PDF file - 1147KB, 2 x 106 MDA-MB-231 or MDA-MB-435 cells were injected into 6- to 7-week-old female NOD/SCID or nu/nu mice via mammary fat pad. The control cohort (n=4) received weekly saline subcutaneous injections, while three other cohorts (n=4 per group) received rHuEPO doses of 300U/kg, 600U/kg or 1200U/kg. (A-D) Data from MDA-MB-231 breast cancer cells in NOD/SCID mice. (E-H) Results from MDA-MB-435 breast cancer cells injected into nude (nu/nu) mice. (A,E) Primary tumor growth of human breast cancer cells injected into mice over time. EPO treatment was initiated at 3 weeks post injection (black arrow). Data are presented as mean SEM. (B,F) Incidence of lung metastasis (% of mice in each treatment group that developed metastases). (C,G) Size (diameter) of lung metastases scored. Data are presented as mean SEM. (D,H) Number of lung metastases observed. Data are presented as the mean SEM. For all panels, # denotes a statistically significant difference relative to the untreated control cohort (p<0.05).

PDF file - 119KB, Supplemental Table 1- Cell lines used. Supplemental Table 2 - PCR primers and cycling conditions. Supplemental Table 3 - Anti Human EPOR antibodies used. Supplemental Table 4 - In Vivo hemoglobin measurements.

PDF file - 145KB, EPOR and JAK2 amplification was normalized to GAPDH intensity for mBM, UT7/EPO, 3T3, H838, MDA-MB-435, MDA-MB-231, MDA-MB-468 and 4T1 samples, with (*) or without rHuEPO treatment and subjected to densitometric analysis.

The optimal management for acute tibial deep venous thrombosis (DVT) is not well established, with no consensus in the current literature. The American College of Chest Physicians suggests that treatment depends on the severity of the patient's symptoms and risk factors for thrombus extension, involving either anticoagulation or serial duplex ultrasound examinations. The purpose of this study was to evaluate the outcomes of tibial DVT managed with and without anticoagulation and to identify factors that influenced these outcomes in patients with tibial DVT. We performed a retrospective chart review of all patients who were diagnosed with tibial DVT with at least one duplex ultrasound study within 30 days after diagnosis from January 2009 to December 2014. Excluded were patients with concomitant proximal DVT. Primary outcomes were clot propagation and new DVT. Patients were divided into those who received full-dose anticoagulation and those who did not. Additional information including patient demographics, comorbidities, and risk factors were also gathered and analyzed using Fisher exact test and a multivariate logistic regression model. There were 141 patients diagnosed with tibial DVT, of whom 115 met inclusion criteria. Seventy-five patients were treated with anticoagulation and 40 patients were not. Clot propagation occurred in 13 of 75 (17.3%) in the anticoagulation group and 7 of 40 (17.5%) in the no-anticoagulation treatment group (P = 1.000). Of the variables and risk factors analyzed, none were shown to be statistically significant in relation to clot progression. Multivariate regression analysis reiterated that anticoagulation had no effect on progression when accounting for other risk factors. In our retrospective study, no individual risk factors were found to be prognostic for clot propagation. Patients treated with anticoagulation did not appear to fare better than those who did not receive anticoagulation with respect to clot propagation. Therefore, until a larger, more adequately powered prospective study can be performed, we will continue to manage patients with tibial DVT without anticoagulation, reserving this therapy only for those patients who have a history of cancer or demonstrate clot propagation on serial scanning with duplex ultrasound.

The optimal management for tibial deep vein thrombosis (tDVT) is not well established. The purpose of this study was to evaluate the outcomes of tDVT managed with and without anticoagulation and to identify factors that influenced these outcomes. We performed a retrospective record review of all patients who were diagnosed with tDVT that had at least one duplex ultrasound (DU) study within 30 days following diagnosis from January 2009 to December 2014. Excluded were patients with concomitant proximal DVT. Primary outcomes evaluated were clot propagation and/or new DVT as measured with DU. Patients were divided into two groups: those who received full-dose anticoagulation and those who did not. Additional information including patient demographics, comorbidities, recent travel, trauma, or surgery, body mass index, history of prior DVT, ambulatory, and inpatient status were also gathered. The data were analyzed using the Fisher exact test. During the study period, 141 patients were diagnosed with tDVT, of whom 115 met inclusion criteria. Seventy-five patients were treated with anticoagulation, and 40 patients were not. Clot propagation occurred in 21 patients, 13 of 75 (17.3%) in the anticoagulation group and eight of 40 (20%) in the no anticoagulation treatment group. None of the variables and risk factors analyzed were shown to be statistically significant in relation to clot progression. A history of cancer was very nearly significant (P = .06). In our retrospective study, no individual risk factors were found to be prognostic for clot propagation. Patients treated with anticoagulation did not appear to fare better than those that did not receive anticoagulation with respect to clot propagation. Therefore, until a larger, more adequately powered prospective study can be performed, we will continue to manage patients with tDVT without anticoagulation, reserving this therapy for only those patients who have a history of cancer and or demonstrate clot propagation on serial scanning with duplex ultrasound imaging.

Mobilization of hematopoietic stem cells is achieved with hematopoietic growth factors with or without chemotherapy or other agents. Although studies comparing granulocyte colony‐stimulating factor (G‐CSF) alone to combined regimens demonstrate an increase in stem cell yield in the latter, mobilization with G‐CSF alone is still effective and has been widely practiced. We conducted a retrospective cohort study of consecutive patients at our institution who underwent at least one mobilization attempt with G‐CSF between January 2000 and December 2008 to identify the proportion of patients failing one or more mobilization attempts and the potential predictors of mobilization failure with this regime. Out of 293 patients, 251 (86.6%) were successfully mobilized and 244 (83.6%) underwent hematopoietic stem cell transplantation. Median yield was 3.55 × 106 CD34+ cells/kg. On univariate analysis, mobilization success was influenced by degree of previous treatment and underlying diagnosis (P < 0.001 each) but not by age (P = 0.114), sex (P = 0.860), or radiotherapy (P = 0.454). A diagnosis of non‐Hodgkin's lymphoma (NHL) and number of previous chemotherapy regimens were predictors of failure on multivariate analysis. CD34+ yield was influenced by diagnosis and previous chemotherapy (P < 0.001 each). Mobilization with G‐CSF alone yields adequate collections for most patients; however, heavily pretreated NHL patients with one failed attempt had high rates of remobilization failure and should be considered for alternative regimens. J. Clin. Apheresis 28:285–292, 2013. © 2013 Wiley Periodicals, Inc.

Abstract 2107This icon denotes a clinically relevant abstract

Abstract Purpose: Erythropoiesis-stimulating agents (ESA) are used clinically for treating cancer-related anemia. Recent clinical trials have reported increased adverse events and reduced survival in ESA-treated breast cancer patients receiving chemotherapy, potentially related to erythropoietin (EPO)-induced cancer progression. However, minimal preclinical data are available about the impact of EPO on metastatic cell behavior and/or the metastatic process, and this was the goal of our study. Experimental Design: Breast cancer cell lines were treated with recombinant human EPO (rHuEPO) and screened for expression of EPO receptors (EPOR). MDA-MB-231 and MDA-MB-435 cell lines were used for functional assays in vitro (two-dimensional/three-dimensional growth and survival) and in vivo (tumorigenicity and metastasis), in the presence or absence of EPO and/or cytotoxic agents. Results: A large variation in EPOR expression across cell lines was observed. In vitro, rHuEPO had a protective effect on radiation-treated MDA-MB-435 cells (P < 0.05); however, rHuEPO treatment alone or combined with chemotherapy or hypoxia did not influence cell survival. In vivo, rHuEPO increased lung metastases in immunocompromised mice injected with MDA-MB-231 or MDA-MB-435 cells and treated with chemotherapy relative to mice treated with chemotherapy alone (P < 0.05). Conclusions: The lack of an in vitro effect of rHuEPO highlights the importance of in vivo studies to delineate the effects of EPO on the metastatic process. These studies may begin to uncover the underlying functional explanation for the observed EPO-related adverse events and decreased survival in ESA-treated metastatic breast cancer patients undergoing chemotherapy. Clin Cancer Res; 17(19); 6151–62. ©2011 AACR.

Abstract Abstract 4445 Background Mobilization of hematopoetic stem cells (HSC) in patients undergoing AHSCT for hematological malignancies is usually done using hematopoietic growth factors such as G-CSF with or without concurrent use of chemotherapy or other agents such as plerixafor. While studies comparing G-CSF alone to combination regimens demonstrate an increase in the yield of stem cells in the latter case, mobilization with G-CSF alone is still effective and represents the standard of care at our institution. Therefore, we aimed to identify potential predictors of mobilization failure with G-CSF alone in patients undergoing AHSCT for hematological malignancies and for which alternative regimens might be considered. Patients and Methods We conducted a single centre retrospective case-control study of all consecutive patients who underwent at least one mobilization attempt with G-CSF for an AHSCT at the London Health Sciences Centre in London, Ontario, Canada between January 2000 and December 2008. The mobilization regimen consisted of G-CSF 10 μ g/kg/day for 4 days with collection on days 5 and 6. The primary outcome was successful mobilization defined as the collection of at least 2.0×109 CD34+cells/kg. The secondary outcome was the mean yield of stem cells mobilized. Groups (successful vs. unsuccessful mobilization) were compared using unpaired Student's t, Mann-Whitney U, χ2 or Fisher's exact tests, as appropriate. Logistic regression analysis was conducted using an unsuccessful mobilization as the dependent variable. CD34+cells/kg yields were compared using unpaired Student's t tests or one-way ANOVA. Results During the study period, mobilization was attempted in 293 patients (134 MM, 57 HD, 86 NHL, 17 Other). The mean age was 47.5±12.3 years. 251 patients (86.6%; 95%CI 82.3, 90.1) were successfully mobilized and 244 (83.6; 95%CI 78.9, 87.4) underwent AHSCT. The median yield was 3.55 ×106CD34+ cells/kg (Interquartile range 2.50–5.30). On univariate analysis, mobilization success was influenced by the number of previous chemotherapy regimens and underlying diagnosis (P<0.001 each), but not by age (P=0.114), sex (P=0.860) or prior radiotherapy (P=0.454). A diagnosis of NHL and number of previous chemotherapy regimens were predictors of unsuccessful mobilization on multivariate analysis (Table 1). CD34+cells/kg yield was influenced by diagnosis and previous chemotherapy (P <0.001 each). The percentage of patients with successful and unsuccessful mobilization using G-CSF alone according to diagnosis and number of attempts is shown in Table 2. Conclusions HSC mobilization with G-CSF alone yields adequate collections for most patients. Patients with NHL and patients treated with 2 or more previous chemotherapy regimens that fail an initial mobilization attempt have higher failure rates and can be considered for alternate mobilization regimes. Disclosures: Howson-Jan: Merck: Research Funding.

Although lymphatic dissemination is a major route for breast cancer metastasis, there has been little work to determine what factors control the ability of tumor cells to survive, establish and show progressive growth in a lymph node environment. This information is of particular relevance now, in the era of sentinel lymph node biopsy, where smaller intranodal tumor deposits are being detected earlier in the course of disease, the clinical relevance of which is uncertain. In this study, we compared differentially expressed genes in cell lines of high (468LN) vs. low (468GFP) lymphatic metastatic ability, and related these to clinical literature on genes associated with lymphatic metastatic ability and prognosis, to identify genes of potential clinical relevance. This approach revealed differential expression of a set of genes associated with 'cancer stem cell-like' properties, as well as networks of genes potentially associated with survival and autonomous growth. We explored these differences functionally and found that 468LN cells have a higher proportion of cells with a cancer stem cell-like (CD44+/CD24-) phenotype, have a higher clonogenic potential and a greater ability to survive, establish and grow in a foreign (lymph node and 3D Matrigel) microenvironment, relative to 468GFP cells. Differentially expressed genes which reflect these functions provide candidates for investigation as potential targets for therapy directed against early lymphatic metastasis.