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Abstract 723: Cortical Bone Derived Stem Cells Modulates T Cell Response After Myocardial Injury

Rationale: The ischemic environment following myocardial infarction induces a harsh, pro-inflammatory state that facilitates the removal of necrotic tissue. Manipulation of T cell responses after cardiac injury directly impacts the outcome of the reparative response. Previous studies have demonstrated Cortical Bone Derived Stem Cells (CBSCs) improve cardiac function after myocardial infarction. CBSCs produce a diverse paracrine profile; whether this can lead to the modulation of T cell response by manipulating T cell subsets and allowing for increased cardiac wound healing post-MI remains unknown. Objective: To determine whether CBSCs secrete a paracrine profile that promotes wound healing by modulating T cell response following myocardial injury. Methods and Results: CD3+ T-lymphocytes were isolated from the spleen of C57BL/6J male mice ages 8 to 12 weeks via Fluorescent Activated Cell Sorting (FACs). CBSC secretome was introduced to T cell populations via a transwell culture system in the presence of CD3/28 coated beads with IL2. T-regulatory (Treg) populations were quantified via FACs 24 hours after in vitro CBSC exposure. Treg populations exposed to CBSC secretome in hypoxic (1% O2) conditions expanded to 26.9% of the CD4+ compartment, a 4.5 fold increase compared to control conditions. This result was further confirmed via qRT-PCR analysis. T cell culture secretome was quantified via proteome profiling of 111 soluble cytokine proteins 24 hours post CBSC exposure. CBSCs express a robust chemotaxis signature that promotes immune cell recruitment. T cells cultured in the presence of CBSC (pre-conditioned or co-culture conditions) exhibit increased CCL5, CCL22, CD40, CXCL1, and Lipocalin-2. C-reactive protein, a pro-inflammatory marker, was decreased by ~5.5 fold in the presence of CBSCs, promoting wound healing. Conclusions: CBSCs secrete a diverse paracrine profile that promotes Treg expansion during hypoxic conditions. Expansion of Treg populations post-CBSC exposure in the post-IR heart leads to increased inflammation resolution and improved myocardial wound healing, allowing for increased cardiac function and repressed cardiac remodeling.

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Long-Term Survival after Complete Surgical Resection and Adjuvant Immunotherapy for Distant Melanoma Metastases.

This phase III study was undertaken to evaluate the efficacy of an allogeneic whole-cell vaccine (Canvaxin™) plus bacillus Calmette-Guerin (BCG) after complete resection of stage IV melanoma. After complete resection of ≤5 distant metastases, patients were randomly assigned to BCG+Canvaxin (BCG/Cv) or BCG+placebo (BCG/Pl). The primary endpoint was overall survival (OS); secondary endpoints were disease-free survival (DFS), and immune response measured by skin test (ClinicalTrials.gov identifier: NCT00052156). Beginning in May 1998, 496 patients were randomized. In April 2005, the Data Safety Monitoring Board recommended stopping enrollment due to a low probability of efficacy. At that time, median OS and 5-year OS rate were 38.6 months and 44.9%, respectively, for BCG/Pl versus 31.4 months and 39.6% in the BCG/Cv group (hazard ratio (HR), 1.18; p=0.250). Follow-up was extended at several trial sites through March 2010. Median OS and 5-year and 10-year survival was 39.1 months, 43.3 and 33.3%, respectively, for BCG/Pl versus 34.9 months, 42.5 and 36.4%, in the BCG/Cv group (HR 1.053; p=0.696). Median DFS, 5- and 10-year DFS were 7.6 months, 23.8 and 21.7%, respectively, for BCG/Pl versus 8.5 months, 30.0%, and 30.0%, respectively, for the BCG/Cv group (HR 0.882; p=0.260). Positive DTH skin testing correlated with increased survival. In this, the largest study of postsurgical adjuvant therapy for stage IV melanoma reported to date, BCG/Cv did not improve outcomes over BCG/placebo. Favorable long-term survival among study patients suggests that metastasectomy should be considered for selected patients with stage IV melanoma.

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