Surgical Delay-Associated Mortality Risk Varies by Subtype in Loco-Regional Breast Cancer Patients in SEER-Medicare.

Substantial evidence supports that delay of surgery after breast cancer diagnosis is associated with increased mortality risk, leading to the introduction of a new Commission on Cancer quality measure for receipt of surgery within 60 days of diagnosis for non-neoadjuvant patients. Breast cancer subtype is a critical prognostic factor and determines treatment options; however, it remains unknown whether surgical delay-associated breast cancer-specific mortality (BCSM) risk differs by subtype. This retrospective cohort study aimed to assess whether the impact of delayed surgery on survival varies by subtype (hormone [HR]+/HER2-, HR-/HER2-, and HER2+) in patients with loco-regional breast cancer who received surgery as their first treatment between 2010-2017 using the SEER-Medicare. Continuous time to surgery from diagnostic biopsy (TTS; days) in reference to TTS = 30 days. BCSM were evaluated as flexibly dependent on continuous time (days) to surgery from diagnosis (TTS) using Cox proportional hazards and Fine and Gray competing-risk regression models, respectively, by HR status. Inverse propensity score-weighting was used to adjust for demographic, clinical, and treatment variables impacting TTS. Adjusted BCSM risk grew with increasing TTS across all subtypes, however, the pattern and extent of the association varied. HR+/HER2- patients exhibited the most pronounced increase in BCSM risk associated with TTS, with approximately exponential growth after 42 days, with adjusted subdistribution hazard ratios (sHR) of 1.21 (95% CI: 1.06-1.37) at TTS = 60 days, 1.79 (95% CI: 1.40-2.29) at TTS = 90 days, and 2.83 (95% CI: 1.76-4.55) at TTS = 120 days. In contrast, both HER2 + and HR-/HER2- patients showed slower, approximately linear growth in sHR, although non-significant in HR-HER2-.

Needle biopsy accelerates pro-metastatic changes and systemic dissemination in breast cancer: Implications for mortality by surgery delay.

Increased breast cancer (BC) mortality risk posed by delayed surgical resection of tumor after diagnosis is a growing concern, yet the underlying mechanisms remain unknown. Our cohort analyses of early-stage BC patients reveal the emergence of a significantly rising mortality risk when the biopsy-to-surgery interval was extended beyond 53 days. Additionally, histology of post-biopsy tumors shows prolonged retention of a metastasis-permissive wound stroma dominated by M2-like macrophages capable of promoting cancer cell epithelial-to-mesenchymal transition and angiogenesis. We show that needle biopsy promotes systemic dissemination of cancer cells through a mechanism of sustained activation of the COX-2/PGE2/EP2 feedforward loop, which favors M2 polarization and its associated pro-metastatic changes but are abrogated by oral treatment with COX-2 or EP2 inhibitors in estrogen-receptor-positive (ER+) syngeneic mouse tumor models. Therefore, we conclude that needle biopsy of ER+ BC provokes progressive pro-metastatic changes, which may explain the mortality risk posed by surgery delay after diagnosis.

Increased breast cancer mortality due to treatment delay and needle biopsy type: a retrospective analysis of SEER-medicare.

BACKGROUND: Substantial evidence indicates that delay of first treatment after diagnosis is associated with poorer survival outcomes in breast cancer. Accordingly, the Commission on Cancer introduced a quality measure for receipt of therapeutic surgery within 60 days of diagnostic biopsy for stage I-III breast cancer patients in the non-neoadjuvant setting. It is unknown, however, what may contribute to mortality associated with treatment delay. Therefore, we investigated whether biopsy type moderates the effect of the mortality risk posed by treatment delay. METHODS: Retrospective analysis of 31,306 women with stage I-III breast cancer diagnosed between 2003 and 2013 selected from the SEER-Medicare database was performed to determine whether needle biopsy type [core needle biopsy (CNB) or vacuum-assisted biopsy (VAB)] impacts time to treatment (TTT)-associated survival outcomes. Multivariable Fine-Gray competing risk survival models, adjusted for inverse propensity score weights, were used to determine the association between biopsy type, TTT, and breast cancer-specific mortality (BCSM). RESULTS: TTT ≥ 60 days was associated with 45% higher risk of BCSM (sHR = 1.45, 95% CI 1.24-1.69) compared to those with TTT < 60 days in stage I-III cases. Independent of TTT, CNB was associated with 28% higher risk of BCSM compared to VAB in stage II-III cases (sHR = 1.28, 95% CI 1.11-1.36), translating to a 2.7% and 4.0% absolute difference in BCSM at 5 and 10 years, respectively. However, in stage I cases, the BCSM risk was not associated with type of biopsy. CONCLUSIONS: Our results suggest that treatment delay ≥ 60 days is independently associated with poorer survival outcomes in breast cancer patients. In stage II-III, CNB is associated with higher BCSM than VAB. However, type of biopsy does not underlie TTT-associated breast cancer mortality risk.

EWI2 and its relatives in Tetraspanin-enriched membrane domains regulate malignancy.

Experimental studies on immunoglobulin superfamily (IgSF) member EWI2 reveal that it suppresses a variety of solid malignant tumors including brain, lung, skin, and prostate cancers in animal models and inhibits tumor cell movement and growth in vitro. While EWI2 appears to support myeloid leukemia in mouse models and maintain leukemia stem cells. Bioinformatics analyses suggest that EWI2 gene expression is downregulated in glioblastoma but upregulated in melanoma, pancreatic cancer, and liver cancer. The mechanism of action for EWI2 is linked to its inhibition of growth factor receptors and cell adhesion proteins through its associated tetraspanin-enriched membrane domains (TEMDs), by altering the cell surface clustering and endolysosome trafficking/turnover of these transmembrane proteins. Recent studies also show that EWI2 modulates the nuclear translocation of ERK and TFEB to change the activities of these gene expression regulators. For EWI2 relatives including FPRP, IgSF3, and CD101, although their roles in malignant diseases are not fully clear and remain to be determined experimentally, FPRP and IgSF3 likely promote the progression of solid malignant tumors while CD101 seems to modulate immune cells of tumor microenvironment. Distinctive from other tumor regulators, the impacts of EWI subfamily members on solid malignant tumors are likely to be context dependent. In other words, the effect of a given EWI subfamily member on a tumor probably depends on the molecular network and composition of TEMDs in that tumor. Collectively, EWI2 and its relatives are emerged as important regulators of malignant diseases with promising potentials to become anti-cancer therapeutics and cancer therapy targets.

Sustained Inflammation of Breast Tumors after Needle Biopsy.

INTRODUCTION: Needle biopsy is essential for definitive diagnosis of breast malignancy. Significant histologic changes due to tissue damage have been reported in solid tumors. This study investigated the association between time from needle biopsy and inflammation in breast tumors. METHODS: A total of 73 stage I-II invasive breast cancer cases diagnosed by image-guided needle biopsy who had surgery as their first definitive treatment were retrospectively analyzed. Time from biopsy to surgical excision ranged from 8 to 252 days. Histological sections of surgically resected tumors with a visible needle tract were reviewed by histologic evaluation. Data were analyzed by McNemar's test for proportional differences, and the Benjamini-Hochberg procedure was used to assess the association between immune cell prevalence and clinical variables. RESULTS: Characteristic histology changes, including foreign body giant-cell reaction, synovial-cell metaplasia, desmoplastic repair changes, granulation tissue, fat necrosis, and inflammation, were frequently detected adjacent to the needle tract. Spatial comparison indicated that a higher proportion of cases had neutrophils, eosinophils, and macrophages adjacent to the needle tract than tumors distant from it. The presence of inflammatory cells adjacent to the needle tract was not associated with time from biopsy or subtype. Still, plasma cells were associated with residual carrier material from biopsy markers. CONCLUSION: Macrophages and eosinophils are highly abundant and retained adjacent to the needle tract regardless of time from the biopsy.

EWI2 prevents EGFR from clustering and endocytosis to reduce tumor cell movement and proliferation.

EWI2 is a transmembrane immunoglobulin superfamily (IgSF) protein that physically associates with tetraspanins and integrins. It inhibits cancer cells by influencing the interactions among membrane molecules including the tetraspanins and integrins. The present study revealed that, upon EWI2 silencing or ablation, the elevated movement and proliferation of cancer cells in vitro and increased cancer metastatic potential and malignancy in vivo are associated with (i) increases in clustering, endocytosis, and then activation of EGFR and (ii) enhancement of Erk MAP kinase signaling. These changes in signaling make cancer cells (i) undergo partial epithelial-to-mesenchymal (EMT) for more tumor progression and (ii) proliferate faster for better tumor formation. Inhibition of EGFR or Erk kinase can abrogate the cancer cell phenotypes resulting from EWI2 removal. Thus, to inhibit cancer cells, EWI2 prevents EGFR from clustering and endocytosis to restrain its activation and signaling.

Spatial Metrics of Interaction between CD163-Positive Macrophages and Cancer Cells and Progression-Free Survival in Chemo-Treated Breast Cancer.

Tumor-associated macrophages (TAMs) promote progression of breast cancer and other solid malignancies via immunosuppressive, pro-angiogenic and pro-metastatic effects. Tumor-promoting TAMs tend to express M2-like macrophage markers, including CD163. Histopathological assessments suggest that the density of CD163-positive TAMs within the tumor microenvironment is associated with reduced efficacy of chemotherapy and unfavorable prognosis. However, previous analyses have required research-oriented pathologists to visually enumerate CD163+ TAMs, which is both laborious and subjective and hampers clinical implementation. Objective, operator-independent image analysis methods to quantify TAM-associated information are needed. In addition, since M2-like TAMs exert local effects on cancer cells through direct juxtacrine cell-to-cell interactions, paracrine signaling, and metabolic factors, we hypothesized that spatial metrics of adjacency of M2-like TAMs to breast cancer cells will have further information value. Immunofluorescence histo-cytometry of CD163+ TAMs was performed retrospectively on tumor microarrays of 443 cases of invasive breast cancer from patients who subsequently received adjuvant chemotherapy. An objective and automated algorithm was developed to phenotype CD163+ TAMs and calculate their density within the tumor stroma and derive several spatial metrics of interaction with cancer cells. Shorter progression-free survival was associated with a high density of CD163+ TAMs, shorter median cancer-to-CD163+ nearest neighbor distance, and a high number of either directly adjacent CD163+ TAMs (within juxtacrine proximity <12 µm to cancer cells) or communicating CD163+ TAMs (within paracrine communication distance <250 µm to cancer cells) after multivariable adjustment for clinical and pathological risk factors and correction for optimistic bias due to dichotomization.

Prolonged Time from Diagnosis to Breast-Conserving Surgery is Associated with Upstaging in Hormone Receptor-Positive Invasive Ductal Breast Carcinoma.

BACKGROUND: Time to surgery (TTS) has been suggested to have an association with mortality in early-stage breast cancer. OBJECTIVE: This study aims to determine the association between TTS and preoperative disease progression in tumor size or nodal status among women diagnosed with clinical T1N0M0 ductal breast cancer. METHODS: Women diagnosed with clinical T1N0M0 ductal breast cancer who had breast-conserving surgery as their first definitive treatment between 2010 and 2016 (n = 90,405) were analyzed using the National Cancer Database. Separate multivariable logistic regression models for hormone receptor (HR)-positive and HR-negative patients, adjusted for clinical and demographic variables, were used to assess the relationship between TTS and upstaging of tumor size (T-upstaging) or nodal status (N-upstaging). RESULTS: T-upstaging occurred in 6.76% of HR-positive patients and 11.00% of HR-negative patients, while N-upstaging occurred in 12.69% and 10.75% of HR-positive and HR-negative patients, respectively. Among HR-positive patients, odds of T-upstaging were higher for 61-90 days TTS (odds ratio [OR] 1.18, 95% confidence interval [CI] 1.05-1.34) and ≥91 days TTS (OR 1.47, 95% CI 1.17-1.84) compared with ≤30 days TTS, and odds of N- upstaging were higher for ≥91 days TTS (OR 1.35, 95% CI 1.13-1.62). No association between TTS and either T- or N-upstaging was found among HR-negative patients. Other clinical and demographic variables, including grade, tumor location, and race/ethnicity, were associated with both T- and N-upstaging. CONCLUSION: TTS ≥61 and ≥91 days was a significant predictor of T- and N-upstaging, respectively, in HR-positive patients; however, TTS was not associated with upstaging in HR-negative breast cancer. Delays in surgery may contribute to measurable disease progression in T1N0M0 ductal breast cancer.

Functional Blockade of E-Selectin in Tumor-Associated Vessels Enhances Anti-Tumor Effect of Doxorubicin in Breast Cancer.

Chemotherapy is a mainstay of treatment for solid tumors. However, little is known about how therapy-induced immune cell infiltration may affect therapy response. We found substantial CD45+ immune cell density adjacent to E-selectin expressing inflamed vessels in doxorubicin (DOX)-treated residual human breast tumors. While CD45 level was significantly elevated in DOX-treated wildtype mice, it remained unchanged in DOX-treated tumors from E-selectin null mice. Similarly, intravenous administration of anti-E-selectin aptamer (ESTA) resulted in a significant reduction in CD45+ immune cell density in DOX-treated residual tumors, which coincided with a delay in tumor growth and lung metastasis in MMTV-pyMT mice. Additionally, both tumor infiltrating T-lymphocytes and tumor associated-macrophages were skewed towards TH2 in DOX-treated residual breast tumors; however, ESTA suppressed these changes. This study suggests that DOX treatment instigates de novo intratumoral infiltration of immune cells through E-selectin, and functional blockade of E-selectin may reduce residual tumor burden as well as metastasis through suppression of TH2 shift.

Prolactin Induced Protein (PIP) is a potential biomarker for early stage and malignant breast cancer.

OBJECTIVES: Breast cancer (BC) is the second leading cause of cancer-related mortality in women. Bioinformatic analysis and expression screening showed that Prolactin Induced Protein (PIP) was differentially expressed in BC. The objective of this investigation was to characterize the expression pattern of PIP, an aspartyl proteinase, in malignant and non-malignant breast tissues. MATERIALS AND METHODS: Real time quantitative PCR was employed to analyze PIP and androgen receptor (AR) mRNA levels in BC cell lines and 190 normal tissues and tumor samples. The tumor specimens were categorized based on TNM classification, anatomic stage, histologic grade and molecular subtype and expression pattern evaluated. To detect protein levels, immunohistochemistry followed by semi quantitative scoring was employed in the examination of 517 normal, benign, and invasive BC tissues. RESULTS: We observed substantial downregulation of PIP transcription in cancer samples compared to normal breast tissue. mRNA levels were significantly downregulated (93 fold, P < 0.005) in advanced grades compared to lower grades. Transcript levels were also significantly lower (22 fold, P < 0.05) in triple negative tumors compared to hormone receptor positive tumors. Significant downregulation was observed in early stage samples of triple negative and hormone receptor positive tumors. Though PIP protein showed a wide range of expression levels in BC, early stage samples showed significant downregulation. CONCLUSIONS: PIP mRNA is significantly downregulated in early stage BC compared to normal breast tissue. Consequently, low PIP mRNA expression in BC tissues could potentially be used as a tissue based biomarker to assist pathologists in confirmation of early stage BC diagnosis.

Aptamer Therapeutics in Cancer: Current and Future.

Aptamer-related technologies represent a revolutionary advancement in the capacity to rapidly develop new classes of targeting ligands. Structurally distinct RNA and DNA oligonucleotides, aptamers mimic small, protein-binding molecules and exhibit high binding affinity and selectivity. Although their molecular weight is relatively small-approximately one-tenth that of monoclonal antibodies-their complex tertiary folded structures create sufficient recognition surface area for tight interaction with target molecules. Additionally, unlike antibodies, aptamers can be readily chemically synthesized and modified. In addition, aptamers' long storage period and low immunogenicity are favorable properties for clinical utility. Due to their flexibility of chemical modification, aptamers are conjugated to other chemical entities including chemotherapeutic agents, siRNA, nanoparticles, and solid phase surfaces for therapeutic and diagnostic applications. However, as relatively small sized oligonucleotides, aptamers present several challenges for successful clinical translation. Their short plasma half-lives due to nuclease degradation and rapid renal excretion necessitate further structural modification of aptamers for clinical application. Since the US Food and Drug Administration (FDA) approval of the first aptamer drug, Macugen® (pegaptanib), which treats wet-age-related macular degeneration, several aptamer therapeutics for oncology have followed and shown promise in pre-clinical models as well as clinical trials. This review discusses the advantages and challenges of aptamers and introduces therapeutic aptamers under investigation and in clinical trials for cancer treatments.

Circulating Tumor Cells Accurately Predicting Progressive Disease After Treatment in a Patient with Non-small Cell Lung Cancer Showing Response on Scans.

Lung cancer is the leading cause of cancer-related deaths worldwide. Most patients present with advanced inoperable disease. Traditionally, responses to treatments are evaluated using different imaging modalities, which can sometimes be confusing. This is particularly more relevant in stage 3 disease where, after radiation therapy, persistent tumors on scans can represent active disease or scar tissue. We have been evaluating role of circulating tumor cells (CTCs) in that setting. Here we present the case of a 68-year-old male with stage 3 disease whose primary tumor responded to chemoradiotherapy on imaging, but whose CTC count was higher than the pre-treatment value. The patient later developed liver metastases. In this case, the CTC count more accurately predicted the patient's prognosis and highlights the need for exploration of the CTC count as a tool supplemental to imaging modalities.

Pathologic evaluation of tumor-associated macrophage density and vessel inflammation in invasive breast carcinomas.

Tumor-associated macrophages (TAMs) are major constituents of the tumor microenvironment in solid tumors and have been implicated as mediators of tumor progression, invasion and metastasis. Correspondingly, accumulation of TAMs is associated with unfavorable clinical outcomes in numerous types of solid tumors. E-selectin is a hallmark of inflammation and a key adhesion molecule that accommodates the initial contact of circulating immune cells with the inflamed vessel surface. Currently, the association between E-selectin and TAMs is not fully elucidated; therefore, the present study investigated the association between vessel inflammation, TAM infiltration, and clinical outcome in breast cancer. A total of 53 procedure-naïve invasive breast cancer cases were immunohistochemically analyzed for the presence of cluster of differentiation (CD)68+ TAMs, E-selectin+ vessels and tumor inflammation. The association between CD68 and E-selectin expression, and tumor inflammation as well as overall survival was evaluated using Kaplan-Meier survival curves and multivariable Cox's proportional hazards regression analysis. The abundance of TAMs was identified to be positively associated with tumor inflammation, estrogen receptor and E-selectin expression levels. A greater prevalence of TAMs and tumor inflammation was significantly associated with shorter overall survival times. E-selectin expression levels were significantly higher in tumor vessels among elderly patients, but were not associated with overall survival. The abundance of TAMs was associated with the presence of E-selectin-expressing inflamed tumor vessels and tumor inflammation, as well as overall survival in patients with invasive breast carcinoma.

E-selectin Targeting PEGylated-thioaptamer Prevents Breast Cancer Metastases.

E-selectin is an adhesion molecule expressed on the luminal surface of inflamed blood vessels that mediates hematogenous metastasis by assisting shear-resistant adhesion of circulating tumor cells to the vessel surface under dynamic blood flow. Previously, we developed an E-selectin antagonistic thioaptamer (ESTA) for the prevention of hematogenous metastasis through the blockade of CD44high breast cancer cells (BCa) adhesion to E-selectin-expressing premetastatic endothelial niche. The current study focuses on developing a PEGylated E-selectin targeting thioaptamer with improved pharmaceutical properties. A serial deletion of stem-loops reveled that loop-1 and -2 (ESTA7) are the minimally effective backbone structure necessary to obtain inhibition of the E-selectin/CD44 interaction and shear resistant adhesion of CD44high BCa to E-selectin-expressing human endothelial cells (HMVECs) at a level equal to ESTA. Chemical conjugation of methoxy-polyethylene-glycol (PEG) at the sizes of 5 and 10 kDa did not interfere with ESTA7-mediated shear-resistant adhesion. However, in vivo study demonstrated that only 10 kDa PEG-conjugated ESTA7 (ESTA7-p10) retains the activity to inhibit metastases at a level equal to parental ESTA. Additionally, a single intravenous injection of ESTA7-p10 inhibited the development of lung, brain, and bone metastases of MDA-MB-231, through the blockade of E-selectin. Moreover, PEGylation led to an extension of elimination half-life and increase of AUC, resulting in superior inhibition of metastasis development compared to parental ESTA with a longer interval between dosing in a spontaneous metastasis model. Lastly, repeated intravenous administration of ESTA7-p10 was tolerated in mice, highlighting the potential prophylactic application of ESTA7-p10 for metastasis prevention.

Improving vascular maturation using noncoding RNAs increases antitumor effect of chemotherapy.

Current antiangiogenesis therapy relies on inhibiting newly developed immature tumor blood vessels and starving tumor cells. This strategy has shown transient and modest efficacy. Here, we report a better approach to target cancer-associated endothelial cells (ECs), reverse permeability and leakiness of tumor blood vessels, and improve delivery of chemotherapeutic agents to the tumor. First, we identified deregulated microRNAs (miRs) from patient-derived cancer-associated ECs. Silencing these miRs led to decreased vascular permeability and increased maturation of blood vessels. Next, we screened a thioaptamer (TA) library to identify TAs selective for tumor-associated ECs. An annexin A2-targeted TA was identified and used for delivery of miR106b-5p and miR30c-5p inhibitors, resulting in vascular maturation and antitumor effects without inducing hypoxia. These findings could have implications for improving vascular-targeted therapy.