High-fat diet and obesity are associated with differential angiogenic gene expression in epithelial ovarian cancer.

OBJECTIVE: We sought to evaluate the association between diet and angiogenic biomarkers in KpB mice, and the association between these markers, body mass index (BMI), and overall survival (OS) in high-grade serous cancers (HGSC). METHODS: Tumors previously obtained from KpB mice subjected to high-fat diets (HFD, n = 10) or low-fat diets (LFD, n = 10) were evaluated for angiogenesis based on CD-31 microvessel density (MVD). Data from prior microarray analysis (Agilent 244 K arrays) conducted in 10 mice were utilized to assess associations between diet and angiogenetic biomarkers. Agilent (mouse) and Affymetrix Human Genome U133a probes were linked to 162 angiogenic-related genes. The associations between biomarkers, BMI, and OS were evaluated in an HGSC internal database (IDB) (n = 40). Genes with unadjusted p < 0.05 were evaluated for association with OS in the TCGA-OV database (n = 339). RESULTS: There was no association between CD-31 and diet in mice (p = 0.66). Sixteen angiogenic-related genes passed the p < 0.05 threshold for association with HFD vs. LFD. Transforming growth factor-alpha (TGFA) demonstrated 72% higher expression in HFD vs. LFD mice (p = 0.04). Similar to the mouse study, in our HGSC IDB, higher TGFA expression correlated with higher BMI (p = 0.01) and shorter survival (p = 0.001). In the TCGA-OV dataset, BMI data was not available and there was no association between TGFA and OS (p = 0.48). CONCLUSIONS: HFD and obesity may promote tumor progression via differential modulation of TGFA. We were unable to confirm this finding in the TCGA dataset. Further evaluation of TGFA is needed to determine if this is a target unique to obesity-driven HGSC.

Evaluating immune response and metabolic related biomarkers pre-allogenic hematopoietic stem cell transplant in acute myeloid leukemia.

Although hematopoietic stem cell transplantation (HCT) is the only curative treatment for acute myeloid leukemia (AML), it is associated with significant treatment related morbidity and mortality. There is great need for predictive biomarkers associated with overall survival (OS) and clinical outcomes. We hypothesized that circulating metabolic, inflammatory, and immune molecules have potential as predictive biomarkers for AML patients who receive HCT treatment. This retrospective study was designed with an exploratory approach to comprehensively characterize immune, inflammatory, and metabolomic biomarkers. We identified patients with AML who underwent HCT and had existing baseline plasma samples. Using those samples (n = 34), we studied 65 blood based metabolomic and 61 immune/inflammatory related biomarkers, comparing patients with either long-term OS (≥ 3 years) or short-term OS (OS ≤ 1 years). We also compared the immune/inflammatory response and metabolomic biomarkers in younger vs. older AML patients (≤30 years vs. ≥ 55 years old). In addition, the biomarker profiles were analyzed for their association with clinical outcomes, namely OS, chronic graft versus host disease (cGVHD), acute graft versus host disease (aGVHD), infection and relapse. Several baseline biomarkers were elevated in older versus younger patients, and baseline levels were lower for three markers (IL13, SAA, CRP) in patients with OS ≥ 3 years. We also identified immune/inflammatory response markers associated with aGVHD (IL-9, Eotaxin-3), cGVHD (Flt-1), infection (D-dimer), or relapse (IL-17D, bFGF, Eotaxin-3). Evaluation of metabolic markers demonstrated higher baseline levels of medium- and long-chain acylcarnitines (AC) in older patients, association with aGVHD (lactate, long-chain AC), and cGVHD (medium-chain AC). These differentially expressed profiles merit further evaluation as predictive biomarkers.

Financial incentives to increase stool collection rates for microbiome studies in adult bone marrow transplant patients.

INTRODUCTION: In order to study the role of the microbiome in hematopoietic stem cell transplantation (HCT), researchers collect stool samples from patients at various time points throughout HCT. However, stool collection requires active subject participation and may be limited by patient reluctance to handling stool. METHODS: We performed a prospective study on the impact of financial incentives on stool collection rates. The intervention group consisted of allogeneic HCT patients from 05/2017-05/2018 who were compensated with a $10 gas gift card for each stool sample. The intervention group was compared to a historical control group of allogeneic HCT patients from 11/2016-05/2017 who provided stool samples before the incentive was implemented. To control for possible changes in collections over time, we also compared a contemporaneous control group of autologous HCT patients from 05/2017-05/2018 with a historical control group of autologous HCT patients from 11/2016-05/2017; neither autologous HCT group was compensated. The collection rate was defined as the number of samples provided divided by the number of time points we attempted to obtain stool. RESULTS: There were 35 allogeneic HCT patients in the intervention group, 19 allogeneic HCT patients in the historical control group, 142 autologous HCT patients in the contemporaneous control group (that did not receive a financial incentive), and 75 autologous HCT patients in the historical control group. Allogeneic HCT patients in the intervention group had significantly higher average overall collection rates when compared to the historical control group allogeneic HCT patients (80% vs 37%, p<0.0001). There were no significant differences in overall average collection rates between the autologous HCT patients in the contemporaneous control and historical control groups (36% vs 32%, p = 0.2760). CONCLUSION: Our results demonstrate that a modest incentive can significantly increase collection rates. These results may help to inform the design of future studies involving stool collection.

Geriatric Assessment Reveals Actionable Impairments in Hematopoietic Stem Cell Transplantation Candidates Age 18 to 80 Years.

Allogeneic hematopoietic stem cell transplantation (HCT) is a potentially curative treatment for both malignant and nonmalignant hematologic diseases; however, reported rates of treatment-related mortality approach 30%. Outcomes are worse in patients who begin HCT with functional impairments. To detect such impairments, a geriatric assessment (GA) is recommended in adults age ≥65 years. Younger HCT candidates also may be impaired because of chemotherapy regimens pre-HCT. Therefore, we hypothesized that GA can be beneficial for adult patients of all ages and subsequently created a clinical pretransplantation optimization program to assess all HCT candidates using a modified GA. One-hundred fifty-seven patients were evaluated in 4 functional domains- physical, cognitive, nutritional, and psychological-at 2 time points prior to HCT-new patient evaluation (NPE) and sign-off (SO)-between October 2017 and January 2020. At NPE, 80.9% of the patients had at least 1 domain with a functional impairment, and physical (P = .006), cognitive (P = .04), and psychological (P = .04) impairments were associated with an increased likelihood of not proceeding to HCT. In addition, patients age 18 to 39 years were more likely than older patients to have a physical function impairment (P = .001). Between NPE and SO, 51.9% of the patients had resolution of 1 or more impairments, and nutritional impairment at SO was predictive of worse overall survival (P = .01). Our study shows that GA can identify functional impairments in patients of all ages. Early identification of impairments could facilitate referrals to supportive care and resolution of impairments prior to HCT, suggesting that GA could be recommended for HCT candidates of all ages.

Obesity and altered angiogenic-related gene expression in endometrial cancer.

OBJECTIVES: Evaluate association between obesity and angiogenic-related gene expression in endometrial cancer (EC). Evaluate interaction between diet and metformin on angiogenic-related gene expression. METHODS: We evaluated the association between 168 human angiogenic-related genes and body mass index (BMI) in the TCGA Uterine Corpus Endometrial Carcinoma cohort (endometrioid endometrial cancer (EEC) cohort n = 290, and copy number high cohort n = 55), an independent validation cohort from Gynecologic Cancer Center of Excellence (GYN-COE) (n = 62) and corresponding 185 homologous mouse genes in an LKB1fl/flp53fl/fl mouse model of EC (n = 20). Mice received 60% of calories from fat in a high-fat diet (HFD), mimicking diet-induced obesity, versus 10% of calories from fat in a low-fat diet (LFD). After tumor growth, HFD (n = 5) and LFD (n = 5) mice were treated with metformin (200 mg/kg/day) or control. Whole transcriptome analysis of mouse tumors was performed using RNA-Seq. RESULTS: At a false-discovery rate of 10%, twenty-one angiogenic-related genes were differentially expressed with respect to BMI when adjusting for grade in the TCGA EEC cohort. Evaluation of these genes in the mouse model control group revealed association between increased Edil3 expression in HFD versus LFD mice (2.5-fold change (FC); unadjusted p = 0.03). An interaction was observed for expression of Edil3 between diet and metformin treatment (unadjusted p = 0.009). Association between BMI and increased expression of EDIL3 was validated in one of four EDIL3 probesets in the GYN-COE cohort (p = 0.0011, adjusted p = 0.0342). CONCLUSIONS: Obesity may promote tumor progression via differential modulation of angiogenic pathways in EEC. Our exploratory findings demonstrated that EDIL3 may be a candidate gene of interest.

A phase 2 trial of the somatostatin analog pasireotide to prevent GI toxicity and acute GVHD in allogeneic hematopoietic stem cell transplant.

BACKGROUND: Allogeneic hematopoietic stem cell transplantation (HCT) is an often curative intent treatment, however it is associated with significant gastrointestinal (GI) toxicity and treatment related mortality. Graft-versus-host disease is a significant contributor to transplant-related mortality. We performed a phase 2 trial of the somatostatin analog pasireotide to prevent gastrointestinal toxicity and GVHD after myeloablative allogeneic HCT. METHODS: Patients received 0.9mg pasireotide every 12 hours from the day prior to conditioning through day +4 after HCT (or a maximum of 14 days). The primary outcomes were grade 3-4 gastrointestinal toxicity through day 30 and acute GVHD. Secondary outcomes were chronic GVHD, overall survival and relapse free survival at one year. Stool and blood samples were collected from before and after HCT for analyses of stool microbiome, local inflammatory markers, and systemic inflammatory and metabolic markers. Results were compared with matched controls. RESULTS: Twenty-six patients received pasireotide and were compared to 52 matched contemporaneous controls using a 1-2 match. Grade 3-4 GI toxicity occurred in 21 (81%) patients who received pasireotide and 35 (67%) controls (p = 0.33). Acute GVHD occurred in 15 (58%) patients in the pasireotide group and 28 (54%) controls (p = 0.94). Chronic GVHD occurred in 16 patients in the pasireotide group (64%) versus 22 patients in the control group (42%) (p = 0.12). Overall survival at 1 year in the pasireotide group was 63% (95% CI: 47%,86%) versus 82% (95% CI: 72%, 93%) in controls (log-rank p = 0.006). Relapse-free survival rate at one year was 40% (95% CI: 25%, 65%) in the pasireotide group versus 78% (95% CI: 68%, 91%) in controls (log-rank p = 0.002). After controlling for the effect of relevant covariates, patients in the pasireotide group had attenuated post-HCT loss of microbial diversity. Analysis of systemic inflammatory markers and metabolomics demonstrated feasibility of such analyses in patients undergoing allogeneic HCT. Baseline level and pre-to-post transplant changes in several inflammatory markers (including MIP1a, MIP1b, TNFa, IL8Pro, and IL6) correlated with likelihood of survival. CONCLUSIONS: Pasireotide did not prevent gastrointestinal toxicity or acute GVHD compared to contemporaneous controls. Pasireotide was associated with numerically higher chronic GVHD and significantly decreased OS and RFS compared to contemporaneous controls. Pasireotide may provide a locally protective effect in the stool microbiome and in local inflammation as measured by stool calprotectin, stool beta-defensin, and stool diversity index.

Decreased Mortality in 1-Year Survivors of Umbilical Cord Blood Transplant vs. Matched Related or Matched Unrelated Donor Transplant in Patients with Hematologic Malignancies.

Allogeneic hematopoietic stem cell transplantation (HCT) has the potential to cure hematologic malignancies but is associated with significant morbidity and mortality. Although deaths during the first year after transplantation are often attributable to treatment toxicities and complications, death after the first year may be due to sequelae of accelerated aging caused by cellular senescence. Cytotoxic therapies and radiation used in cancer treatments and conditioning regimens for HCT can induce aging at the molecular level; HCT patients experience time-dependent effects, such as frailty and aging-associated diseases, more rapidly than people who have not been exposed to these treatments. Consistent with this, recipients of younger cells tend to have decreased markers of aging and improved survival, decreased graft-versus-host disease, and lower relapse rates. Given that umbilical cord blood (UCB) is the youngest donor source available, we studied the outcomes after the first year of UCB transplantation versus matched related donor (MRD) and matched unrelated donor (MUD) transplantation in patients with hematologic malignancies over a 20-year period. In this single-center, retrospective study, we examined the outcomes of all adult patients who underwent their first allogeneic HCT through the Duke Adult Bone Marrow Transplant program from January 1, 1996, to December 31, 2015, to allow for at least 3 years of follow-up. Patients were excluded if they died or were lost to follow-up before day 365 after HCT, received an allogeneic HCT for a disease other than a hematologic malignancy, or received cells from a haploidentical or mismatched adult donor. UCB recipients experienced a better unadjusted overall survival than MRD/MUD recipients (log rank P = .03, median overall survival: UCB not reached, MRD/MUD 7.4 years). After adjusting for selected covariates, UCB recipients who survived at least 1 year after HCT had a hazard of death that was 31% lower than that of MRD/MUD recipients (hazard ratio, 0.69; 95% confidence interval, 0.47-0.99; P = .049). This trend held true in a subset analysis of subjects with acute leukemia. UCB recipients also experienced lower rates of moderate or severe chronic graft-versus-host disease (GVHD) and nonrelapse mortality, and slower time to relapse. UCB and MRD/MUD recipients experienced similar rates of grade 2-4 acute GVHD, chronic GHVD, secondary malignancy, and subsequent allogeneic HCT. UCB is already widely used as a donor source in pediatric HCT; however, adult outcomes and adoption have historically lagged behind in comparison. Recent advancements in UCB transplantation such as the implementation of lower-intensity conditioning regimens, double unit transplants, and ex vivo expansion have improved early mortality, making UCB an increasingly attractive donor source for adults; furthermore, our findings suggest that UCB may actually be a preferred donor source for mitigating late effects of HCT.

H2O2-Driven Anticancer Activity of Mn Porphyrins and the Underlying Molecular Pathways.

Mn(III) ortho-N-alkyl- and N-alkoxyalkyl porphyrins (MnPs) were initially developed as superoxide dismutase (SOD) mimics. These compounds were later shown to react with numerous reactive species (such as ONOO-, H2O2, H2S, CO3 •-, ascorbate, and GSH). Moreover, the ability of MnPs to oxidatively modify activities of numerous proteins has emerged as their major mechanism of action both in normal and in cancer cells. Among those proteins are transcription factors (NF-κB and Nrf2), mitogen-activated protein kinases, MAPKs, antiapoptotic bcl-2, and endogenous antioxidative defenses. The lead Mn porphyrins, namely, MnTE-2-PyP5+ (BMX-010, AEOL10113), MnTnBuOE-2-PyP5+ (BMX-001), and MnTnHex-2-PyP5+, were tested in numerous injuries of normal tissue and cellular and animal cancer models. The wealth of the data led to the progression of MnTnBuOE-2-PyP5+ into four Phase II clinical trials on glioma, head and neck cancer, anal cancer, and multiple brain metastases, while MnTE-2-PyP5+ is in Phase II clinical trial on atopic dermatitis and itch.

Microtransplantation in older patients with AML: A pilot study of safety, efficacy and immunologic effects.

Older AML patients have low remission rates and poor survival outcomes with standard chemotherapy. Microtransplantation (MST) refers to infusion of allogeneic hematopoietic stem cells without substantial engraftment. MST has been shown to improve clinical outcomes compared with chemotherapy alone. This is the first trial reporting on broad correlative studies to define immunologic mechanisms of action of MST in older AML patients. Older patients with newly diagnosed AML were eligible for enrollment, receiving induction chemotherapy with cytarabine (100 mg/m2) on days 1-7 and idarubicin (12 mg/m2) on days 1-3 (7 + 3). MST was administered 24 hours later. Patients with complete response (CR) were eligible for consolidation with high dose cytarabine (HiDAC) and a second cycle of MST. Responses were evaluated according to standard criteria per NCCN. Immune correlative studies were performed. Sixteen patients were enrolled and received 7 + 3 and MST (median age 73 years). Nine (56%) had high-risk and seven (44%) had standard-risk cytogenetics. Ten episodes of CRS were observed. No cases of GVHD or treatment-related mortality were reported. Event-free survival (EFS) was 50% at 6 months and 19% at 1 year. Overall survival (OS) was 63% at 6 months and 44% at 1 year. Donor microchimerism was not detected. Longitudinal changes were noted in NGS, TCR sequencing, and cytokine assays. Addition of MST to induction and consolidation chemotherapy was well tolerated in older AML patients. Inferior survival outcomes in our study may be attributed to a higher proportion of very elderly patients with high-risk features. Potential immunologic mechanisms of activity of MST include attenuation of inflammatory cytokines and emergence of tumor-specific T cell clones.

Differential expression of immune related genes in high-grade ovarian serous carcinoma.

OBJECTIVE: To identify novel immunologic targets and biomarkers associated with overall survival (OS) in high-grade serous ovarian cancer (HGSC). METHODS: In this retrospective study, microarray data from 51 HGSC specimens were analyzed (Affymetrix HG-U133A). A panel of 183 immune/inflammatory response related genes linked to 279 probe sets was constructed a priori and screened. Associations between gene expression and OS were assessed using logrank tests. Multiple testing was addressed within the False Discovery Rate (FDR) framework. For external validation, TCGA Ovarian dataset and five GSE publicly available HGSC datasets were evaluated. RESULTS: In Duke data, 110 probe sets linked to 83 immunologic/inflammatory-related genes were differentially expressed in tumors from long versus short-term HGSC survivors (adjusted p < 0.05). In TCGA, concordant with the results from the Duke discovery cohort, high expression of one probe (IL6R) demonstrated a consistent significance and concordant association with higher expression in long-term HGSC survivors (Duke q-value = 0.022) and improved OS in the TCGA dataset (p-value = 0.015, HR = 0.8). Thirteen genes in GSE14764 (N = 4) and GSE26712 (N = 9) datasets had significant p-values and consistent concordant with Duke Data. Despite the significant associations of gene expression and OS in the individual GSE datasets, in the GSE meta-analysis no genes were consistently concordant and significantly associated with survival. CONCLUSIONS: Evaluation of IL6R expression may be warranted based on higher expression in long-term survivors and association with improved survival in advanced HGSC. The other candidate genes may also be of worthy of further exploration to enhance immuno-oncology drug discovery.

Late Gastrointestinal Complications of Allogeneic Hematopoietic Stem Cell Transplantation in Adults.

Gastrointestinal (GI) complications including graft-versus-host disease (GVHD) are a major cause of morbidity and mortality in allogenic stem transplant recipients. Although several studies have previously looked into the acute GI complications, fewer smaller studies have reported late complications. In this large study we focus on the late (100 days post-transplant) GI complications in allogenic stem transplant recipients. In this single-center, retrospective study of all adult allogenic stem cell transplant recipients who had their transplant at Duke University over a 6-year period, 479 patients underwent allogenic stem cell transplant, of whom 392 recipients survived for at least 100 days post-transplant. Late GI symptoms were noted in 71 patients, prompting endoscopic evaluation. The primary endpoint of our study was the diagnosis of GI-GVHD based on endoscopic findings, whereas overall survival and nonrelapse mortality were the secondary endpoints. Of the 71 patients who underwent endoscopy, 45 (63%) had GI-GVHD. Of these 45 patients, 39 (87%) had late acute GVHD, 1 (2%) had chronic GVHD, and 5 patients (11%) had overlap disease. Of the patients who did not have GVHD, the symptoms were mostly related to infectious and inflammatory causes. Less common causes included drug toxicity, food intolerance, disease relapse, and motility issues. In a multivariate analysis the factors most indicative of GI-GVHD were histologic findings of apoptosis on the tissue specimen (odds ratio, 2.35; 95% confidence interval, 1.18 to 4.70; P = .015) and clinical findings of diarrhea (odds ratio, 5.43; 95% confidence interval, 1.25 to 23.54; P = .024). The median survival time from the first endoscopy was 8.5 months. The incidence of nonrelapse mortality at 6 months was 31% in patients with GI-GVHD and 19% in patients without GI-GVHD (P = .42). All patients with GI-GVHD were on steroid therapy, and 31% of them received total parenteral nutrition. In our population close to one-fifth of allogenic transplant recipients experienced late GI complications, warranting endoscopic evaluation. Most of these patients were found to have GI-GVHD that had a high incidence of nonrelapse mortality at 6 months and close to one-third of these patients needed total parenteral nutrition.

Subtype-Specific Radiation Response and Therapeutic Effect of FAS Death Receptor Modulation in Human Breast Cancer.

Breast cancer is the most common malignancy diagnosed among women and represents a heterogeneous group of subtypes. Radiation therapy is a critical component of treatment for breast cancer patients. However, little is known about radiation response among these intrinsic subtypes. In previous studies, we identified a significant induction of FAS after irradiation in biologically favorable breast cancer patients and breast cancer cell lines. Here, we expanded our study and investigated radiation response in a mouse model of breast cancer. MCF7 (luminal), HCC1954 (HER2+) or SUM159 (basal) cells were implanted orthotopically into the dorsal mammary fat pad of nude mice. These mice were then treated with different doses of radiation to assess tumor growth control. We further investigated the therapeutic effect of FAS modulation by silencing FAS in radiation-responsive tumors and injecting FAS agonist antibody into radiation-resistant tumors. Exposure to radiation inhibited MCF7, and to a lesser extent HCC1954 tumor growth in a dose-dependent manner. In contrast, SUM159 tumors were resistant to radiation. The estimated TCD50 values were 19.3 Gy for MCF7 and 44.9 Gy for SUM159. Radiation induced FAS expression in MCF7 tumors, but not SUM159 tumors. We found that silencing of FAS did not negatively impact radiation response in MCF7 tumors, possibly due to compensation by other apoptotic pathways. On the other hand, FAS activating antibody in combination with radiation treatment delayed SUM159 and HCC1954 tumor growth. However, it did not reach statistical significance compared to radiation treatment alone. Our results suggest that there is intrinsic variation in radiation response among breast cancer subtypes. FAS activation concurrent with radiation slows tumor growth in the radiation-resistant subtypes, but the effect was not significant. Alternative subtype-specific modulators of radiation response are under investigation.

FAS Death Receptor: A Breast Cancer Subtype-Specific Radiation Response Biomarker and Potential Therapeutic Target.

Although a standardized approach to radiotherapy has been used to treat breast cancer, regardless of subtype (e.g., luminal, basal), recent clinical data suggest that radiation response may vary significantly among subtypes. We hypothesized that this clinical variability may be due, in part, to differences in cellular radiation response. In this study, we utilized RNA samples for microarray analysis from two sources: 1. Paired pre- and postirradiation breast tumor tissue from 32 early-stage breast cancer patients treated in our unique preoperative radiation Phase I trial; and 2. Sixteen biologically diverse breast tumor cell lines exposed to 0 and 5 Gy irradiation. The transcriptome response to radiation exposure was derived by comparing gene expression in samples before and after irradiation. Genes with the highest coefficient of variation were selected for further evaluation and validated at the RNA and protein level. Gene editing and agonistic antibody treatment were performed to assess the impact of gene modulation on radiation response. Gene expression in our cohort of luminal breast cancer patients was distinctly different before and after irradiation. Further, two distinct patterns of gene expression were observed in our biologically diverse group of breast cancer cell lines pre- versus postirradiation. Cell lines that showed significant change after irradiation were largely luminal subtype, while gene expression in the basal and HER2+ cell lines was minimally impacted. The 100 genes with the most significant response to radiation in patients were identified and analyzed for differential patterns of expression in the radiation-responsive versus nonresponsive cell lines. Fourteen genes were identified as significant, including FAS, a member of the tumor necrosis factor receptor family known to play a critical role in programed cell death. Modulation of FAS in breast cancer cell lines altered radiation response phenotype and enhanced radiation sensitivity in radioresistant basal cell lines. Our findings suggest that cell-type-specific, radiation-induced FAS contributes to subtype-specific breast cancer radiation response and that activation of FAS pathways may be exploited for biologically tailored radiotherapy.

Prognostic significance of differential expression of angiogenic genes in women with high-grade serous ovarian carcinoma.

OBJECTIVES: To identify angiogenic biomarkers associated with tumor angiogenesis and clinical outcome in high-grade serous ovarian cancer (HGSC). METHODS: 51 HGSC samples were analyzed using Affymetrix HG-U133A microarray. Microvessel density (MVD) counts were determined using CD31 and CD105. Associations between mRNA expression levels and overall survival were assessed using rank score statistic. Effect size was estimated as a hazard ratio (HR) under a proportional hazard model. The Storey q-value method was used to account for multiple testing within the false-discovery rate (FDR) framework. Publicly available databases including TCGA and GSE were used for external confirmation. RESULTS: Thirty-one angiogenic-related genes were significantly associated with survival (q≤0.05). Of these 31 genes, 4 were also associated with outcome in the TCGA data: AKT1 (q=0.02; TCGA p=0.01, HR=0.8), CD44 (q=0.003; TCGA p=0.05, HR=0.9), EPHB2 (q=0.01; TCGA p=0.05, HR=1.2), and ERBB2 (q=0.02; TCGA p=0.05, HR=1.2). While 5 were associated with outcome in the GSE database: FLT1 (q=0.03; GSE26712 p=0.01, HR=3.1); PF4 (q=0.02; GSE26712 p=0.01, HR=3.0); NRP1 (q=0.02; GSE26712 p<0.04, HR>1.4); COL4A3 (q=0.04; GSE26712 p=0.03, HR=1.3); and ANGPTL3 (q=0.02; GSE14764 p=0.02, HR=1.5). High AKT1 and CD44 were associated with longer survival. In contrast, high expression of EPHB2, ERBB2, FLT1; PF4, NRP1, COL4A3, and ANGPTL3 were associated with shorter survival. CD105-MVD and CD31-MVD were not significantly associated with angiogenic gene expression. CONCLUSIONS: Thirty-one angiogenic-related genes were associated with survival in advanced HGSC and nine of these genes were confirmed in independent publicly available databases.

Preoperative Single-Fraction Partial Breast Radiation Therapy: A Novel Phase 1, Dose-Escalation Protocol With Radiation Response Biomarkers.

PURPOSE: Women with biologically favorable early-stage breast cancer are increasingly treated with accelerated partial breast radiation (PBI). However, treatment-related morbidities have been linked to the large postoperative treatment volumes required for external beam PBI. Relative to external beam delivery, alternative PBI techniques require equipment that is not universally available. To address these issues, we designed a phase 1 trial utilizing widely available technology to 1) evaluate the safety of a single radiation treatment delivered preoperatively to the small-volume, intact breast tumor and 2) identify imaging and genomic markers of radiation response. METHODS AND MATERIALS: Women aged ≥55 years with clinically node-negative, estrogen receptor-positive, and/or progesterone receptor-positive HER2-, T1 invasive carcinomas, or low- to intermediate-grade in situ disease ≤2 cm were enrolled (n=32). Intensity modulated radiation therapy was used to deliver 15 Gy (n=8), 18 Gy (n=8), or 21 Gy (n=16) to the tumor with a 1.5-cm margin. Lumpectomy was performed within 10 days. Paired pre- and postradiation magnetic resonance images and patient tumor samples were analyzed. RESULTS: No dose-limiting toxicity was observed. At a median follow-up of 23 months, there have been no recurrences. Physician-rated cosmetic outcomes were good/excellent, and chronic toxicities were grade 1 to 2 (fibrosis, hyperpigmentation) in patients receiving preoperative radiation only. Evidence of dose-dependent changes in vascular permeability, cell density, and expression of genes regulating immunity and cell death were seen in response to radiation. CONCLUSIONS: Preoperative single-dose radiation therapy to intact breast tumors is well tolerated. Radiation response is marked by early indicators of cell death in this biologically favorable patient cohort. This study represents a first step toward a novel partial breast radiation approach. Preoperative radiation should be tested in future clinical trials because it has the potential to challenge the current treatment paradigm and provide a path forward to identify radiation response biomarkers.

Exercise behavior and patient-reported outcomes in women with early breast cancer receiving locoregional radiation therapy.

PURPOSE: Radiation therapy is associated with acute treatment-related complications that can lead to decreased quality of life (QOL). Exercise has been shown in other cancer treatment settings to improve negative outcomes. We conducted a prospective pilot study to explore the association between exercise, patient-reported outcomes, and acute radiation therapy toxicities. METHODS AND MATERIALS: Women receiving curative breast radiation therapy were enrolled. Each patient completed an exercise behavior/QOL survey before or during the first week of treatment and again during the last week of treatment. Exercise behavior was quantified with the Godin Leisure Time Exercise Questionnaire (metabolic equivalent [MET] hours per week). Measurements to evaluate upper extremity lymphedema and shoulder range of motion were completed. Skin toxicity was assessed weekly. Patient-reported outcomes were measured using standardized questionnaires. RESULTS: Forty-five patients were enrolled. Mean patient age was 54 (range, 28-73) years. Mean METs in the exercise cohort (≥9 METs/wk) was 21 per week (range, 11-38, n = 14); 3 per week (range, 0-8, n = 25) in the nonexercise cohort (<9 METs/wk). Women in the exercise cohort showed improvements in treatment-induced quality of life and fatigue (not significant) despite more extensive surgical, medical, and radiation treatment. No differences in treatment-related toxicities, pain, or sleep scores were noted. Lymphedema was mild (<3 cm) in the entire patient cohort. CONCLUSIONS: The vast majority of current exercise oncology literature implicates physical activity as an independent predictor of QOL in cancer patients. Our study noted similar trends, but they were not statistically significant. This may be due to our finding that patient-reported outcomes with radiation therapy are relatively high compared with other treatment modalities and remain stable throughout treatment. Thus, it may be that radiation therapy has a limited impact on QOL in breast cancer patients. Exercise may be best used as a targeted therapy in patients at high risk for poor QOL or radiation-related toxicities at baseline.

Genomic profiling in locally advanced and inflammatory breast cancer and its link to DCE-MRI and overall survival.

PURPOSE: We have previously reported that dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) perfusion patterns obtained from locally advanced breast cancer (LABC) patients prior to neoadjuvant therapy predicted pathologic clinical response. Genomic analyses were also independently conducted on the same patient population. This retrospective study was performed to test two hypotheses: (1) gene expression profiles are associated with DCE-MRI perfusion patterns, and (2) association between long-term overall survival data and gene expression profiles can lead to the identification of novel predictive biomarkers. METHODS: We utilised RNA microarray and DCE-MRI data from 47 LABC patients, including 13 inflammatory breast cancer (IBC) patients. Association between gene expression profile and DCE-MRI perfusion patterns (centrifugal and centripetal) was determined by Wilcoxon rank sum test. Association between gene expression level and survival was assessed using a Cox rank score test. Additional genomic analysis of the IBC subset was conducted, with a period of follow-up of up to 11 years. Associations between gene expression and overall survival were further assessed in The Cancer Genome Atlas Data Portal. RESULTS: Differences in gene expression profiles were seen between centrifugal and centripetal perfusion patterns in the sulphotransferase family, cytosolic, 1 A, phenol-preferring, members 1 and 2 (SULT1A1, SULT1A2), poly (ADP-ribose) polymerase, member 6 (PARP6), and metastasis tumour antigen1 (MTA1). In the IBC subset our analyses demonstrated that differential expression of 45 genes was associated with long-term survival. CONCLUSIONS: Here we have demonstrated an association between DCE-MRI perfusion patterns and gene expression profiles. In addition we have reported on candidate prognostic biomarkers in IBC patients, with some of the genes being significantly associated with survival in IBC and LABC.

Differential Angiogenic Gene Expression in TP53 Wild-Type and Mutant Ovarian Cancer Cell Lines.

OBJECTIVES: Underlying mechanisms regulating angiogenesis in ovarian cancer have not been completely elucidated. Evidence suggests that the TP53 tumor suppressor pathway and tumor microenvironment play integral roles. We utilized microarray technology to study the interaction between TP53 mutational status and hypoxia on angiogenic gene expression. METHODS: Affymetrix U133A arrays were analyzed for angiogenic gene expression in 19 ovarian cancer cell lines stratified both by TP53 mutation status and A2780 wild-type (wt) TP53 vs. mutated (m) TP53 cell lines after treatment under hypoxic conditions or with ionizing radiation. RESULTS: Twenty-eight differentially expressed angiogenic genes were identified in the mTP53 cell lines compared to wtTP53 lines. Five genes were upregulated in mTP53 cells: 40% involved in extracellular matrix (ECM) degradation [matrix metalloproteinase 10 (MMP10)/15] and 60% in angiogenesis (fibroblast growth factor receptor 3/VEGFA/ephrin receptor-B4). Twenty-three genes were upregulated in wtTP53: nearly 22% were ECM constituents or involved in ECM degradation; over 40% were growth factors or mediators of angiogenesis. Five genes were upregulated in the A2780mTP53 cells: 40% involved in ECM remodeling (MMP10, ADAMTS1), 40% with pro-angiogenic activity (EFNB2, factor 2 receptor), and 20% with anti-angiogenic properties (ADAMTS1). Three genes were upregulated in hypoxia treated cells compared to controls: one with anti-angiogenic activity (angiopoietin-like 4) and two with pro-angiogenic activity (VEGFA, EFNA3). No significant gene fold changes were noted after exposure to radiation. Four genes continued to demonstrate significant differential expression (p ≤ 0.05) after adjusting for multiple comparisons. These genes included endoglin upregulation in wt lines (pro-angiogenesis) and upregulation of FGF20 (growth factor), ADAMTS1 (anti-angiogenesis) and MMP10 (ECM degradation) in mTP53 cell lines. CONCLUSION: Our exploratory findings indicate that non-overlapping angiogenic pathways may be altered by TP53 mutations and hypoxic conditions in the tumor microenvironment. Further evaluation is needed for confirmation.

A novel monoclonal antibody to secreted frizzled-related protein 2 inhibits tumor growth.

Secreted frizzled-related protein 2 (SFRP2) is overexpressed in human angiosarcoma and breast cancer and stimulates angiogenesis via activation of the calcineurin/NFATc3 pathway. There are conflicting reports in the literature as to whether SFRP2 is an antagonist or agonist of ß-catenin. The aims of these studies were to assess the effects of SFRP2 antagonism on tumor growth and Wnt-signaling and to evaluate whether SFRP2 is a viable therapeutic target. The antiangiogenic and antitumor properties of SFRP2 monoclonal antibody (mAb) were assessed using in vitro proliferation, migration, tube formation assays, and in vivo angiosarcoma and triple-negative breast cancer models. Wnt-signaling was assessed in endothelial and tumor cells treated with SFRP2 mAb using Western blotting. Pharmacokinetic and biodistribution data were generated in tumor-bearing and nontumor-bearing mice. SFRP2 mAb was shown to induce antitumor and antiangiogenic effects in vitro and inhibit activation of ß-catenin and nuclear factor of activated T-cells c3 (NFATc3) in endothelial and tumor cells. Treatment of SVR angiosarcoma allografts in nude mice with the SFRP2 mAb decreased tumor volume by 58% compared with control (P = 0.004). Treatment of MDA-MB-231 breast carcinoma xenografts with SFRP2 mAb decreased tumor volume by 52% (P = 0.03) compared with control, whereas bevacizumab did not significantly reduce tumor volume. Pharmacokinetic studies show the antibody is long circulating in the blood and preferentially accumulates in SFRP2-positive tumors. In conclusion, antagonizing SFRP2 inhibits activation of ß-catenin and NFATc3 in endothelial and tumor cells and is a novel therapeutic approach for inhibiting angiosarcoma and triple-negative breast cancer.

Systemic markers of oxidative status and colorectal adenomatous polyps.

PURPOSE: Oxidative damage has been implicated in carcinogenesis. We hypothesized that elevated systemic oxidative status would be associated with later occurrence of colorectal adenomatous polyps, a precursor of colorectal cancer. METHODS: We examined the prospective association between four systemic markers of oxidative status and colorectal adenomatous polyps within a nondiabetic subcohort of the Insulin Resistance Atherosclerosis Study (n = 425). Urine samples were collected from 1992 to 1994 and colorectal adenomas prevalence were assessed in 2002 to 2004. Oxidative status markers were assessed, which included four F(2)-isoprostanes (F(2)-IsoPs) from the classes III and IV: iPF2α-III, 2,3-dinor-iPF2α-III (a metabolite of iPF2α-III), iPF2α-VI, and 8,12-iso-iPF2α-VI. All biomarkers were quantified using liquid chromatography-tandem mass spectrometry. Prospective associations were assessed using multivariate logistic regression analysis. RESULTS: The adjusted odds ratio (OR) (95% confidence interval [CI]) for occurrence of colorectal adenomatous polyps and scaled to 1 SD of F(2)-IsoP distribution were 1.16 (95% CI, 0.88-1.50), 0.88 (95% CI, 0.63-1.17), 1.04 (95% CI, 0.80-1.34), and 1.16 (95% CI, 0.90-1.48) for iPF2α-III, iPF2α-VI, 8,12-iso-iPF2α-VI, and 2,3-dinor-iPF2α-III, respectively. CONCLUSIONS: The lack of association between F(2)-IsoPs and adenomatous polyps does not support the hypothesis that elevated oxidative status is associated with colorectal adenomatous polyp occurrence during a 10-year period of follow-up.