to 62 % patients treated with these drugs experiment progression during the first year of treatment. Some molecular mechanisms have been proposed to be responsible for this resistance, such as activation of alternative signaling pathways (through ERBB receptors and non-ERBB receptors or increased expression of ligands and alterations in HER2 signaling components). In this article, we will review the influence of genetic markers in non-HER2 signaling pathways investigated to date as cause of resistance to HER2-targeted drugs in HER2-positive BC patients. GRB7, included in the 17q12 amplicon, has been associated to poor prognosis in BC patients. Biomarkers like EPHAR and SRC, have demonstrated clinical relevance and prognostic value in HER2-positive BC patients. Non-invasive biomarkers, such as elevated IGF1 serum levels have been revealed as interesting biomarkers to be considered as predictors of trastuzumab clinical outcomes in BC patients. However, the prognostic value of most of the biomarkers investigated to date, such as HER3, IGF1R, PIK3CA, or AKT1 cannot be fully established yet, since results have not been conclusive.

Moderate HER2 expression as a prognostic factor in hormone receptor positive breast cancer.

Overexpression and/or amplification of human epidermal growth factor receptor 2 (HER2) is associated with poor prognosis in breast cancer and predicts response to anti-HER2 therapy in breast cancer. The prognostic relevance of moderate expression of HER2 is unclear. Data of 9872 patients with primary nonmetastatic breast cancer from the cancer registries of Magdeburg and Halle, Germany, were analyzed retrospectively. A total of 5907 patients with complete data sets including follow-up were eligible for analysis. HER2 status was determined as recommended by international guidelines. Of 5907 patients investigated, 5023 (68.4%) had HER2 0 and 1+ expression and 884 (12.0%) had HER2 (2+)/HER2- expression. Patients with hormone receptor positive (HR+) and HER2 (2+) tumors had a shorter median disease-free survival (DFS; P<0.0001) and breast cancer specific survival (BCSS; P=0.019) than HR+ patients with HER2 (0/1+) tumors. Among patients with HR- breast cancer there was no significant difference between HER2 (2+) and HER2 (0/1+) tumors. In multivariate analysis after adjustment for other prognostic factors, HER2 (2+) status remained an unfavorable prognostic factor for DFS (hazard ratio (HR)=1.217, 95% CI=1.052-1.408; P=0.008) but not for BCSS (HR=1.045, 95% CI=0.926-1.178; P=0.474). The HER2 (2+) status is an unfavorable prognostic factor for survival of patients with HR+ breast cancer. The impact of anti-HER2 therapy in this group of patients should be evaluated.

Resistance to Anti-HER2 Therapies in Breast Cancer.

HER2 is amplified or overexpressed in 20% to 25% of breast cancers. HER2 is a redundant, robust, and powerful signaling pathway that represents an attractive therapeutic target. Anti-HER2 therapy in the clinic has resulted in significant improvements in patient outcomes and, in recent years, combinations of anti-HER2 therapies have been explored and carry great promise. However, treatment resistance remains a problem. Resistance can be mediated, among others, by pathway redundancy, reactivation, or the utilization of escape pathways. Understanding mechanisms of resistance can lead to better therapeutic strategies to overcome resistance and optimize outcomes.

Abstract P4-15-13: CDK8 inhibition potentiates anti-ER and anti-HER2 therapies in breast cancer

Abstract CDK8, along with its paralog CDK19, is a cyclin dependent kinase which, in contrast to other members of the CDK family does not regulate cell cycle progression. CDK8 acts as a pleiotropic transcription regulator potentiating the induction of transcription by several transcription factors. Immunohistochemical staining of breast tissue arrays and bioinformatics analysis of gene expression microarray data of breast cancer patients revealed that CDK8 is overexpressed in breast cancer and that higher CDK8 expression correlates with the failure of systemic therapy. Small-molecule selective inhibitors of CDK8 and CDK19 (Senexin A and Senexin B) inhibited the mitogenic effects of estrogen and estrogen-dependent transcription in estrogen receptor (ER)+ breast cancer cell lines. CDK8/19 inhibitors had a cytostatic effect on different ER+ cell lines, and this growth inhibition was synergistic with the effect of the anti-estrogen fulvestrant, particularly in ER+ cell lines resistant to estrogen deprivation. Some of the ER+ cell lines sensitive to CDK8/19 inhibition also express HER2, and therefore we tested CDK8/19 inhibitors in combination with the HER2 and EGFR tyrosine kinase inhibitor lapatinib and an anti-HER2 monoclonal antibody, a biosimilar of trastuzumab. CDK8/19 inhibition produced a synergistic decrease in cell growth with both HER2 inhibitors; this effect was especially pronounced with a trastuzumab biosimilar. Surprisingly, the synergistic effect with HER2 inhibitors was observed in both ER+ HER2+ and ER-HER2+ cell lines, suggesting an effect on a HER2-complementing molecular target other than ER. Interestingly, CDK8/19 inhibition also synergized with trastuzumab biosimilar in a breast cancer cell line that exhibits innate resistance to trastuzumab, suggesting that CDK8/19 inhibition can overcome trastuzumab resistance in breast cancer. These results suggest that combining anti-estrogen and anti-CDK8 therapy may be more effective than conventional hormone therapy for ER positive breast cancer and that combining anti-HER2 and anti-CDK8 therapy is a rational potential treatment for HER2+ breast cancer, regardless of ER status or sensitivity to trastuzumab. Citation Format: Martina S McDermott, Chang-uk Lim, Mengqian Chen, Alexander Chumanevich, James F Catroppo, Balazs Gyorffy, David Oliver, Igor B Roninson, Eugenia V Broude. CDK8 inhibition potentiates anti-ER and anti-HER2 therapies in breast cancer [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P4-15-13.

Moderate level of HER2 expression and its prognostic significance in breast cancer with intermediate grade.

Overexpression of human epidermal growth factor receptor 2 (HER2) is an important prognostic and predictive marker of response to anti-HER2 therapy in breast cancer. Our goal was to analyze the prognostic significance of moderate expression of HER2 in breast cancer with intermediate differentiation grade. We performed a multicenter retrospective register study of 8494 patients with primary non-metastatic breast cancer admitted between 2000 and 2011 to eight Clinics in Saxony-Anhalt, federal state of Germany. Patients were divided into three groups according to their HER2 score: 4073 were classified as HER2 negative (HER2 0 and 1+), 822 HER2 moderate (HER2 2+/HER2), and 1238 HER2 positive (HER2 3+ or HER2 2+/HER2+). HER2-positive cases were excluded from analysis. Tumors with moderate HER2 (HER2 2+) expression demonstrated an aggressive behavior and worse patient survival compared with HER2 0 and 1+ status. HER2 2+ status was associated with shorter median overall survival (OS) (P < 0.0001) in breast cancer patients with an intermediate grade of differentiation. Comparing low-grade and high-grade tumors, HER2 moderate expression did not significantly influence patient survival. In multivariate analysis after adjustment for other prognostic factors HER2 2+ status remained an unfavorable prognostic factor for OS (HR 1.224, 95 % CI 1.059-1.415, P = 0.006) in breast cancer patients with an intermediate grade of differentiation. HER2 2+ status is an unfavorable prognostic factor regarding the OS of breast cancer patients with intermediate grade of differentiation and could be used to identify patients, who may benefit from adjuvant therapy.

Abstract 3331: CD24 promotes HER2 signaling pathways and CD24 inhibition sensitizes anti-HER2 therapy in breast cancer

Abstract Human epidermal growth factor receptor 2 (HER2)-overexpressing breast cancers are treated with HER2-targeted agents such as trastuzumab and lapatinib, which suppress HER2-initiated activation of the phosphatidylinositol 3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) cascades. CD24 is a glycosyl phosphatidylinositol (GPI)-anchored membrane protein and its overexpression is correlated with a poor prognosis in breast cancer patients. It has been reported that CD24 tends to be highly expressed in the HR-HER2+ breast cancer subgroup, which is negative for the hormone receptors (HRs) and positive for HER2. To determine the relationship between CD24 and HER2, we overexpressed HER2 in MDA-MB-231 breast cancer cells, which were negative for CD24 and HER2. We found that the expression of CD24 was increased by the overexpression of HER2. Flow cytometry analysis showed that the expression of CD24 was higher in the HER2-positive fraction than in the HER2-negative fraction. Conversely, in BT474 cells which express both CD24 and HER2, CD24 knockdown reduced the expression of HER2, while the knockdown of HER2 did not affect the expression of CD24. Furthermore, the knockdown of CD24 decreased the phosphorylation of Akt, which is known as a downstream molecule of PI3K and is related to cell survival. Accordingly, CD24 plays an important role in the regulation of the HER2-initiated signaling pathway. Based on these findings, we speculated that the expression of CD24 promotes a resistance to HER2-targeted therapies, leading to tumor progression in HER2-overexpressing cancers. We found that the knockdown of CD24 enhanced the sensitivity to lapatinib treatment. Consequently, we suggest using an anti-CD24 therapy to enhance the HER2-targeted therapy. Citation Format: Yoshimi Arima, Mari Hosonaga, Hideyuki Saya. CD24 promotes HER2 signaling pathways and CD24 inhibition sensitizes anti-HER2 therapy in breast cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3331. doi:10.1158/1538-7445.AM2014-3331

PRKACA mediates resistance to HER2-targeted therapy in breast cancer cells and restores anti-apoptotic signaling.

Targeting HER2 with antibodies or small molecule inhibitors in HER2-positive breast cancer leads to improved survival, but resistance is a common clinical problem. To uncover novel mechanisms of resistance to anti-HER2 therapy in breast cancer, we performed a kinase open reading frame (ORF) screen to identify genes that rescue HER2-amplified breast cancer cells from HER2 inhibition or suppression. In addition to multiple members of the MAPK and PI3K signaling pathways, we discovered that expression of the survival kinases PRKACA and PIM1 rescued cells from anti-HER2 therapy. Furthermore, we observed elevated PRKACA expression in trastuzumab-resistant breast cancer samples, indicating that this pathway is activated in breast cancers that are clinically resistant to trastuzumab-containing therapy. We found that neither PRKACA nor PIM1 restored MAPK or PI3K activation after lapatinib or trastuzumab treatment, but rather inactivated the pro-apoptotic protein BAD, thereby permitting survival signaling through BCL-XL. Pharmacological blockade of BCL-XL/BCL-2 partially abrogated the rescue effects conferred by PRKACA and PIM1, and sensitized cells to lapatinib treatment. These observations suggest that combined targeting of HER2 and the BCL-XL/BCL-2 anti-apoptotic pathway may increase responses to anti-HER2 therapy in breast cancer and decrease the emergence of resistant disease.