Epithelial Expressed B7-H4 Drives Differential Immunotherapy Response in Murine and Human Breast Cancer.

Combinations of immune checkpoint inhibitors (ICI, including anti-PD-1/PD-L1) and chemotherapy have been FDA approved for metastatic and early-stage triple-negative breast cancer (TNBC), but most patients do not benefit. B7-H4 is a B7 family ligand with proposed immunosuppressive functions being explored as a cancer immunotherapy target and may be associated with anti-PD-L1 resistance. However, little is known about its regulation and effect on immune cell function in breast cancers. We assessed murine and human breast cancer cells to identify regulation mechanisms of B7-H4 in vitro. We used an immunocompetent anti-PD-L1-sensitive orthotopic mammary cancer model and induced ectopic expression of B7-H4. We assessed therapy response and transcriptional changes at baseline and under treatment with anti-PD-L1. We observed B7-H4 was highly associated with epithelial cell status and transcription factors and found to be regulated by PI3K activity. EMT6 tumors with cell-surface B7-H4 expression were more resistant to immunotherapy. In addition, tumor-infiltrating immune cells had reduced immune activation signaling based on transcriptomic analysis. Paradoxically, in human breast cancer, B7-H4 expression was associated with survival benefit for patients with metastatic TNBC treated with carboplatin plus anti-PD-L1 and was associated with no change in response or survival for patients with early breast cancer receiving chemotherapy plus anti-PD-1. While B7-H4 induces tumor resistance to anti-PD-L1 in murine models, there are alternative mechanisms of signaling and function in human cancers. In addition, the strong correlation of B7-H4 to epithelial cell markers suggests a potential regulatory mechanism of B7-H4 independent of PD-L1. SIGNIFICANCE: This translational study confirms the association of B7-H4 expression with a cold immune microenvironment in breast cancer and offers preclinical studies demonstrating a potential role for B7-H4 in suppressing response to checkpoint therapy. However, analysis of two clinical trials with checkpoint inhibitors in the early and metastatic settings argue against B7-H4 as being a mechanism of clinical resistance to checkpoints, with clear implications for its candidacy as a therapeutic target.

An Unlikely Source of Iodine Uptake: A Bronchogenic Cyst Masquerading as Metastatic Thyroid Cancer.

Radioactive iodine therapy and posttreatment scanning are essential components of differentiated thyroid carcinoma treatment and detection of metastatic disease. False-positive results can be seen on an I-131 scan and are important for clinicians to be aware of. Here, we present a case of a 33-year-old female with follicular thyroid carcinoma who was noted to have an area of moderate uptake in the chest on a whole-body scan following remnant ablation with 30 mCi of I-131 (1.11GBq) concerning for a metastatic hilar lymph node. This was determined to be a mediastinal bronchogenic cyst on surgical pathology. It has been previously proposed that the expression of sodium iodide symporters in some bronchogenic cysts could be the mechanism by which iodine uptake is seen within them. We were able to demonstrate positive immunohistochemical staining for both sodium iodide symporter and the associated paired box gene 8 transcription factor in the cyst sample, which supports the proposed theory.

Endogenous pAKT activity is associated with response to AKT inhibition alone and in combination with immune checkpoint inhibition in murine models of TNBC.

Triple-negative breast cancer (TNBC) is a heterogeneous and challenging-to-treat breast cancer subtype. The clinical introduction of immune checkpoint inhibitors (ICI) for TNBC has had mixed results, and very few patients achieved a durable response. The PI3K/AKT pathway is frequently mutated in breast cancer. Given the important roles of the PI3K pathway in immune and tumor cell signaling, there is an interest in using inhibitors of this pathway to increase the response to ICI. This study sought to determine if AKT inhibition could enhance the response to ICI in murine TNBC models. We further sought to understand underlying mechanisms of response or non-response to AKT inhibition in combination with ICI. Using four murine TNBC-like cell lines and corresponding orthotopic mouse tumor models, we found that hyperactivity of the PI3K pathway, as evidenced by levels of phospho-AKT rather than PI3K pathway mutational status, was associated with response to AKT inhibition alone and in combination with ICI. Additional mutations in other growth regulatory pathways could override the response of PI3K pathway mutant tumors to AKT inhibition. Furthermore, we observed that AKT inhibition enhanced the response to ICI in an already sensitive model. However, AKT inhibition failed to convert ICI-resistant tumors, to responsive tumors. These findings suggest that analysis of both the mutational status and phospho-AKT protein levels may be beneficial in predicting which TNBC tumors will respond to AKT inhibition in combination with ICI.

AXL/WRNIP1 Mediates Replication Stress Response and Promotes Therapy Resistance and Metachronous Metastasis in HER2+ Breast Cancer.

Therapy resistance and metastatic progression are primary causes of cancer-related mortality. Disseminated tumor cells possess adaptive traits that enable them to reprogram their metabolism, maintain stemness, and resist cell death, facilitating their persistence to drive recurrence. The survival of disseminated tumor cells also depends on their ability to modulate replication stress in response to therapy while colonizing inhospitable microenvironments. In this study, we discovered that the nuclear translocation of AXL, a TAM receptor tyrosine kinase, and its interaction with WRNIP1, a DNA replication stress response factor, promotes the survival of HER2+ breast cancer cells that are resistant to HER2-targeted therapy and metastasize to the brain. In preclinical models, knocking down or pharmacologically inhibiting AXL or WRNIP1 attenuated protection of stalled replication forks. Furthermore, deficiency or inhibition of AXL and WRNIP1 also prolonged metastatic latency and delayed relapse. Together, these findings suggest that targeting the replication stress response, which is a shared adaptive mechanism in therapy-resistant and metastasis-initiating cells, could reduce metachronous metastasis and enhance the response to standard-of-care therapies. SIGNIFICANCE: Nuclear AXL and WRNIP1 interact and mediate replication stress response, promote therapy resistance, and support metastatic progression, indicating that targeting the AXL/WRNIP1 axis is a potentially viable therapeutic strategy for breast cancer.

Atezolizumab in Combination With Carboplatin and Survival Outcomes in Patients With Metastatic Triple-Negative Breast Cancer: The TBCRC 043 Phase 2 Randomized Clinical Trial.

Importance: Agents targeting programmed death ligand 1 (PD-L1) have demonstrated efficacy in triple-negative breast cancer (TNBC) when combined with chemotherapy and are now the standard of care in patients with PD-L1-positive metastatic disease. In contrast to microtubule-targeting agents, the effect of combining platinum compounds with programmed cell death 1 (PD-1)/PD-L1 immunotherapy has not been extensively determined. Objective: To evaluate the efficacy of atezolizumab with carboplatin in patients with metastatic TNBC. Design, Setting, and Participants: This phase 2 randomized clinical trial was conducted in 6 centers from August 2017 to June 2021. Interventions: Patients with metastatic TNBC were randomized to receive carboplatin area under the curve (AUC) 6 alone or with atezolizumab, 1200 mg, every 3 weeks until disease progression or unacceptable toxic effects with a 3-year duration of follow-up. Main Outcome and Measures: The primary end point was investigator-assessed progression-free survival (PFS). Secondary end points included overall response rate (ORR), clinical benefit rate (CBR), and overall survival (OS). Other objectives included correlation of response with tumor PD-L1 levels, tumor-infiltrating lymphocytes (TILs), tumor DNA- and RNA-sequenced biomarkers, TNBC subtyping, and multiplex analyses of immune markers. Results: All 106 patients with metastatic TNBC who were enrolled were female with a mean (range) age of 55 (27-79) years, of which 12 (19%) identified as African American/Black, 1 (1%) as Asian, 73 (69%) as White, and 11 (10%) as unknown. Patients were randomized and received either carboplatin (n = 50) or carboplatin and atezolizumab (n = 56). The combination improved PFS (hazard ratio [HR], 0.66; 95% CI, 0.44-1.01; P = .05) from a median of 2.2 to 4.1 months, increased ORR from 8.0% (95% CI, 3.2%-18.8%) to 30.4% (95% CI, 19.9%-43.3%), increased CBR at 6 months from 18.0% (95% CI, 9.8%-30.1%) to 37.5% (95% CI, 26.0%-50.6%), and improved OS (HR, 0.60; 95% CI, 0.37-0.96; P = .03) from a median of 8.6 to 12.6 months. Subgroup analysis showed PD-L1-positive tumors did not benefit more from adding atezolizumab (HR, 0.62; 95% CI, 0.23-1.65; P = .35). Patients with high TILs (HR, 0.12; 95% CI, 0.30-0.50), high mutation burden (HR, 0.50; 95% CI, 0.23-1.06), and prior chemotherapy (HR, 0.59; 95% CI, 0.36-0.95) received greater benefit on the combination. Patients with obesity and patients with more than 125 mg/dL on-treatment blood glucose levels were associated with better PFS (HR, 0.35; 95% CI, 0.10-1.80) on the combination. TNBC subtypes benefited from adding atezolizumab, except the luminal androgen receptor subtype. Conclusions and Relevance: In this randomized clinical trial, the addition of atezolizumab to carboplatin significantly improved survival of patients with metastatic TNBC regardless of PD-L1 status. Further, lower risk of disease progression was associated with increased TILs, higher mutation burden, obesity, and uncontrolled blood glucose levels. Trial Registration: ClinicalTrials.gov Identifier: NCT03206203.

NKG2A Is a Therapeutic Vulnerability in Immunotherapy Resistant MHC-I Heterogeneous Triple-Negative Breast Cancer.

Despite the success of immune checkpoint inhibition (ICI) in treating cancer, patients with triple-negative breast cancer (TNBC) often develop resistance to therapy, and the underlying mechanisms are unclear. MHC-I expression is essential for antigen presentation and T-cell-directed immunotherapy responses. This study demonstrates that TNBC patients display intratumor heterogeneity in regional MHC-I expression. In murine models, loss of MHC-I negates antitumor immunity and ICI response, whereas intratumor MHC-I heterogeneity leads to increased infiltration of natural killer (NK) cells in an IFNγ-dependent manner. Using spatial technologies, MHC-I heterogeneity is associated with clinical resistance to anti-programmed death (PD) L1 therapy and increased NK:T-cell ratios in human breast tumors. MHC-I heterogeneous tumors require NKG2A to suppress NK-cell function. Combining anti-NKG2A and anti-PD-L1 therapies restores complete response in heterogeneous MHC-I murine models, dependent on the presence of activated, tumor-infiltrating NK and CD8+ T cells. These results suggest that similar strategies may enhance patient benefit in clinical trials. SIGNIFICANCE: Clinical resistance to immunotherapy is common in breast cancer, and many patients will likely require combination therapy to maximize immunotherapeutic benefit. This study demonstrates that heterogeneous MHC-I expression drives resistance to anti-PD-L1 therapy and exposes NKG2A on NK cells as a target to overcome resistance. This article is featured in Selected Articles from This Issue, p. 201.

MYC Deregulation and PTEN Loss Model Tumor and Stromal Heterogeneity of Aggressive Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) patients have a poor prognosis and few treatment options. Mouse models of TNBC are important for development of new therapies, however, few mouse models represent the complexity of TNBC. Here, we develop a female TNBC murine model by mimicking two common TNBC mutations with high co-occurrence: amplification of the oncogene MYC and deletion of the tumor suppressor PTEN. This Myc;Ptenfl model develops heterogeneous triple-negative mammary tumors that display histological and molecular features commonly found in human TNBC. Our research involves deep molecular and spatial analyses on Myc;Ptenfl tumors including bulk and single-cell RNA-sequencing, and multiplex tissue-imaging. Through comparison with human TNBC, we demonstrate that this genetic mouse model develops mammary tumors with differential survival and therapeutic responses that closely resemble the inter- and intra-tumoral and microenvironmental heterogeneity of human TNBC, providing a pre-clinical tool for assessing the spectrum of patient TNBC biology and drug response.

Limiting mitochondrial plasticity by targeting DRP1 induces metabolic reprogramming and reduces breast cancer brain metastases.

Disseminated tumor cells with metabolic flexibility to utilize available nutrients in distal organs persist, but the precise mechanisms that facilitate metabolic adaptations remain unclear. Here we show fragmented mitochondrial puncta in latent brain metastatic (Lat) cells enable fatty acid oxidation (FAO) to sustain cellular bioenergetics and maintain redox homeostasis. Depleting the enriched dynamin-related protein 1 (DRP1) and limiting mitochondrial plasticity in Lat cells results in increased lipid droplet accumulation, impaired FAO and attenuated metastasis. Likewise, pharmacological inhibition of DRP1 using a small-molecule brain-permeable inhibitor attenuated metastatic burden in preclinical models. In agreement with these findings, increased phospho-DRP1 expression was observed in metachronous brain metastasis compared with patient-matched primary tumors. Overall, our findings reveal the pivotal role of mitochondrial plasticity in supporting the survival of Lat cells and highlight the therapeutic potential of targeting cellular plasticity programs in combination with tumor-specific alterations to prevent metastatic recurrences.

Rare Primary Fallopian Tube Grade 2 Neuroendocrine Tumor: A Case Report With Review of Literature.

Neuroendocrine neoplasms commonly arise from the gastrointestinal tract and lungs. Less commonly, they may occur in the gynecologic tract, typically within the ovary of a mature cystic teratoma. Primary neuroendocrine neoplasms of the fallopian tube are exceptionally rare and only a total of 11 cases have been reported in the literature. We describe the first case to our knowledge of a primary grade 2 neuroendocrine tumor of the fallopian tube in a 47-yr-old female. In this report, we describe the case's unique presentation, review the published literature on primary neuroendocrine neoplasms of the fallopian tube, discuss the treatment options, and speculate on their origin and histogenesis.

Robust biomarker discovery through multiplatform multiplex image analysis of breast cancer clinical cohorts.

Spatial profiling of tissues promises to elucidate tumor-microenvironment interactions and enable development of spatial biomarkers to predict patient response to immunotherapy and other therapeutics. However, spatial biomarker discovery is often carried out on a single patient cohort or imaging technology, limiting statistical power and increasing the likelihood of technical artifacts. In order to analyze multiple patient cohorts profiled on different platforms, we developed methods for comparative data analysis from three disparate multiplex imaging technologies: 1) cyclic immunofluorescence data we generated from 102 breast cancer patients with clinical follow-up, in addition to publicly available 2) imaging mass cytometry and 3) multiplex ion-beam imaging data. We demonstrate similar single-cell phenotyping results across breast cancer patient cohorts imaged with these three technologies and identify cellular abundance and proximity-based biomarkers with prognostic value across platforms. In multiple platforms, we identified lymphocyte infiltration as independently associated with longer survival in triple negative and high-proliferation breast tumors. Then, a comparison of nine spatial analysis methods revealed robust spatial biomarkers. In estrogen receptor-positive disease, quiescent stromal cells close to tumor were more abundant in good prognosis tumors while tumor neighborhoods of mixed fibroblast phenotypes were enriched in poor prognosis tumors. In triple-negative breast cancer (TNBC), macrophage proximity to tumor and B cell proximity to T cells were greater in good prognosis tumors, while tumor neighborhoods of vimentin-positive fibroblasts were enriched in poor prognosis tumors. We also tested previously published spatial biomarkers in our ensemble cohort, reproducing the positive prognostic value of isolated lymphocytes and lymphocyte occupancy and failing to reproduce the prognostic value of tumor-immune mixing score in TNBC. In conclusion, we demonstrate assembly of larger clinical cohorts from diverse platforms to aid in prognostic spatial biomarker identification and validation.

Multiomics in primary and metastatic breast tumors from the AURORA US network finds microenvironment and epigenetic drivers of metastasis.

The AURORA US Metastasis Project was established with the goal to identify molecular features associated with metastasis. We assayed 55 females with metastatic breast cancer (51 primary cancers and 102 metastases) by RNA sequencing, tumor/germline DNA exome and low-pass whole-genome sequencing and global DNA methylation microarrays. Expression subtype changes were observed in ~30% of samples and were coincident with DNA clonality shifts, especially involving HER2. Downregulation of estrogen receptor (ER)-mediated cell-cell adhesion genes through DNA methylation mechanisms was observed in metastases. Microenvironment differences varied according to tumor subtype; the ER+/luminal subtype had lower fibroblast and endothelial content, while triple-negative breast cancer/basal metastases showed a decrease in B and T cells. In 17% of metastases, DNA hypermethylation and/or focal deletions were identified near HLA-A and were associated with reduced expression and lower immune cell infiltrates, especially in brain and liver metastases. These findings could have implications for treating individuals with metastatic breast cancer with immune- and HER2-targeting therapies.

Obesity and metabolic syndrome are associated with short-term endocrine therapy resistance in early ER + breast cancer.

PURPOSE: Increased body mass index (BMI) and metabolic syndrome (MS) are associated with increased breast cancer recurrence risk. Whether this is due to intrinsic tumor biology or modifiable factors of the obese state remains incompletely understood. METHODS: Oncotype DX Recurrence Scores of 751 patients were stratified by BMI to assess association with tumor-intrinsic recurrence risk. Cellular proliferation by Ki67 after 10-21 days of presurgical letrozole treatment was used to stratify endocrine therapy response (sensitive-ln(Ki67) < 1; intermediate-ln(Ki67)1-2; resistant-ln(Ki67) > = 2). BMI at the time of surgery and MS variables were collected retrospectively for 143 patients to analyze association between therapy response and BMI/MS. Additionally, PI3K pathway signaling was evaluated by immunohistochemistry of phosphorylated Akt and S6. RESULTS: There was no significant association between BMI and recurrence score (p = 0.99), and risk score distribution was similar across BMI groups. However, BMI was associated with short-term endocrine therapy resistance, with a significant enrichment of intermediate and resistant tumors in patients with obesity (55%, p = 0.0392). Similarly, the relative risk of an endocrine therapy-resistant tumor was 1.4-fold greater for patients with MS (p = 0.0197). In evaluating PI3K pathway mediators, we found patients with 3 or more MS criteria had more tumors with pAkt scores above the median (p = 0.0436). There were no significant differences in S6 activation. CONCLUSION: Our findings suggest the association between obesity/metabolic syndrome and breast cancer recurrence is better reflected by response to treatment than tumor-intrinsic properties, suggesting interventions to reverse obesity and/or MS may improve outcomes for breast cancer recurrence.

Polycomb repressor complex 2 suppresses interferon-responsive MHC-II expression in melanoma cells and is associated with anti-PD-1 resistance.

BACKGROUND: Despite the remarkable success of immunotherapy in treating melanoma, understanding of the underlying mechanisms of resistance remains limited. Emerging evidence suggests that upregulation of tumor-specific major histocompatibility complex-II (tsMHC-II) serves as a predictive marker for the response to anti-programmed death-1 (PD-1)/programmed death ligand 1 (PD-L1) therapy in various cancer types. The genetic and epigenetic pathways modulating tsMHC-II expression remain incompletely characterized. Here, we provide evidence that polycomb repressive complex 2 (PRC2)/EZH2 signaling and resulting H3K27 hypermethylation suppresses tsMHC-II. METHODS: RNA sequencing data from tumor biopsies from patients with cutaneous melanoma treated with or without anti-PD-1, targeted inhibition assays, and assays for transposase-accessible chromatin with sequencing were used to observe the relationship between EZH2 inhibition and interferon (IFN)-γ inducibility within the MHC-II pathway. RESULTS: We find that increased EZH2 pathway messenger RNA (mRNA) expression correlates with reduced mRNA expression of both presentation and T-cell genes. Notably, targeted inhibition assays revealed that inhibition of EZH2 influences the expression dynamics and inducibility of the MHC-II pathway following IFN-γ stimulation. Additionally, our analysis of patients with metastatic melanoma revealed a significant inverse association between PRC2-related gene expression and response to anti-PD-1 therapy. CONCLUSIONS: Collectively, our findings demonstrate that EZH2 inhibition leads to enhanced MHC-II expression potentially resulting from improved chromatin accessibility at CIITA, the master regulator of MHC-II. These insights shed light on the molecular mechanisms involved in tsMHC-II suppression and highlight the potential of targeting EZH2 as a therapeutic strategy to improve immunotherapy efficacy.

Peripheral Blood Monocyte Abundance Predicts Outcomes in Patients with Breast Cancer.

Biomarkers of response are needed in breast cancer to stratify patients to appropriate therapies and avoid unnecessary toxicity. We used peripheral blood gene expression and cell type abundance to identify biomarkers of response and recurrence in neoadjuvant chemotherapy treated breast cancer patients. We identified a signature of interferon and complement response that was higher in the blood of patients with pathologic complete response. This signature was preferentially expressed by monocytes in single cell RNA sequencing. Monocytes are routinely measured clinically, enabling examination of clinically measured monocytes in multiple independent cohorts. We found that peripheral monocytes were higher in patients with good outcomes in four cohorts of breast cancer patients. Blood gene expression and cell type abundance biomarkers may be useful for prognostication in breast cancer. Significance: Biomarkers are needed in breast cancer to identify patients at risk for recurrence. Blood is an attractive site for biomarker identification due to the relative ease of longitudinal sampling. Our study suggests that blood-based gene expression and cell type abundance biomarkers may have clinical utility in breast cancer.

FGFR1 Antibody Validation and Characterization of FGFR1 Protein Expression in ER+ Breast Cancer.

Clinical trials in patients with ER+ breast cancer with or without FGFR pathway somatic alterations have shown limited clinical benefit from treatment with FGFR tyrosine kinase inhibitors alone or in combination with endocrine therapy. This is likely because of an inadequate predictive biomarker to select appropriate patients. In this study, we evaluated 4 anti-FGFR1 antibodies in breast cancer cell lines and patient-derived xenografts with FGFR1 amplification. We correlated D8E4 expression in 209 tumors from postmenopausal patients with stage I-III operable ER+ breast cancer with FGFR1 amplification status as determined by fluorescence in situ hybridization. FGFR1 amplification was identified in 10% of tumors (21/209), 80% of which exhibited membranous FGFR1 expression; however, only 50% of amplified cases showed strong, complete membranous staining (3+) based on established criteria to score HER2 by immunohistochemistry. These findings suggest the combined evaluation of FGFR1 status by immunohistochemistry and fluorescence in situ hybridization may need to be incorporated into the selection of patients for trials with FGFR inhibitors.

Combined Dusp4 and p53 loss with Dbf4 amplification drives tumorigenesis via cell cycle restriction and replication stress escape in breast cancer.

AIM: Deregulated signaling pathways are a hallmark feature of oncogenesis and driver of tumor progression. Dual specificity protein phosphatase 4 (DUSP4) is a critical negative regulator of the mitogen-activated protein kinase (MAPK) pathway and is often deleted or epigenetically silenced in tumors. DUSP4 alterations lead to hyperactivation of MAPK signaling in many cancers, including breast cancer, which often harbor mutations in cell cycle checkpoint genes, particularly in TP53. METHODS: Using a genetically engineered mouse model, we generated mammary-specific Dusp4-deleted primary epithelial cells to investigate the necessary conditions in which DUSP4 loss may drive breast cancer oncogenesis. RESULTS: We found that Dusp4 loss alone is insufficient in mediating tumorigenesis, but alternatively converges with loss in Trp53 and MYC amplification to induce tumorigenesis primarily through chromosome 5 amplification, which specifically upregulates Dbf4, a cell cycle gene that promotes cellular replication by mediating cell cycle checkpoint escape. CONCLUSIONS: This study identifies a novel mechanism for breast tumorigenesis implicating Dusp4 loss and p53 mutations in cellular acquisition of Dbf4 upregulation as a driver of cellular replication and cell cycle checkpoint escape.

Atypical Ductal Hyperplasia-Ductal Carcinoma In Situ Spectrum: Diagnostic Considerations and Treatment Impact in the Era of Deescalation.

As the first node in treatment algorithms for breast disease, pathologists have the potential to play a critical role in refining appropriate therapy for lesions in the atypical ducal hyperplasia-ductal carcinoma in situ (ADH-DCIS) spectrum by conservatively approaching diagnosis of lesions limited in size on core needle biopsy. Appropriate efforts to downgrade the diagnosis of lesions at the borderline of ADH and DCIS will certainly lead to more breast conservation and avoid the common morbidities of mastectomy, sentinel node biopsy, and radiation therapy. Whether results of clinical trials of active surveillance will successfully identify a subset of women who may successfully forgo even limited breast-conserving surgery is eagerly anticipated. Given the increasing concern that a significant number of women with DCIS are overtreated, identification of patients at very low risk for progression who may forgo surgery and radiation therapy safely is of significant interest.

Metabolic diversity within breast cancer brain-tropic cells determines metastatic fitness.

HER2+ breast cancer patients are presented with either synchronous (S-BM), latent (Lat), or metachronous (M-BM) brain metastases. However, the basis for disparate metastatic fitness among disseminated tumor cells of similar oncotype within a distal organ remains unknown. Here, employing brain metastatic models, we show that metabolic diversity and plasticity within brain-tropic cells determine metastatic fitness. Lactate secreted by aggressive metastatic cells or lactate supplementation to mice bearing Lat cells limits innate immunosurveillance and triggers overt metastasis. Attenuating lactate metabolism in S-BM impedes metastasis, while M-BM adapt and survive as residual disease. In contrast to S-BM, Lat and M-BM survive in equilibrium with innate immunosurveillance, oxidize glutamine, and maintain cellular redox homeostasis through the anionic amino acid transporter xCT. Moreover, xCT expression is significantly higher in matched M-BM brain metastatic samples compared to primary tumors from HER2+ breast cancer patients. Inhibiting xCT function attenuates residual disease and recurrence in these preclinical models.

Subacromial Bursal Tissue and Surrounding Matrix of Patients Undergoing Rotator Cuff Repair Contains Progenitor Cells.

PURPOSE: To build upon previous literature to identify a complete analysis of cellular contents of subacromial bursal tissue as well as the matrix surrounding the rotator cuff. METHODS: Samples of subacromial bursal tissue and surrounding matrix milieu from above the rotator cuff tendon and above the rotator cuff muscle bellies were obtained from 10 patients undergoing arthroscopic rotator cuff repair. Samples were analyzed using fluorescent-activated cell sorting and histologic analysis with staining protocols (Oil Red O, Alcian Blue, and Picro-Sirius Red), for identification of matrix components, including fat, proteoglycans, and collagen. RESULTS: Progenitor cells and fibroblast-type cells were present in significant amounts in subacromial bursal tissue in both tissues obtained from over the tendinous and muscle belly portions. Markers for neural tissue, myeloid cells, and megakaryocytes also were present to a lesser extent. There were prominent amounts of fat and proteoglycans present in the matrix, based on ImageJ analysis of stained histologic slides. CONCLUSIONS: The subacromial bursal tissue and surrounding matrix of patients undergoing rotator cuff repair contains progenitor cells in significant concentrations both over the tendon and muscle belly of the rotator cuff. CLINICAL RELEVANCE: This presence of progenitor cells, in particular, in the subacromial bursal tissue provides a potential basis for future applications of augmentation purposes in rotator cuff healing, and calls into question the practice of routine bursectomy. As the potential role of bursal tissue contents in growth and regeneration in the setting of rotator cuff healing is more well understood, maintaining this tissue may become more relevant. Concentration of these cellular components for use in autologous re-implantation is also an avenue of interest.

A Phase Ib Dose Escalation Trial of RO4929097 (a γ-secretase inhibitor) in Combination with Exemestane in Patients with ER + Metastatic Breast Cancer (MBC).

PRECLINICAL STUDIES: have demonstrated a complex cross-talk between Notch and estrogen signaling in ERα-positive breast cancer. Gamma-secretase inhibitors (GSIs) are investigational agents that block the cleavage and activation of Notch receptors. In animal models of endocrine-resistant breast cancer, combinations of tamoxifen and GSIs produce additive or synergistic efficacy, while decreasing the intestinal toxicity of GSIs. However, results of a clinical trial of a GSI-endocrine therapy combination in the metastatic setting have not been published to date, nor had the safety of such combinations been investigated with longer term treatment. We conducted a phase 1b dose escalation trial (NCT01149356) of GSI RO4929097 with exemestane in patients with ERα+, metastatic breast cancer (MBC) STUDY OBJECTIVES: To determine the safety, tolerability and maximum tolerated dose (MTD) or recommended phase 2 dose (RP2D) of RO4929097 when administered in combination with exemestane in patients with estrogen receptor positive metastatic breast cancer RESULTS: We enrolled 15 patients with MBC. Of 14 evaluable patients, one had a partial response, 6 had stable disease and 7 progressive disease. Twenty % of patients had stable disease for ≥ 6 months. Common toxicities included nausea (73.3%), anorexia (60%), hyperglycemia (53.3%), hypophosphatemia (46.7%), fatigue (66.7%) and cough (33.0%). Grade 3 toxicities were uncommon, and included hypophosphatemia (13%) and rash (6.3%). Rash was the only DLT observed at 140 mg/d. Results suggest a possible recommended phase 2 dose of 90 mg/d. Ten patients with evaluable archival tissue showed expression of PKCα, which correlated with expression of Notch4. Mammospheres from a PKCα-expressing, endocrine-resistant T47D cell line were inhibited by a GSI-fulvestrant combination CONCLUSIONS: Our data indicate that combinations including endocrine therapy and Notch inhibitors deserve further investigation in endocrine-resistant ERα-positive breast cancer.