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For many patients with breast cancer, symptomatic bone metastases appear after years of latency. How micrometastatic lesions remain dormant and undetectable before initiating colonization is unclear. Here, we describe a mechanism involved in bone metastatic latency of oestrogen receptor-positive (ER+) breast cancer. Using an in vivo genome-wide short hairpin RNA screening, we identified the kinase MSK1 as an important regulator of metastatic dormancy in breast cancer. In patients with ER+ breast cancer, low MSK1 expression associates with early metastasis. We show that MSK1 downregulation impairs the differentiation of breast cancer cells, increasing their bone homing and growth capacities. MSK1 controls the expression of genes required for luminal cell differentiation, including the GATA3 and FOXA1 transcription factors, by modulating their promoter chromatin status. Our results indicate that MSK1 prevents metastatic progression of ER+ breast cancer, suggesting that stratifying patients with breast cancer as high or low risk for early relapse based on MSK1 expression could improve prognosis. Gawrzak et al. show that MSK1 regulates bone metastatic dormancy of ER+ breast cancer. MSK1 affects histone modifications at luminal transcription factor promoters to prevent cell differentiation and bone homing.

This consensus statement revises and updates the recommendations for biomarkers use in the diagnosis and treatment of breast cancer, and is a joint initiative of the Spanish Society of Medical Oncology and the Spanish Society of Pathology. This expert group recommends determining in all cases of breast cancer the histologic grade and the alpha-estrogen receptor (ER), progesterone receptor, Ki-67 and HER2 status, in order to assist prognosis and establish therapeutic options, including hormone therapy, chemotherapy and anti-HER2 therapy. One of the four available genetic prognostic platforms (MammaPrint®, Oncotype DX®, Prosigna® or EndoPredict®) may be used in node-negative ER-positive patients to establish a prognostic category and decide with the patient whether adjuvant treatment may be limited to hormonal therapy. Newer technologies including next-generation sequencing, liquid biopsy, tumour-infiltrating lymphocytes or PD-1 determination are at this point investigational.

Background: There is limited prospective data for advanced breast cancer (ABC) patients (pts), unresectable locally or metastatic, treated as per clinical practice. RegistEM, an academic study conducted by a non-profit collaborative group, will provide epidemiological, pathological and clinical data as per routine practice, including treatments for different disease settings, collected from pts’ medical records. Understanding the real distribution of BC subtypes is the primary objective. This is a non-interventional cohort study that will enroll approximately 1,867 pts with advanced disease diagnosed from January 2016 to December 2019, either after recurrence or as first diagnosis (de novo), in 38 Spanish sites representative of the whole country geography and whose investigators are GEICAM members. Biological samples (primary and/or metastatic tumor tissue, and blood) collection is part of study procedures. Methods: In this analysis we describe the characteristics of 229 pts with HR+/HER2- BC documented in a metastatic tumor lesion after disease recurrence and who received adjuvant endocrine therapy (ET). This subpopulation has been identify as an interesting group from a clinical perspective for detailed analysis. Three subgroups of pts have been considered for this analysis based on their disease-free interval (DFI) since the diagnosis of early disease: 50% pts recurring at ≥84 mo and only 1.3% pts at ≤12 mo. Most pts were Caucasian (99.1%) and female (99.6%), and at ABC diagnosis, 75.5% were postmenopausal and their median age was 59 years (33-88); median age was higher in pts with DFI ≥60 mo. Family history of BC and/or ovarian cancer was reported in 31.4% pts, with a slightly higher percentage in pts with DFI 50% pts recurred at ≥84 mo. Bone is the most frequent metastatic location in all DFI subgroups. The most common 1st- and 2nd-line therapies were the ET/BT combination, with AI/CDKi and FUL/CDKi, respectively. The median TTP to 1st-line therapy was superior in pts with DFI ≥60 mo. Citation Format: Isabel Alvarez, Angel Guerrero- Zotano, Sara López-Tarruella, Purificación Martínez, Marta Mori, Catalina Falo, Silvia Antolin, Cesar A. Rodríguez, Mireia Margeli, Isabel Garau, Ariadna Tibau, Diana Moreno, Josefina Cruz, M. Jose Echarri, Juan Lao, Alvaro Rodríguez-Lescure, Juan Jose Miralles, Susana Bezares, Federico Rojo, Carlos Jara. Characteristics of HR+/HER2- patients with recurrent disease from a prospective registry of unresectable locally advanced or metastatic breast cancer: GEICAM/2014-03 (RegistEM) [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS7-08.

The absence of established predictive markers with value to anticipate response to neoadjuvant 5-fluorouracil (5-FU)-based chemoradiotherapy (CRT) represents a current major challenge in locally advanced rectal cancer (LARC). The tumor suppressor microRNA (miR)-199b has been reported to play a key role determining 5-FU sensitivity of colorectal cancer cells through the regulation of several signaling pathways, and has emerged as a novel molecular target to overcome the 5-FU resistant phenotype. Moreover, miR-199b downregulation was described as a common alteration that predicts lack of response to preoperative CRT in LARC but this issue needs to be confirmed in independent larger cohorts. Here, we evaluate the clinical impact of miR-199b in LARC and perform additional analyses to further clarify its potential relevance as novel marker in this disease. Thus, miR-199b expression was quantified by real-time-PCR in a cohort of 185 LARC patients, observing this miR downregulated in 22.2% of cases and significantly associated with higher tumor size (p = 0.026) and positive lymph node after CRT (p = 0.005), and higher pathological stage (p = 0.004). Notably, this alteration showed a strong independent predictive value of poor pathological response to neoadjuvant CRT (p = 0.004). Moreover, the subgroup of cases with low miR-199b levels had a markedly shorter overall (p &lt, 001) and event-free survival (p &lt, 0.001), and multivariate analyses showed that miR-199b deregulation represents an independent prognosticator for patient outcome in LARC. Interestingly, the prognostic impact of this miR was strongly significant in both younger and elderly patients, and was very effective determining patient recurrence (p = 0.004). Finally, we compared miR-199b expression profiles in a set of cases with pre and post-treatment samples available, observing that only a minimal response leads to miR-199b increase levels, further suggesting its potential clinical and therapeutic relevance as a promising marker and novel molecular target for the management of LARC.

Introduction HER2+ correlates with aggressive phenotype and poor prognosis. The use of several antiHER2 drugs has dramatically improved prognosis and survival of HER2+ breast cancer (BC) patients. However, ≈20% of these patients will present resistance to antiHER2 therapies and relapse. The purpose of this study is to explore the role of AXL in antiHER2 treatment resistance and its potential as a druggable biomarker in HER2+ BC. Methods We generated 2 trastuzumab (T) resistant models from a HER2+ BC patient. The first one was a PDX derived resistant cell line established by chronic treatment for 6 months with T in vitro (TR1 and TR2). The second resistant model (TR4) was established by chronic treatment with T for 4 months and two serial passages in mice. The prognostic value of AXL was evaluated in primary tumor cohort of HER2+ BC patients treated by standard guidelines (n=51). AXL was also explored in a cohort from PAMELA trial, a neoadjuvant phase II study. Patients were given lapatinib and T for 18 weeks (Luminal-HER2+ patients were additionally given hormonotherapy). Tumor samples were collected at baseline, day 14 and surgery (n= 96). Results PDX resistant cell lines (TR1 and TR2) and resistant tumors (TR4) exhibit upregulation of AXL in comparison to the parental cells (PDX118). In vitro, the combination of an AXL inhibitor (TP-0903) + T resensitized resistant cells to T. In 3D organoids models from TR4, the combination decreased cell number in organoids more than both single agents. In TR4 in vivo model, TP-0903 + T completely abrogate tumor growth for more than 2 months.To validate our preclinical findings, we analyzed the level of AXL in our retrospectively cohort of patients. AXL level was significantly upregulated in patients who relapsed (p Conclusion AXL level was correlated with poor outcome of HER2+ BC patients. On our preclinical models, we showed that combination of TP-0903 and T abrogate tumor growth in acquired resistant models to T in PDXs. We postulate AXL as a promising new therapeutic strategy to overcome resistance to antiHER2 therapies Citation Format: Anna Adam-Artigues, Raimundo Cervera, Enrique Javier Arenas, Fara Brasó-Maristany, Alex Martinez-Sabadell, Eduardo Tormo, Cristina Hernando, Maria Teresa Martinez, Soraya Simon, Jesús Poveda, Octavio Burgués, Ana Rovira, Federico Rojo, Joan Albanell, Begoña Bermejo, Ana Lluch, Pilar Eroles, Aleix Prat, Joaquin Arribas, Juan Miguel Cejalvo. AXL is a potential druggable target in trastuzumab resistance in HER2+ breast cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1075.

e12539 Background: The Notch signaling pathway plays an important role in cell-differentiation, survival, proliferation, stem-cell renewal, and in determining cell fate during development and morphogenesis. The dysregulation of the Notch pathways contributes to carcinogenesis, cancer stem cell renewal, angiogenesis, and chemo-resistance. Elevated levels of Notch receptors and ligands have been associated with cancer-progression and poor survival. A less explored function of Notch pathways in cancer is their role in leukocyte homing and activation. Understanding their role and relationship with immune infiltrates is an area of interest in cancer research. Methods: The expression of four different Notch genes (Notch1, Notch2, Notch3 and Notch4) was explored in relation with A luminal breast cancer patient outcome using transcriptomic data (Affymetrix dataset, exploratory cohort) and the METABRIC study (validation cohort). The TIMER online tool was used to explore the association of the identified notch and immune infiltration, and the TCGA and METABRIC studies to analyze the correlation between notch1 - 4 expression and genomic signatures of immune activation. Results: We identified 2 individual genes called Notch1 and Notch2, which predict favorable prognosis in luminal A breast cancer. Their expression positively correlated with the presence of immune infiltrates within the tumor (dendritic cells, CD4+ T cells, neutrophils, CD8+ T cells and B cells), with markers of T cell activation and antigen presentation, and with gene signatures of immune surveillance (cytotoxic T lymphocyte activation and IFN gamma signature). By contrast Notch3 and Notch4 which predicted for detrimental outcome were not associated with any of these parameters. Conclusions: Our analysis identifies a Notch signature composed of 2 genes with potential to recognize immune infiltrated and activated A luminal phenotype breast cancers with favorable prognosis.

1083 Background: Triple-negative breast cancer (TNBC) is an aggressive breast cancer (BC) subtype comprising approximately 15% of BC. Conventional cytotoxic chemotherapies continue to be the mainstay for treatment of this BC, which lacks targetable markers. In this context, microRNAs have been described to have an important role. The aim of this work was to elucidate the function of miR-503-5p in doxorubicin resistance in TNBC. Methods: miR-503-5p expression was evaluated in the TNBC cell line with acquired resistance to doxorubicin (MDA-MB-231R) and its parental cell line (MDA-MB-231), by qRT-PCR. Studies of gain/loss of function of miR-503-5p were carried out in MDA-MB-231 and MDA-MB-231R cells by transient transfection of mimics and inhibitors. Cells were treated with doxorubicin, and viability was measured by flow cytometry and MTT assay. The role of miR-503-5p was also evaluated in vivo by Chicken Chorioallantoic Membrane (CAM) assay. MDA-MB-231 cells transfected with miR-503-5p mimic or scramble miRNA were inoculated onto the CAM of fertilized chicken eggs. After 48 hours, tumours were treated with doxorubicin or supplemented media for 48 hours and tumour growth was measured. miR-503-5p expression was quantified by qRT-PCR in a retrospective cohort of 74 TNBC patients treated with anthracycline + taxane regimens. Overall survival analysis for miR-503-5p in TNBC patients from METABRIC dataset was evaluated by the KM plotter online tool. Results: miR-503-5p was significantly upregulated in the resistant MDA-MB-231R TNBC cell line when compared to its parental cell line MDA-MB-231 (̃3.5-fold; p< 0.0001). Then, gain/loss function assays showed that upregulation of miR-503-5p in MDA-MB-231 cells increased resistance to doxorubicin ( p< 0.0001) and its downregulation in MDA-MB-231R cells had the opposite effect ( p< 0.0001). Moreover, the role of miR-503-5p was also confirmed in the CAM assay in vivo model, where miR-503-5p overexpression inhibited the effect of doxorubicin. In our cohort of patients, miR-503-5p expression levels in core biopsies sampled before preoperative chemotherapy were associated with residual cancer burden (RCB). miR-503-5p expression was significantly higher in patients with poor response to chemotherapy (RCB II and III; median, 95% CI: 0.00055, 0.00024 - 0.00136) than in patients with good response (RCB 0 and I; median, 95% CI: 0.00018, 0.00011 - 0.00034; p = 0.036). Moreover, we confirmed that TNBC patients with high expression of miR-503-5p had worse overall survival than patients with low expression ( p= 0.016). Conclusions: We identified miR-503-5p as a modulator of doxorubicin resistance in TNBC. Our in vitro findings are supported by the clinical data of TNBC patients and in vivo assays. Hence, the inhibition of miR-503-5p may be a promising strategy to improve chemotherapeutic efficacy. Moreover, the expression levels of miR-503-5p may be used as a biomarker for therapy response in TNBC.

519 Background: About one third of patients with hormone-receptor-positive (HR+), HER2‐ primary breast cancer with residual invasive disease after neoadjuvant chemotherapy will relapse despite adjuvant endocrine therapy. Therapeutic inhibition of cyclin-dependent kinase 4 and 6 (CDK 4/6) by palbociclib combined with endocrine therapy demonstrated highly relevant efficacy in metastatic breast cancer. The phase III PENELOPE-B (NCT01864746) study did not show a significant benefit from palbociclib in women with centrally confirmed HR+, HER2- primary breast cancer without a pathological complete response after taxane‐containing neoadjuvant chemotherapy and at high-risk of relapse (CPS‐EG score ≥3 or 2 and ypN+) for the primary endpoint (Loibl et al. JCO 2021). Methods: After completion of neoadjuvant chemotherapy and locoregional therapy, PENELOPE-B patients were randomized (1:1) to receive 13 cycles (1 year) of palbociclib 125mg daily or placebo on days 1-21 in a 28d cycle in addition to standard endocrine therapy. Analysis of the primary endpoint of invasive disease-free survival (iDFS) was planned after 290 events. Secondary objective included iDFS in luminal-B group by treatment. Gene expression in post-neoadjuvant surgical residual tumor tissue samples was profiled using the HTG EdgeSeq Oncology Biomarker Panel targeting 2559 genes (HTG Molecular Diagnostics Inc.). Based on 91 genes of this panel the AIMS subtype (Paquet & Hallett, JNCI 2014) was calculated. Results: Gene expressions were measured in tumors from 906 of 1250 (72%) PENELOPE-B patients; 663 had LumA subtype, 64 LumB, 135 NormL, 16 BasalL, and 28 HER2E. Compared to LumA the LumB patients were older, had higher post-neoadjuvant Ki-67, higher risk status (CPS-EG), and higher grade; no significant correlation was found for the region of participating sites, cT, ypT, and ypN. Patients with LumB tumors had an estimated 3-year iDFS of 71.9% with palbociclib vs 44.8% with placebo HR = 0.50 (0.24-1.05); outcome was similar in patients with LumA tumors (3-year iDFS 83.9% vs 79.5%, HR = 0.93 (0.68-1.28), interaction p = 0.132); this was confirmed in multivariable analyses. Ki-67 by IHC and proliferation biomarkers from the HTG panel also showed no significant interaction with treatment. Conclusions: PENELOPE-B did not show a benefit from the addition of 1 year palbociclib to endocrine therapy compared to placebo in the total enrolled high-risk primary breast cancer population. However, the small group of luminal-B tumors (n = 64) derived benefit from palbociclib, although without a statistically significant interaction. Further investigation is required in a larger cohort to validate a palbociclib benefit that might be confined to this group.

e12600 Background: In early-stage HER2+ breast cancer (BC), escalation or de-escalation of systemic treatment is an unmet need. Integration of promising biomarkers into risk scoring will further help progressing in the field. We aim to develop a prognostic signature that integrates two miRNAs (A and B) and quantitative and qualitative clinical variables in patients diagnosed with HER2+ BC. Methods: This study was conducted in a retrospective cohort of 45 HER2+ BC patients. Patients received standard treatment for localized disease. We calculated a prognostic signature for disease-free survival (DFS) using principal components analysis for mixed data combining clinicopathological data (Ki67 and axillary lymph node [pN0, pN1, pN2, pN3]) and expression of two microRNAs (we used mir-16 as housekeeping). Multiple DFS prognostic signatures were calculated and goodness of fit was evaluated by means of Akaike’s Information Criterion (AIC) to perform Cox model selection. Signature was then dichotomized into “high risk” and “low risk” using maximally selected Log-Rank statistics by Hothorn and Lausen, as method for optimal cut-off. Kaplan-Meier curves, Log-Rank test and Breslow test were used to ascertain statistical differences in the probability of DFS between high and low risk groups. MiRNA targeted genes were selected and used to perform functional enrichment analysis with the KEGG pathway database. To select significant terms/pathways, p-values were adjusted by the Benjamini-Hochberg method (p < 0.05). Results: MiR-A and miR-B expression was higher in primary tumor of patients who relapse compared to those free of disease after treatment (p = 0.018 and 0.004, respectively). Both miRNAs were strongly correlated (r = 0.84). This signature was significantly associated with relapse of the disease (HR 1.72; CI 95%: 1.243–2.382; p < 0.01, AIC = 114.02). The optimal cut-off of this score was obtained and patients were classified into high and low risk groups. Median DFS of the high-risk was 44 months while it has been not reached yet across the low risk after a median follow-up of 67 months (HR 8.39; p = 0.005, AIC = 111.784). Significant differences in survival between both groups were found (log rank test p < 0.001; Breslow test p = 0.002). miR-A and miR-B functional enrichment analysis returned 55 significant pathways. Interestingly, P53 pathway, apoptosis and cell cycle which are closely related to tumorigenesis and treatment response, were in the top 5 enriched pathways. Conclusions: Both miRNAs included in this signature are related to important biological pathways associated to BC progression. Our new prognostic signature identifies patients with early-stage, HER2+ BC who might be candidates for escalated or de-escalated systemic treatment. This signature was able to classify patients for DFS in high or low risk groups at the moment of BC diagnosis. Further investigations to validate the value of this new signature are on-going.

Background Nanoparticle albumin-bound paclitaxel (nab-Paclitaxel) is an alternative to standard taxanes for breast cancer (BC) treatment. We evaluated nab-Paclitaxel efficacy as neoadjuvant treatment for early estrogen receptor-positive (ER+), human epidermal growth factor receptor 2-negative (HER2-) disease. Materials and methods Women with ER+, HER2-, stage II-III BC were treated preoperatively with four cycles of weekly nab-Paclitaxel (150 mg/m2), 3 weeks on and 1 week off. We hypothesized that poor pathological response rate (residual cancer burden [RCB] III; Symmans criteria) would be ≤16%. Results Eighty-one patients with a median age of 47 years were treated; 64.2% were premenopausal, and 69% of tumors were stage II. Residual cancer burden III rate was 28.4% (95% confidence interval [CI]: 18.6%-38.2%), RCB 0+I (good response) rate was 24.7% (95% CI: 15.3%-34.1%) and RCB 0 (complete response) rate was 7.4% (95% CI: 1.7%-13.1%). Objective response rate by magnetic resonance imaging was 76.5% and rate of conversion to breast conserving surgery was 40.0%. The most frequent grade 3 and 4 toxicity was neutropenia (12.3% and 3.7% of patients, respectively), without any febrile neutropenia. Sensory neuropathy grade 2 and 3 were seen in 25.9% and 2.5% of patients, respectively. Tumor secreted protein, acidic, cysteine-rich (SPARC) overexpression was significantly associated with RCB 0 (odds ratio: 0.079; 95% CI: 0.009-0.689; p = .0216). Conclusion Despite failing to confirm an RCB III rate ≤16% in nab-Paclitaxel-treated patients, the RCB 0+I rate indicates a significant drug antitumor activity with low rates of grade 3-4 toxicity. Our exploratory biomarker analysis suggests a potential predictive role of complete response for SPARC. Confirmatory analyses are warranted, adapting dose and schedule to decrease peripheral neurotoxicity. (Trial registration: European Clinical Trials Database study number: 2011-004476-10; ClinicalTrials.gov: NCT01565499). Implications for practice The pathological response rate (residual cancer burden [RCB]; Symmans criteria) of nanoparticle albumin-bound paclitaxel administered as neoadjuvant treatment for early estrogen receptor-positive, human epidermal growth factor receptor 2-negative disease was evaluated. Whereas poor response (RCB III) was 24.7%, similar to that for docetaxel, good response (RCB 0+I) reached 23.0%, far superior to the 13% for docetaxel, while keeping toxicity low. Exploratory biomarker analysis suggests secreted protein, acidic, cysteine-rich overexpression in tumor cells as a potential predictor of complete response (RCB 0). Findings point to an encouraging single-agent neoadjuvant treatment with low toxicity, which warrants future research and development.

Caveolin-1 (CAV-1), the main scaffold protein in caveolae, is frequently deregulated in human cancer. Of importance, this protein has been described to show tumor suppressor or oncogenic properties depending on the cell type and the stage of the disease. In fact, its role in colorectal cancer (CRC) remains to be fully clarified due to discrepancies in the literature. We analyzed CAV-1 by western blot in a set of early-stage CRC patients with paired tumor tissue and normal colonic mucosa available. CAV-1 mRNA and expression levels of miR-124, 133 and 802 were quantified by real-time PCR. We found CAV-1 strongly downregulated in 76.2% of tumor samples and associated with the subgroup of elderly patients (p = 0.027). We observed by real-time PCR a lack of correlation between CAV-1 mRNA and protein levels in some cases with CAV-1 downregulated by western blot, and miR-124 deregulation was identified as a potential contributing alteration to decrease CAV-1 protein expression. CAV-1 is commonly downregulated in patients with primary CRC, which suggests its tumor suppressor role in early stages of this disease. Moreover, based on our findings, the previous discrepancies observed in different studies to date could be due to a complex posttranscriptional CAV-1 regulation.

Background: Meta-analysis of clinical trials has shown that adjuvant bisphosphonates reduce bone metastases and improve survival in postmenopausal (PM) pts1. However, we are unable to select pts most likely to benefit. To address this, the recently identified early breast cancer bone relapse biomarker, 16q23(MAF) gain (MAF+)2, was tested retrospectively in the large prospectively randomized AZURE trial3 of standard adjuvant therapy +/- zoledronic acid (ZOL) to determine the prognostic value of MAF and its potential to predict the effects of ZOL on disease outcomes. Materials and methods: All analyses were performed with ethics approval and consent. The biomarker analysis was completed on TMAs from primary tumors. Quadruplicate cores of breast tumor tissue were arrayed across replicate TMAs. MAF+ was detected using a validated (MAF/D16Z3) FISH test (Inbiomotion SL, Barcelona, Spain). A central laboratory (Targos, Kassel, Germany) validated the assay for analytic and diagnostic performance, established acceptance criteria, included appropriate quality controls for each assay, and performed the analyses in a blinded fashion. A copy number cut-off ≥2.5 was preset for MAF+. Invasive disease free (IDFS), overall (OS) survival and time to bone metastases multivariate analyses were performed in control and ZOL pts separately. Subsequently, interactions between MAF+ and effects of ZOL on disease outcomes by menopausal status were evaluated. Results: 1769 of the 3360 AZURE pts donated primary tumor samples. Median follow-up was 84 months. 865 pts (49%) had 2 FISH evaluable cores and were included in the analysis of which 184 (21%) had MAF+ tumors. Tumors that were MAF+ were more likely to be of higher grade, ER-ve and Her2+. In control pts, MAF was not prognostic for IDFS or OS although there were differences in IDFS by menopause (HR for MAF-/MAF+ in PM=0.47 [95%CI 0.25-0.88]; HR in non-PM=1.58 [0.82-3.03], test for interaction (TFI) by menopause P=0.007). In ZOL pts, MAF was prognostic for IDFS (HR=0.52 [0.36-0.75] and OS (HR=0.48 [0.31-0.75]). There were insufficient bone events (19 MAF+, 73 MAF-) in this sample set to reliably assess the impact of MAF+ on relapse in bone. In pts with MAF- tumors, ZOL was associated with improved IDFS (HR=0.74 [0.56-0.98]) and OS (HR=0.78 [0.55-1.10]). However, the effects of ZOL in MAF+ were profoundly influenced by menopausal status with possibly better outcomes in PM women (HR for IDFS=0.74 [0.35-1.58]) but clearly worse IDFS and OS outcomes in ZOL treated MAF+ pts who were non-PM (HR for IDFS 2.46 [1.23-4.92], TFI by treatment P=0.002 and HR for OS=2.27 [1.04-4.93], TFI by treatment P=0.032). The interactions between disease outcomes, ZOL use and menopause were driven largely by an association between MAF+ and an increased risk of extra-skeletal recurrence with the use of ZOL in women who were not PM. Conclusions: Absence of MAF amplification is associated with improved disease outcomes with adjuvant ZOL. However, the use of adjuvant ZOL in women with MAF+ tumors who are not PM at the start of treatment is associated with extraskeletal spread and worse DFS and OS. 1EBCTCG Lancet 2015;386:1353–1361; 2Pavlovic M et al JNCI 2015;107(12):djv256; 3Coleman RE et al Lancet Oncol 2014;15:997-1006. Citation Format: Coleman R, Hall A, Bell R, Cameron D, Marshall H, Jean-Mairet J, Tercero J, Rojo F, Albanell J, Gomis R. Impact of MAF gene amplification on disease recurrence and effects of adjuvant zoledronic acid in early breast cancer [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P1-09-01.

PURPOSE: Limited knowledge exists on the detection of breast cancer stem cell (BCSC)-related mutations in circulating free DNA (cfDNA) from patients with advanced cancers. Identification of new cancer biomarkers may allow for earlier detection of disease progression and treatment strategy modifications. METHODS: We conducted a prospective study to determine the feasibility and prognostic utility of droplet digital polymerase chain reaction (ddPCR)-based BCSC gene mutation analysis of cfDNA in patients with breast cancer. RESULTS: Detection of quantitative BCSC gene mutation in cfDNA by ddPCR mirrors disease progression and thus may represent a valuable and cost-effective measure of tumor burden. We have previously shown that hematological and neurological expressed 1-like (HN1L), ribosomal protein L39 (RPL39), and myeloid leukemia factor 2 (MLF2) are novel targets for BCSC self-renewal, and targeting these genetic alterations could be useful for personalized genomic-based therapy. CONCLUSION: BCSC mutation detection in cfDNA may have important implications for diagnosis, prognosis, and serial monitoring.

The mechanisms of chemotherapy resistance in triple negative breast cancer remain unclear, and so, new molecules which might mediate this resistance could optimize treatment response. Here we analyzed the involvement of the miRNA-449 family in the response to doxorubicin. The cell viability, cell-cycle phases, and the expression of in silico target genes and proteins of sensitive/resistant triple negative breast cancer cell lines were evaluated in response to doxorubicin treatment and after gain/loss of miRNAs-449 function achieved by transient transfection. Triple negative breast cancer patients were selected for ex vivo experiments and to evaluate gene and miRNAs expression changes after treatment, as well as survival analysis by Kaplan-Meier. Doxorubicin treatment upregulated miRNAs-449 and DNA-damage responder factors E2F1 and E2F3 in triple negative breast cancer sensitive breast cancer cells, while expression remained unaltered in resistant ones. In vitro overexpression of miRNAs-449 sensitized cells to the treatment and significantly reduced the resistance to doxorubicin. These changes showed also a strong effect on cell cycle regulation. Finally, elevated levels of miRNA-449a associated significantly with better survival in chemotherapy-treated triple negative breast cancer patients. These results reveal for the first time the involvement of the miRNA-449 family in doxorubicin resistance and their predictive and prognostic value in triple negative breast cancer patients.

SET nuclear proto-oncogene (SET) deregulation is a novel molecular target in metastatic colorectal cancer (CRC). However, its role in CRC progression and its potential clinical impact in early-stage CRC patients remain unknown. Here, we studied the biological effects of SET on migration using wound-healing and transwell assays, and anchorage-independent cell growth using soft agar colony formation assays after ectopic SET modulation. SET was analyzed by immuno-staining in 231 early-stage CRC patients, and miR-199b expression was quantified by real-time PCR in a set of CRC patients. Interestingly, SET enhances cell migration, markedly affects the colony-forming ability, promotes epithelial to mesenchymal transition, and induces the expression of the MYC proto-oncogene (c-MYC) in CRC cells. SET overexpression was detected in 15.4% of cases and was associated with worse Eastern Cooperative Oncology Group (ECOG) status (p = 0.021) and relapse in stage-II CRC patients (p = 0.008). Moreover, SET overexpression predicted shorter overall survival (p < 0.001) and time to metastasis (p < 0.001), and its prognostic value was particularly evident in elderly patients. MiR-199b downregulation was identified as a molecular mechanism to deregulate SET in patients with localized disease. In conclusion, SET overexpression is a common alteration in early-stage CRC, playing an oncogenic role associated with progression and aggressiveness, and portends a poor outcome. Thus, SET emerges as a novel potential molecular target with clinical impact in early-stage in CRC.

Purpose EndoPredict (EPclin) is a prognostic test validated to inform decisions on adjuvant chemotherapy to endocrine therapy alone for patients with oestrogen receptor-positive, HER2-negative breast cancer. Here, we determine the performance of EPclin for estimating 10-year distant recurrence-free interval (DRFI) rates for those who received adjuvant endocrine therapy (ET) alone compared to those with chemotherapy plus endocrine therapy (ET + C). Methods A total of 3746 women were included in this joint analysis. 2630 patients received 5 years of ET alone (ABCSG-6/8, TransATAC) and 1116 patients received ET + C (GEICAM 2003-02/9906). The primary objective was to evaluate the ability of EPclin to provide an estimate of the 10-year DR rate as a continuous function of EPclin separately for ET alone and ET + C. Cox proportional hazard models were used for these analyses. Results EPclin was highly prognostic for DR in women who received ET alone (HR 2.79 (2.49–3.13), P

PURPOSE RAS and BRAF mutations can be detected as a mechanism of acquired resistance in circulating tumor (ct) DNA in patients with metastatic colorectal cancer treated with anti–epidermal growth factor receptor therapy. METHODS RAS and BRAF mutational status was assessed in ctDNA in a baseline plasma sample and a serum sample collected at the time of the last available determination (named secondary extraction) from patients with KRAS exon 2 wild-type metastatic colorectal cancer treated in two first-line prospective biomarker-designed clinical trials (PULSE, ClinicalTrials.gov identifier: NCT01288339; and POSIBA, ClincialTrials.gov identifier: NCT01276379). RESULTS Analysis of extended RAS and BRAF in tissue and plasma from 178 patients with KRAS exon 2 wild-type metastatic colorectal cancer showed a sensitivity of 64.1% and a specificity of 90%. The median overall survival (OS) of baseline patients with RAS and BRAF mutations in ctDNA was 22.3 months (95% CI, 15.6 to 29 months) and 8.9 months (95% CI, 6.3 to 11.4 months), respectively, which was significantly inferior to the median OS of 40.4 months (95% CI, 35.9 to 44.9 months) in two patients with wild-type disease ( P < .001). Acquisition of RAS/BRAF mutations occurred in nine of 63 patients (14%) with progressive disease (PD; ie, blood draw within 1 month before PD or after PD) compared with six of 73 patients (8%) with no PD or blood extraction for ctDNA analysis before 1 month of PD ( P = .47). Median OS in patients with RAS/BRAF acquisition was 23.9 months (95% CI, 19.7 to 27.9 months) compared with 40.6 months (95% CI, not reached to not reached) in patients who remained free of mutations ( P = .016). CONCLUSION Our results confirm that baseline RAS and BRAF testing in ctDNA discriminates survival. The emergence of RAS/BRAF mutations has limited relevance for the time to progression to anti–epidermal growth factor receptor therapy.

The introduction of targeted treatments and more recently immune checkpoint inhibitors (ICI) to the treatment of metastatic non-small cell lung cancer (NSCLC) has dramatically changed the prognosis of selected patients. For patients with oncogene-addicted metastatic NSCLC harbouring an epidermal growth factor receptor (EGFR) or v-Raf murine sarcoma viral oncogene homologue B1 (BRAF) mutation or an anaplastic lymphoma kinase (ALK) or ROS proto-oncogene 1, receptor tyrosine kinase (ROS1) gene alteration (translocation, fusion, amplification) mutation-specific tyrosine kinase inhibitors (TKI) are already first-line standard treatment, while targeted treatment for other driver mutations affecting MET, RET, human epidermal growth factor receptor (HER) 2, tropomyosin receptor kinases (TRK) 1–3 and others are currently under investigation. The role of ICI in these patient subgroups is currently under debate. This article summarises a round-table discussion organised by ESMO Open in Vienna in July 2018. It reviews current clinical data on ICI treatment in patients with metastatic oncogene-addicted NSCLC and discusses molecular diagnostic assessment, potential biomarkers and radiological methods for response evaluation of ICI treatment. The round-table panel concluded ICI should only be considered in patients with oncogene-addicted NSCLC after exhaustion of effective targeted therapies and in some cases possibly after all other therapies including chemotherapies. More clinical trials on combination therapies and biomarkers for ICI therapy based on the specific differing characteristics of oncogene-addicted NSCLC need to be conducted.

Advanced bladder cancer is associated with a poor prognosis and limited treatment options. The PI3K/AKT/mTOR pathway is frequently activated in this disease and can be a potential therapeutic target for treatment intervention. We studied the antitumor efficacy of a new targeted therapy, TAK-228 (oral mTORC1/2 inhibitor), in preclinical models of bladder cancer. We evaluated the effects of TAK-228 in combination with a PI3Kα inhibitor (TAK-117) or with chemotherapy (paclitaxel). We used six bladder cancer cell lines and in vivo xenografts models. TAK-228 strongly inhibited cell proliferation in vitro, and reduced tumor growth and angiogenesis in vivo. Three possible biomarkers of response to TAK-228 (basal levels of 4E-BP1, p-4E-BP1/4E-BP1 ratio, or eIF4E/4E-BP1 ratio) were identified. The combination of TAK-228 and TAK-117 had synergistic effects in vitro and in vivo. Furthermore, TAK-228 demonstrated greater efficiency when combined with paclitaxel. TAK-228 also showed ex vivo activity in tumor tissue from patients with treatment-naïve bladder cancer. TAK-228 is a promising investigational agent that induces a strong effect on cell proliferation, tumor growth, and angiogenesis in bladder cancer models. High synergistic effects were observed with TAK-228 combined with a PI3K inhibitor or with chemotherapy. These results are currently being investigated in a clinic trial of TAK-228 plus paclitaxel in patients with metastatic bladder cancer (NCT03745911). Implications: Strong synergistic effects were observed when combining TAK-228 with TAK-117 (a PI3Kα inhibitor) or with paclitaxel chemotherapy. A phase II study at our institution is currently evaluating the efficacy of TAK-228 combined with paclitaxel in patients with metastatic bladder cancer.

Background: Changes in the clinical subtype (CS) and intrinsic subtype (IS) between breast cancer (BC) metastases and corresponding primary tumours have been reported. However, their relationship with tumour genomic changes remains poorly characterised. Here, we analysed the association between genomic remodelling and subtype conversion in paired primary and metastatic BC samples. Methods: A total of 57 paired primary and metastatic tumours from GEICAM/2009-03 (ConvertHER, NCT01377363) study participants with centrally assessed CS (n = 57) and IS (n = 46) were analysed. Targeted capture and next-generation sequencing of 202 genes on formalin-fixed paraffin-embedded samples was performed. The cancer cell fraction (CCF) of mutations in primary and metastatic pairs was estimated as a surrogate of tumour clonal architecture. Changes in mutation CCF between matched primary and metastatic tumours were analysed in the presence or absence of subtype conversion. Findings: CS conversion occurred in 24.6% and IS conversion occurred in 36.9% of metastases. Primary tumours and metastases had a median of 11 (range, 3-29) and 9 (range, 1-38) mutations, respectively (P = 0.05). Overall, mutations in metastases showed a higher estimated CCF than in primary tumours (median CCF, 0.51 and 0.47, respectively; P = 0.042), consistent with increased clonal homogeneity. The increase in mutation CCF was significant in CS-converted (P = 0.04) but not in IS-converted (P = 0.48) metastases. Clonal remodelling was highest in metastases from hormone receptorepositive and human epidermal growth factor 2 (HER2)-positive tumours (P = 0.006). Conclusions(1): Mutations in BC metastases showed significantly higher estimated CCF than primary tumours. CCF changes were more prominent in metastases with CS conversion. Our findings suggest that changes in BC subtypes are linked to clonal remodelling during BC evolution. (C) 2019 Elsevier Ltd. All rights reserved.

Natural killer (NK) cells can orchestrate effective antitumor immunity. The presence of tumor-infiltrating NK cells in diagnostic biopsies predicts pathologic complete response (pCR) to HER2-specific therapeutic antibodies in patients with primary breast cancer. Here, we analyzed whether diversity in circulating NK cells might influence tumor infiltration and HER2-specific therapeutic antibody efficacy. We found that numbers of circulating CD57+ NK cells inversely correlated with pCR to HER2-specific antibody treatment in patients with primary breast cancer independently of age, traditional clinicopathologic factors, and CD16A 158F/V genotype. This association was uncoupled from the expression of other NK-cell receptors, the presence of adaptive NK cells, or changes in major T-cell subsets, reminiscent of cytomegalovirus-induced immunomodulation. NK-cell activation against trastuzumab-coated HER2+ breast cancer cells was comparable in patients with high and low proportions of CD57+ NK cells. However, circulating CD57+ NK cells displayed decreased CXCR3 expression and CD16A-induced IL2-dependent proliferation in vitro. Presence of CD57+ NK cells was reduced in breast tumor–associated infiltrates as compared with paired peripheral blood samples, suggesting deficient homing, proliferation, and/or survival of NK cells in the tumor niche. Indeed, numbers of circulating CD57+ were inversely related to tumor-infiltrating NK-cell numbers. Our data reveal that NK-cell differentiation influences their antitumor potential and that CD57+ NK cells may be a biomarker useful for tailoring HER2 antibody–based therapeutic strategies in breast cancer.

[EN] BACKGROUND: Liquid biopsy offers a minimally invasive alternative to tissue-based evaluation of mutational status in cancer. The goal of the present study was to evaluate the aggregate performance of OncoBEAM RAS mutation analysis in plasma of colorectal cancer (CRC) patients at 10 hospital laboratories in Spain where this technology is routinely implemented. METHODS: Circulating cell-free DNA from plasma was examined for RAS mutations using the OncoBEAM platform at each hospital laboratory. Results were then compared to those obtained from DNA extracted from tumour tissue from the same patient. RESULTS: The overall percentage agreement between plasma-based and tissue-based RAS mutation testing of the 236 participants was 89% (210/236; kappa, 0.770 (95% CI: 0.689-0.852)). Re-analysis of tissue from all discordant cases by BEAMing revealed two false negative and five false positive tumour tissue RAS results, with a final concordance of 92%. Plasma false negative results were found more frequently in patients with exclusive lung metastatic disease. CONCLUSIONS: In this first prospective real-world RAS mutation performance comparison study, a high overall agreement was observed between results obtained from plasma and tissue samples. Overall, these findings indicate that the plasma-based BEAMing assay is a viable solution for rapid delivery of RAS mutation status to determine mCRC patient eligibility for anti-EGFR therapy., This work was supported by Sysmex Inostics, Inc as well as the AES Programme [Grant Numbers PI15/00934, PT17/0015/0006]. The funders had no involvement in the writing of this manuscript but did review for medical accuracy prior to submission.

Natural killer (NK) cells can orchestrate effective antitumor immunity. The presence of tumor-infiltrating NK cells in diagnostic biopsies predicts pathologic complete response (pCR) to HER2-specific therapeutic antibodies in patients with primary breast cancer. Here, we analyzed whether diversity in circulating NK cells might influence tumor infiltration and HER2-specific therapeutic antibody efficacy. We found that numbers of circulating CD57

[Background]: An important proportion of HER2-positive metastatic breast cancer patients do not respond to trastuzumab. The combination of dasatinib and trastuzumab has shown to be synergistic in preclinical models., [Methods]: We conducted a phase II trial combining dasatinib 100 mg once daily with trastuzumab 2 mg/kg and paclitaxel 80 mg/m2 weekly. Primary objective was objective response rate (ORR) and secondary included safety, other efficacy parameters and pharmacodynamics in tumour tissue, blood samples and skin biopsies., [Results]: From June 2013 to December 2015, 29 patients were included. Median number of cycles was 12 (1–49). Only 6 patients discontinued due to adverse events. ORR was 79.3% (95% CI 60.3–92), clinical benefit rate 82.8% (95% CI 64.2–94.2). Median time to progression 23.9 months (95% CI 14.9–not reached [NR]), median progression-free survival 23.9 months (95% CI 10.3–NR). No grade 4 toxicity was seen. Grade 3 toxicities included: ejection fraction decrease, neutropenia, hyponatremia, fatigue and sensory neuropathy and one left ventricular systolic dysfunction. Phosphorylated (p)-SRC was reduced in peripheral blood mononuclear cells. Phosphorylated SRC, ERK and AKT were also reduced in epidermal keratinocytes., [Conclusions]: Dasatinib can be safely combined with trastuzumab and paclitaxel. The combination is active with an ORR of almost 80%. Trial registration: NCT01306942, EudraCT 2010-023304-27., The study was financially supported by Bristol-Myers Squibb which also supplied the dasatinib.

Background: NF-κB signalling appears deregulated in breast tumours. The purpose of this study was to determine whether the non-canonical NF-κB pathway, is activated in oestrogen receptor positive (ER+) breast cancer, to identify any correlation between its activity and the clinico-pathological phenotype and to explore whether NF-κB2 and RelB subunits and/or any of their target genes might be used as a predictive marker. Methods: Two independent cohorts of ER+ early breast cancer patients treated with adjuvant endocrine therapy were included in the study. Activation of RelB and NF-κB2 subunits was determined in a training set of 121 patients by measuring DNA-binding activities in nuclear extracts from fresh frozen specimens by an ELISA-based assay. Samples of 15 ER− breast cancer patients were also included in the study. In a large validation cohort of 207 patients, nuclear immunostaining of RelB and NF-κB2 on formalin-fixed paraffin-embedded specimens was performed. Statistical correlation within clinico-pathological factors, disease-free survival (DFS) and overall survival (OS) was evaluated. Publicly available gene expression and survival data have been interrogated aimed to identify target genes. Results: Activation of NF-κB2 and RelB was found in 53.7 and 49.2% of the 121 ER+ tumours analysed, with similar levels to ER− breast tumours analysed in parallel for comparisons. In the validation cohort, we obtained a similar proportion of cases with activation of NF-κB2 and RelB (59.9 and 32.4%), with a 39.6% of co-activation. Multiplexing immunofluorescence in breast cancer tissue confirmed an inverse spatial distribution of ER with NF-κB2 and RelB nuclear expression in tumour cells. Interestingly, NF-κB2 and RelB mRNA expression was inversely correlated with ER gene (ESR1) levels (P

This work has been carried out with the support of the RNA-Reg. CONSOLIDER-Consortium (CSD2009-00080) of the Spanish Ministry of Economy and Competitiveness., Aguilera, O., Muñoz-Sagastibelza, M., Torrejón, B., Borrero-Palacios, A., del Puerto-Nevado, L., Martínez-Useros, J., Rodriguez-Remirez, M., Zazo, S., García, E., Fraga, M., Rojo, F., García-Foncillas, J.

Aberrant activation of the Wnt/β-catenin pathway and polo-like kinase 1 (Plk1) overexpression represent two common events in prostate cancer with relevant functional implications. This minireview analyzes their potential therapeutic significance in prostate cancer based on their role as androgen receptor (AR) signaling regulators and the pivotal role of the tumor suppressor protein phosphatase 2A (PP2A) modulating these pathways.

Introduction: The role of bevacizumab in combination with chemotherapy in metastatic BC is controversial, and no biomarker exists as of today that predicts benefit to this agent. In the LEA clinical trial, a numerical, statistically non-significant benefit from the addition of bevacizumab to endocrine therapy (ET) was observed in the first-line metastatic setting (18.4 vs. 13.8 months of Progression-Free Survival (PFS), p=0.14). Here, we explored various gene expression-based predictors of outcome and benefit to bevacizumab. Methods and materials: LEA trial randomized 380 patients with HR+/HER2- advanced disease to bevacizumab in combination with ET (ET+B) vs. ET alone. Primary endpoint was PFS. Expression of BC selected genes was evaluated in formalin-fixed paraffin-embedded (FFPE) primary tumors using the nCounter platform from patients randomized in Spain that consent for biomarker analyses. The following variables were evaluated: 1) research-based PAM50 intrinsic subtypes (categorical variable; Luminal A, Luminal B, HER2-enriched, Basal-like and Normal-like); 2) research-based PAM50 signatures (continuous variable; scores showing the distant of the gene expression values of an individual sample compared to the centroid gene values for each PAM50 intrinsic subtype); 3) risk of recurrence (ROR) groups (low, medium and high); 4) the 13-gene hypoxia/VEGF signature (continuous); and 5) Ki67 by immunohistochemistry (continuous). Uni- and multivariable Cox models for PFS were used to test the prognostic significance of each variable. To determine whether each variable is predictive of bevacizumab benefit, we tested the interaction term of each variable by treatment arm in a Cox model. Results: Tumor samples from 103 patients were analyzed: 55 (53%) in ET+B arm and 49 (47%) in ET arm. Subtype distribution was as follows: 57 (55.3%) Luminal A, 32 (31.1%) Luminal B, 5 (4.9%) HER2-enriched, 1 (1.0%) Basal-like, and 8 (7.8%) normal-like. In a univariate analysis, Luminal B tumors had a poorer outcome using Luminal A as reference (13.8 vs. 21.3 months, respectively; (hazard ratio, HR=1.80, 95% CI 1.10-2.95, p=0.019). Concordant with this finding, Luminal A signature was associated with a better outcome. Similarly, ROR-P high group showed a poorer outcome than ROR-P low group (8.5 vs. 19.4 months; HR=2.88, 95% CI 1.30-6.35, p=0.009). Neither VEGF-13 signature nor Ki67 were found to be associated with PFS. Similar findings were obtained after adjustment for treatment, age, previous ET, ECOG, visceral disease and number of metastatic sites. In terms of treatment benefit, the HER2-enriched signature was the only variable found predictive of bevacizumab PFS benefit in univariate (p=0.010) and multivariate (p=0.015) analyses. Conclusions: In advanced HR+/HER2- disease, intrinsic subtype (i.e. Luminal A vs. B) independently predicts PFS following first-line ET. In addition, HR+/HER2-negative tumors with high expression of the HER2-enriched signature, a biomarker of estrogen-independence, benefit the most from bevacizumab. Further validation of these prognostic and predictive biomarkers is warranted. Citation Format: Prat A, de la Haba-Rodriguez J, Guerrero Á, García-Sáenz JA, Morales S, Antón A, Muñoz M, Ramos M, Martínez-Jáñez N, Margelí M, Servitja S, Rojo F, Galván P, González S, Cruz J, Sánchez-Rovira P, Perelló A, Rodríguez-Martin C, Casas M, Carrasco E, Caballero R, Martín M. Predicting outcome and benefit to first-line bevacizumab in advanced/metastatic hormone receptor (HR)+/HER2-negative breast cancer (BC) treated with endocrine therapy: A correlative science study from the LEA phase III clinical trial (GEICAM/2006-11_GBG 051). [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 P3-07-42.

e21555 Background: Lung-ONE study aims to determine the clinical utility in making decisions on planned 2nd line therapy in advanced NSCLC based on a comprehensive genomic profile (CGP) test as FoundationOne CDx or FoundationOne Liquid and describe alterations found on these patients. Methods: Lung-ONE included advanced/metastatic NSCLC patients being treated on first-line, with molecular diagnostic wild-type (or unknown) for at least ALK, EGFR, and ROS-1 genes. Patients treated with targeted therapies or immunotherapy were excluded. This interim analysis describes the findings of CGP from patients of Lung-ONE with driver genomic alterations (GAs) and the investigators decision to change the planned second line treatment or not. Patients continue being followed-up for up to 2 years. Results: As of November 15 2019, 152 patients were included. At the time of sampling, 90.7% had received ≥2 different chemotherapy regimens and other therapies. Main testing for ALK and ROS-1 was IHC in 91.4% and 73.8%, respectively; and for EGFR, it was qPCR in 99.3%. Planned 2nd-line based on routine molecular profile was chemotherapy and, mainly checkpoint inhibitors (61.7%). CGP was assayed on 102 (67.1%) tissue specimens and 50 (32.9%) blood samples. Of the 1,581 GAs identified in 141 patients, 541(34.2%) were potential drivers in 131. The most frequently mutated genes were TP53 in 72(55%) patients and KRAS in 52(40%). TMB and MSI signatures were calculated on 82 patients. 21% had < = 16 mut/Mb. Other GAs of interest are presented in table. Therapeutic orientations were recommended for 116/152(76.8%) patients. In 39/151(25.8%) patients the GAs had approved drugs in NSCLC or other tumor types. CGP outcomes, would be used by 27% of oncologists to guide targeted 2nd-line. Conclusions: CGP detected mutations that are informative for choosing personalized treatment in 25%. CGP demonstrates the ability of detect druggable genomic alterations missed at diagnosis opening new treatment options. [Table: see text]

Background: Breast cancer (BC) is a heterogeneous disease. HER2+ BC represents between 15-20% of cases. Trastuzumab, a monoclonal antibody, has been successfully improved clinical benefits in both localized and advanced setting. Despite this evidence, many patients experience resistance to therapy. The objective of this study is to analyze AXL receptor as a potential mechanism of resistance and its implication as a prognostic factor. Methods: We used three breast cancer cell lines with acquired resistance to Trastuzumab. Resistant models were generated by treating parental cells (AU565, SKR3, BT474) with constant dose of Trastuzumab (15μg/mL) for 6 months. Cell viability was estimated by MTT assay. Proteins were assessed by Western blot and genes by qRT-PCR. AXL was downregulated by siRNA and a selective tyrosine kinase (TK) AXL inhibitor (TP-0903). Proteins interaction was assessed by immunoprecipitation and Duolink®PLA. The prognostic value of AXL was evaluated in primary tumor in a cohort of HER2+ BC patients treated with Trastuzumab plus chemotherapy from our center (n=33). Results: Acquired resistant cell lines (RCLs) maintained HER2 amplification. RCLs were more proliferative than sensitive parental cell lines. There was an important up-regulation of AXL (>2.5 fold-change) and epithelial-mesenchymal transition markers (VIM, CDH2, CTNNB1 and FN1) in RCLs (p Citation Format: Anna Adam-Artigues, Eduardo Tormo, Raimundo Cervera, Federico Rojo, Alejandro Pérez-Fidalgo, Sandra Zazo, Paula González-Alonso, Cristina Hernándo, María Teresa Martínez, Valentina Gambardella, Jesús Poveda, Soraya Simón, Belén Ortega, Santiago Moragón, Ana Rovira, Joan Albanell, Octavio Burgués, Begoña Bermejo, Pilar Eroles, Ana Lluch, Juan Miguel Cejalvo. AXL-HER2 dimer as mechanism of anti-HER2 acquired resistance in HER2 amplified brest cancer models: A new step towards precision medicine [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P6-03-05.

Cetuximab is a standard-of-care treatment for RAS wild-type metastatic colorectal cancer (mCRC) but not for those harbor a KRAS mutation since MAPK pathway is constitutively activated. Nevertheless, cetuximab also exerts its effect by its immunomodulatory activity despite the presence of RAS mutation. The aim of this study was to determine the impact of polymorphism FcγRIIIa V158F and killer immunoglobulin-like receptor (KIR) genes on the outcome of mCRC patients with KRAS mutations treated with cetuximab. This multicenter Phase II clinical trial included 70 mCRC patients with KRAS mutated. We found KIR2DS4 gene was significantly associated with OS (HR 2.27; 95% CI, 1.08–4.77; P = 0.03). In non-functional receptor homozygotes the median OS was 2.6 months longer than in carriers of one copy of full receptor. Multivariate analysis confirmed KIR2DS4 as a favorable prognostic marker for OS (HR 6.71) in mCRC patients with KRAS mutation treated with cetuximab. These data support the potential therapeutic of cetuximab in KRAS mutated mCRC carrying non-functional receptor KIR2DS4 since these patients significantly prolong their OS even after heavily treatment. KIR2DS4 typing could be used as predictive marker for identifying RAS mutated patients that could benefit from combination approaches of anti-EGFR monoclonal antibodies and other immunotherapies to overcome the resistance mediated by mutation in RAS., This clinical trial was approved and supported by Merck S.L., an affiliate of Merck KGaA, Darmstadt. Germany [research project number 2010-023580-18, date: 05-06-2014]

We investigated the value of tumor-infiltrating NK (TI-NK) cells and HLA class I tumor expression as biomarkers of response to neoadjuvant anti-HER2 antibody-based treatment in breast cancer.TI-NK cells and HLA-I were determined by IHC in pretreatment tumor biopsies from two cohorts of patients with HER2-positive breast cancer [discovery cohort (TI-NK cells were significantly associated with pCR in the discovery cohort as well as in the validation cohort (This study identifies baseline TI-NK cells as an independent biomarker with great predictive value for pCR to anti-HER2 antibody-based treatment and points to the complementary value of tumor HLA-I status for defining patient prognosis independently of pCR.

Pathology archives with linked clinical data are an invaluable resource for translational research, with the limitation that most cancer samples are formalin-fixed paraffin-embedded (FFPE) tissues. Therefore, FFPE tissues are an important resource for genomic profiling studies but are under-utilised due to the low amount and quality of extracted nucleic acids. We profiled the copy number landscape of 356 breast cancer patients using DNA extracted FFPE tissues by shallow whole genome sequencing. We generated a total of 491 sequencing libraries from 2 kits and obtained data from 98.4% of libraries with 86.4% being of good quality. We generated libraries from as low as 3.8ng of input DNA and found that the success was independent of input DNA amount and quality, processing site and age of the fixed tissues. Since copy number alterations (CNA) play a major role in breast cancer, it is imperative that we are able to use FFPE archives and we have shown in this study that sWGS is a robust method to do such profiling.

Fumonisins are mycotoxins responsible for equine leukoencephalomalacia (ELEM) and they are produced by Fusarium species of high incidence in corn crops. In Jalisco (Mexico), six cases were confirmed by histopathological and neurological findings and fumonisins analysis. This is the first report of fumonisins in corn stover associated to ELEM cases.

This consensus statement revises and updates the recommendations for biomarkers use in the diagnosis and treatment of breast cancer, and is a joint initiative of the Spanish Society of Medical Oncology and the Spanish Society of Pathology. This expert group recommends determining in all cases of breast cancer the histologic grade and the alpha-estrogen receptor (ER), progesterone receptor, Ki-67 and HER2 status, in order to assist prognosis and establish therapeutic options, including hormone therapy, chemotherapy and anti-HER2 therapy. One of the four available genetic prognostic platforms (MammaPrint®, Oncotype DX®, Prosigna® or EndoPredict®) may be used in node-negative ER-positive patients to establish a prognostic category and decide with the patient whether adjuvant treatment may be limited to hormonal therapy. Newer technologies including next-generation sequencing, liquid biopsy, tumour-infiltrating lymphocytes or PD-1 determination are at this point investigational.

Treatment of colorectal cancer (CRC) with monoclonal antibodies against epidermal growth factor receptor requires the assessment of the mutational status of exons 2, 3, and 4 of the NRAS and KRAS oncogenes. Moreover, the mutational status of exon 15 of the BRAF oncogene is a marker of poor prognosis in CRC. The Idylla NRAS-BRAF Mutation Test is a reliable, simple (

INTRODUCTION: The coexpression of pIGF-1R and MMP-7 (double-positive phenotype, DP) correlates with poor overall survival (OS) in KRAS wild-type (WT) (exon 2) metastatic colorectal cancer (mCRC) patients treated with irinotecan-cetuximab in second/third line. METHODS: We analyzed two prospective biomarker design trials of newly diagnosed RAS-WT mCRC patients treated with panitumumab-FOLFOX6 (PULSE trial; NCT01288339) or cetuximab plus either FOLFOX6/FOLFIRI (POSIBA trial; NCT01276379). The main exposure was DP phenotype (DP/ non-DP), as assessed by two independent pathologists. DP cases were defined by immunohistochemistry as N70% expression of moderate or strong intensity for both MMP-7 and pIGF-1R. Primary endpoint: progression-free survival (PFS); secondary endpoints: OS and response rate. PFS and OS were adjusted by baseline characteristics using multivariate Cox models. RESULTS: We analyzed 67 patients (30 non-DP, 37 DP) in the PULSE trial and 181 patients in the POSIBA trial (158 non-DP, 23 DP). Response rates and PFS were similar between groups in both studies. DP was associated with prolonged OS in PULSE (adjusted HR: 0.23; 95% CI: 0.11-0.52; P =. 0004) and with shorter OS in POSIBA (adjusted HR: 1 .67; 95% CI: 0.96-2.90; P =. 07). CONCLUSION: A differential effect of anti-EGFRs on survival by DP phenotype was observed. Panitumumab might be more beneficial for RAS-WT mCRC patients with DP phenotype, whereas cetuximab might improve OS in non-DP., Amgen supported the PULSE trial and Merck supported the POSIBA trial. Neither Amgen nor Merck had any role in the present analysis design, analysis and interpretation of data, writing the report, and the decision to submit the report for publication.

The cellular and molecular basis of stromal cell recruitment, activation and crosstalk in carcinomas is poorly understood, limiting the development of targeted anti-stromal therapies. In mouse models of triple negative breast cancer (TNBC), Hedgehog ligand produced by neoplastic cells reprograms cancer-associated fibroblasts (CAFs) to provide a supportive niche for the acquisition of a chemo-resistant, cancer stem cell (CSC) phenotype via FGF5 expression and production of fibrillar collagen. Stromal treatment of patient-derived xenografts with smoothened inhibitors (SMOi) downregulates CSC markers expression and sensitizes tumors to docetaxel, leading to markedly improved survival and reduced metastatic burden. In the phase I clinical trial EDALINE, 3 of 12 patients with metastatic TNBC derived clinical benefit from combination therapy with the SMOi Sonidegib and docetaxel chemotherapy, with one patient experiencing a complete response. These studies identify Hedgehog signaling to CAFs as a novel mediator of CSC plasticity and an exciting new therapeutic target in TNBC., Stromal cell recruitment, activation and crosstalk with cancer cells is poorly understood. Here, the authors demonstrate that cancer cell-derived Hedgehog ligand triggers stromal remodeling that in turn induces a cancer-stem-cell like, drug-resistant phenotype of nearby cancer cells while treatment with smoothened inhibitors reverses these phenotypes.