Citations

Abstract Medulloblastoma arising from the cerebellum is the most common pediatric brain malignancy, with leptomeningeal metastases often present at diagnosis and recurrence associated with poor clinical outcome. In this study, we used mouse medulloblastoma models to explore the relationship of tumor pathophysiology and dysregulated expression of the NOTCH pathway transcription factor ATOH1, which is present in aggressive medulloblastoma subtypes driven by aberrant Sonic Hedgehog/Patched (SHH/PTCH) signaling. In experiments with conditional ATOH1 mouse mutants crossed to Ptch1+/− mice, which develop SHH-driven medulloblastoma, animals with Atoh1 transgene expression developed highly penetrant medulloblastoma at a young age with extensive leptomeningeal disease and metastasis to the spinal cord and brain, resembling xenografts of human SHH medulloblastoma. Metastatic tumors retained abnormal SHH signaling like tumor xenografts. Conversely, ATOH1 expression was detected consistently in recurrent and metastatic SHH medulloblastoma. Chromatin immunoprecipitation sequencing and gene expression profiling identified candidate ATOH1 targets in tumor cells involved in development and tumorigenesis. Among these targets specific to metastatic tumors, there was an enrichment in those implicated in extracellular matrix remodeling activity, cytoskeletal network and interaction with microenvironment, indicating a shift in transcriptomic and epigenomic landscapes during metastasis. Treatment with bone morphogenetic protein or SHH pathway inhibitors decreased tumor cell proliferation and suppressed metastatic tumor growth, respectively. Our work reveals a dynamic ATOH1-driven molecular cascade underlying medulloblastoma metastasis that offers possible therapeutic opportunities. Cancer Res; 77(14); 3766–77. ©2017 AACR.

Abstract Rationale The release of eosinophil granule proteins in the lungs of patients with asthma has been dogmatically linked with lung remodeling and airway hyperresponsiveness. However, the demonstrated inability of established mouse models to display the eosinophil degranulation occurring in human subjects has prevented a definitive in vivo test of this hypothesis. Objectives To demonstrate in vivo causative links between induced pulmonary histopathologies/lung dysfunction and eosinophil degranulation. Methods A transgenic mouse model of chronic T-helper cell type 2–driven inflammation overexpressing IL-5 from T cells and human eotaxin 2 in the lung (I5/hE2) was used to test the hypothesis that chronic histopathologies and the development of airway hyperresponsiveness occur as a consequence of extensive eosinophil degranulation in the lung parenchyma. Measurement and Main Results Studies targeting specific inflammatory pathways in I5/hE2 mice surprisingly showed that eosinophil-dependent immunoregulative events and not the release of individual secondary granule proteins are the central contributors to T-helper cell type 2–induced pulmonary remodeling and lung dysfunction. Specifically, our studies highlighted a significant role for eosinophil-dependent IL-13 expression. In contrast, extensive degranulation leading to the release of major basic protein-1 or eosinophil peroxidase was not causatively linked to many of the induced pulmonary histopathologies. However, these studies did define a previously unappreciated link between the release of eosinophil peroxidase (but not major basic protein-1) and observed levels of induced airway mucin. Conclusions These data suggest that improvements observed in patients with asthma responding to therapeutic strategies ablating eosinophils may occur as a consequence of targeting immunoregulatory mechanisms and not by simply eliminating the destructive activities of these purportedly end-stage effector cells.

Abstract Background. We previously reported an alternative ESR1 somatic gain-of-function chromosomal translocation event in a patient presenting with aggressive, endocrine therapy resistant estrogen receptor (ER) positive disease, producing an in-frame fusion gene consisting of N-terminal ESR1 and the C-terminus of the Hippo pathway coactivator YAP1 (ESR1-YAP1). We recently identified another ESR1 fusion through RNA sequencing (RNA-seq) in advanced stage ER+ disease from a chest wall recurrence in a male patient that was refractory to multiple lines of treatment. Two examples of fusions discovered in primary breast cancer samples include ESR1 fused in-frame to C-terminal sequences from NOP2 (ESR1-NOP2), identified in a resistant cohort from a RNA-seq screen focused on 81 primary breast cancers from aromatase inhibitor clinical trials, and a second ESR1 fusion, fused in-frame to the entire coding sequence of POLH (ESR1-POLH), that was identified from RNA-seq analysis of 728 Cancer Genome Atlas breast samples. This current study extends our previous characterization of ESR1-YAP1 by comparing functional and pharmacological properties of these three additional ESR1 gene fusion events of both early stage and advanced breast cancers. Methods. In vitro and in vivo experiments were conducted to test ESR1 fusions to induce therapeutic resistance, and metastasis. The transcriptional and binding properties of each fusion was also examined. Pharmacological inhibition with Palbociclib, a cyclin-dependent kinase 4/6 inhibitor, was utilized to assess drug sensitivity in ESR1 fusion containing breast cancer cells and in a patient derived xenograft (PDX) model expressing ESR1-YAP1 (WHIM18). Results. The YAP1 and PCDH11x fusions conferred estrogen-independent and fulvestrant-resistant growth. Immunohistochemistry revealed significantly higher numbers of ER+ cells in lungs of mice xenografted with T47D cells expressing the YAP1 and PCDH11x fusions compared to YFP control, NOP2 and POLH fusions. Results from ChIP-seq and microarray studies suggest that these two fusions promote proliferation and metastasis through genomic action by binding estrogen response elements (ERE) and subsequent gene activation. We thereby define these fusions as “canonical” fusions compared to “non-canonical” NOP2 and POLH fusions, which demonstrated dramatically decreased genomic binding ability. The non-canonical fusions induced genes associated with basal-like breast cancer and promoted HER2, EGFR, and MAPK gene expression signatures in contrast to genes associated with cell cycle/proliferation induced by canonical fusions. The proliferative ability of canonical fusion-containing ER+ cells was inhibited by Palbociclib in a dose-dependent manner. In vivo WHIM18 tumors in mice fed with Palbociclib-containing chow demonstrated significantly reduced tumor volume, growth rate, and weight compared to tumors in mice on control chow. Conclusions. In-frame ERE activating canonical fusions occur in end-stage drug resistant advanced breast cancer and can be added to ESR1 point mutations as a class of recurrent somatic mutation that may cause acquired resistance. Growth induced by these fusions can be antagonized by Palbociclib and is potentially clinically helpful. Citation Format: Lei JT, Shao J, Zhang J, Iglesia M, Cao J, Chan DW, He X, Kosaka Y, Schmidt C, Matsunuma R, Haricharan S, Crowder R, Hoog J, Phommaly C, Goncalves R, Ramalho S, Lai W-C, Hampton O, Rogers A, Tobias E, Parikh P, Davies S, Ma C, Suman V, Hunt K, Watson M, Hoadley KA, Thompson A, Chen X, Perou CM, Creighton CJ, Maher C, Ellis MJ. Recurrent functionally diverse in-frame ESR1 gene fusions drive endocrine resistance in 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 PD2-03.

98 Background: Concomitant with widespread adoption of axillary ultrasound (AUS) with ultrasound-guided needle biopsy (USNB) of suspicious lymph nodes (LN) for preoperative nodal staging of breast cancer patients, utilization of breast MRI, which includes axillary imaging, has increased. Little is known about the added value of MRI imaging of the axilla in this context. We undertook this study to assess the role of breast MRI in preoperative axillary nodal staging. Methods: We studied 988 consecutive invasive breast cancers in patients undergoing primary operation including axillary surgery, without neoadjuvant therapy, from 2010-2011. Results: 505 patients (51%) underwent MRI of which 168 (33%) demonstrated suspicious findings in the axilla. Abnormal axillary MRI findings included cortical thickening, edema, enhancement, hilar effacement, and/or altered shape and size. 114 patients had findings concordant with AUS. 54 patients had suspicious LNs on MRI either without a preceding AUS (33 cases) or after an initially negative AUS (21 cases). Second look AUS was performed in 29 of these cases and was abnormal in 3 (10%) in whom USNB confirmed metastatic adenopathy. Of the 54 cases with MRI-detected suspicious LNs 20 (37%) were node positive at operation with a pN stage of N0 (63%), N0i+ (5%), N1mic (4%), N1 (20%), N2 (6%), N3 (2%); extranodal extension was seen in 7 of 20 node-positive patients (35%). Conclusions: Second look AUS, when performed secondary to suspicious axillary MRI findings, identified LN metastasis preoperatively in 10% of patients. When MRI is done to evaluate the breast in newly diagnosed breast cancer patients, axillary findings can enhance the accuracy of preoperative nodal staging. We recommend second look AUS when MRI demonstrates suspicious axillary LN findings. [Table: see text]

49 Background: Predicting nodal positivity can guide surgical planning in breast cancer. Memorial Sloan-Kettering Cancer Center (MSKCC) and MD Anderson Cancer Center (MDACC) have established nomograms to predict sentinel node positivity. We propose that addition of distance of tumor from the nipple (DFN) can improve their performance. Methods: With IRB approval clinical T1/T2 tumors with pre-biopsy ultrasound were reviewed. MSKCC and MDACC nomogram predictions and the AUC-ROC were calculated. The addition of DFN was examined using multiple logistic regression and comparison of AUC-ROC values. Results: 401 breast cancers with clinical T1 (85%) or T2 (15%) tumors in 395 patients were included; of which 79/401 (19.7%) were node positive. Tumors were significantly closer to the nipple in those with positive nodes. 17/33 (51.5%) tumors within 2 cm of the nipple were node-positive versus 62/368 (16.8%) tumors > 2 cm from the nipple (p<0.0001). The MSKCC and MDACC nomograms each demonstrated good discrimination between node positive and negative patients with AUC-ROC values of 0.71 (95% CI: 0.64-0.77) and 0.74 (95% CI: 0.68-0.81), respectively. When added to the MSKCC nomogram, DFN ≤2 cm contributed significantly (OR 4.78, p=0.0001) to prediction of node positivity and improved the AUC-ROC to 0.73 (95% CI: 0.67-0.80). Similarly, DFN ≤2 cm was significant (OR 4.73, p=0.0002) when added to the MDACC nomogram and improved the AUC to 0.76 (95% CI: 0.70-0.82). Within nomogram probability categories, the proportion with positive nodes was consistently higher with DFN ≤2 cm. Conclusions: Tumor distance from nipple is associated with nodal positivity. When added to nomograms it improves prediction of node positivity. DFN should be considered when considering likelihood of nodal positivity for treatment planning. [Table: see text]