Validation of contrast-enhanced magnetic resonance imaging to monitor regenerative efficacy after cell therapy in a porcine model of convalescent myocardial infarction.

BACKGROUND: Magnetic resonance imaging (MRI) in the CArdiosphere-Derived aUtologous stem CElls to reverse ventricUlar dySfunction (CADUCEUS) trial revealed that cardiosphere-derived cells (CDCs) decrease scar size and increase viable myocardium after myocardial infarction (MI), but MRI has not been validated as an index of regeneration after cell therapy. We tested the validity of contrast-enhanced MRI in quantifying scarred and viable myocardium after cell therapy in a porcine model of convalescent MI. METHODS AND RESULTS: Yucatan minipigs underwent induction of MI and 2-3 weeks later were randomized to receive intracoronary infusion of 12.5×10(6) mismatched allogeneic CDCs or vehicle. Allogeneic CDCs induced mild local mononuclear infiltration but no systemic immunogenicity. MRI revealed that allogeneic CDCs attenuated remodeling, improved global and regional function, decreased scar size, and increased viable myocardium compared with placebo 2 months post-treatment. Extensive histological analysis validated quantitatively the MRI measurements of scar size, scar mass, and viable mass. CDCs neither altered gadolinium contrast myocardial kinetics nor induced changes in vascular density or architecture in viable and scarred myocardium. Histology demonstrated that CDCs lead to cardiomyocyte hyperplasia in the border zone, consistent with the observed stimulation of endogenous regenerative mechanisms (cardiomyocyte cycling, upregulation of endogenous progenitors, angiogenesis). CONCLUSIONS: Contrast-enhanced MRI accurately measures scarred and viable myocardium after cell therapy in a porcine model of convalescent MI. MRI represents a useful tool for assessing dynamic changes in the infarct and monitoring regenerative efficacy.

Targeted degradation of the AML1/MDS1/EVI1 oncoprotein by arsenic trioxide.

Arsenic trioxide (ATO) has been found to be an effective treatment for acute promyelocytic leukemia patients and is being tested for treating other hematologic malignancies. We have previously shown that AML1/MDS1/EVI1 (AME), a fusion gene generated by a t(3;21)(q26;q22) translocation found in patients with chronic myelogenous leukemia during blast phase, myelodysplastic syndrome, or acute myelogenous leukemia (AML), impairs hematopoiesis and eventually induces an AML in mice. Both fusion partners of AME, AML1 and MDS1/EVI1, encode transcription factors and are also targets of a variety of genetic abnormalities in human hematologic malignancies. In addition, aberrant expression of ectopic viral integration site 1 (EVI1) has also been found in solid tumors, such as ovarian and colon cancers. In this study, we examined whether ATO could target AME and related oncoproteins. We found that ATO used at therapeutic levels degrades AME. The ATO treatment induces differentiation and apoptosis in AME leukemic cells in vitro as well as reduces tumor load and increases the survival of mice transplanted with these cells. We further found that ATO targets AME via both myelodysplastic syndrome 1 (MDS1) and EVI1 moieties and degrades EVI1 via the ubiquitin-proteasome pathway and MDS1 in a proteasome-independent manner. Our results suggest that ATO could be used as a part of targeted therapy for AME-, AML1/MDS1-, MDS1/EVI1-, and EVI1-positive human cancers.

Human cardiosphere-derived cells from advanced heart failure patients exhibit augmented functional potency in myocardial repair.

OBJECTIVES: This study sought to compare the regenerative potency of cells derived from healthy and diseased human hearts. BACKGROUND: Results from pre-clinical studies and the CADUCEUS (CArdiosphere-Derived aUtologous stem CElls to reverse ventricUlar dySfunction) trial support the notion that cardiosphere-derived cells (CDCs) from normal and recently infarcted hearts are capable of regenerating healthy heart tissue after myocardial infarction (MI). It is unknown whether CDCs derived from advanced heart failure (HF) patients retain the same regenerative potency. METHODS: In a mouse model of acute MI, we compared the regenerative potential and functional benefits of CDCs derived from 3 groups: 1) non-failing (NF) donor: healthy donor hearts post-transplantation; 2) MI: patients who had an MI 9 to 35 days before biopsy; and 3) HF: advanced cardiomyopathy tissue explanted at cardiac transplantation. RESULTS: Cell growth and phenotype were identical in all 3 groups. Injection of HF CDCs led to the greatest therapeutic benefit in mice, with the highest left ventricular ejection fraction, thickest infarct wall, most viable tissue, and least scar 3 weeks after treatment. In vitro assays revealed that HF CDCs secreted higher levels of stromal cell-derived factor (SDF)-1, which may contribute to the cells' augmented resistance to oxidative stress, enhanced angiogenesis, and improved myocyte survival. Histological analysis indicated that HF CDCs engrafted better, recruited more endogenous stem cells, and induced greater angiogenesis and cardiomyocyte cell-cycle re-entry. CDC-secreted SDF-1 levels correlated with decreases in scar mass over time in CADUCEUS patients treated with autologous CDCs. CONCLUSIONS: CDCs from advanced HF patients exhibit augmented potency in ameliorating ventricular dysfunction post-MI, possibly through SDF-1­mediated mechanisms.

Functional performance of human cardiosphere-derived cells delivered in an in situ polymerizable hyaluronan-gelatin hydrogel.

The vast majority of cells delivered into the heart by conventional means are lost within the first 24 h. Methods are needed to enhance cell retention, so as to minimize loss of precious material and maximize effectiveness of the therapy. We tested a cell-hydrogel delivery strategy. Cardiosphere-derived cells (CDCs) were grown from adult human cardiac biopsy specimens. In situ polymerizable hydrogels made of hyaluronan and porcine gelatin (Hystem(®)-C™) were formulated as a liquid at room temperature so as to gel within 20 min at 37 °C. CDC viability and migration were not compromised in Hystem-C™. Myocardial infarction was created in SCID mice and CDCs were injected intramyocardially in the infarct border zone. Real-time PCR revealed engraftment of CDCs delivered in Hystem-C™ was increased by nearly an order of magnitude. LVEF (left ventricular ejection fraction) deteriorated in the control (PBS only) group over the 3-week time course. Hystem-C™ alone or CDCs alone preserved LVEF relative to baseline, while CDCs delivered in Hystem-C™ resulted in a sizable boost in LVEF. Heart morphometry revealed the greatest attenuation of LV remodeling in the CDC + Hystem-C™ group. Histological analysis suggested cardiovascular differentiation of the CDCs in Hystem-C™. However, the majority of functional benefit is likely from paracrine mechanisms such as tissue preservation and neovascularization. A CDC/hydrogel formulation suitable for catheter-based intramyocardial injection exhibits superior engraftment and functional benefits relative to naked CDCs.

Direct comparison of different stem cell types and subpopulations reveals superior paracrine potency and myocardial repair efficacy with cardiosphere-derived cells.

OBJECTIVES: The goal of this study was to conduct a direct head-to-head comparison of different stem cell types in vitro for various assays of potency and in vivo for functional myocardial repair in the same mouse model of myocardial infarction. BACKGROUND: Adult stem cells of diverse origins (e.g., bone marrow, fat, heart) and antigenic identity have been studied for repair of the damaged heart, but the relative utility of the various cell types remains unclear. METHODS: Human cardiosphere-derived cells (CDCs), bone marrow-derived mesenchymal stem cells, adipose tissue-derived mesenchymal stem cells, and bone marrow mononuclear cells were compared. RESULTS: CDCs revealed a distinctive phenotype with uniform expression of CD105, partial expression of c-kit and CD90, and negligible expression of hematopoietic markers. In vitro, CDCs showed the greatest myogenic differentiation potency, highest angiogenic potential, and relatively high production of various angiogenic and antiapoptotic-secreted factors. In vivo, injection of CDCs into the infarcted mouse hearts resulted in superior improvement of cardiac function, the highest cell engraftment and myogenic differentiation rates, and the least-abnormal heart morphology 3 weeks after treatment. CDC-treated hearts also exhibited the lowest number of apoptotic cells. The c-kit(+) subpopulation purified from CDCs produced lower levels of paracrine factors and inferior functional benefit when compared with unsorted CDCs. To validate the comparison of cells from various human donors, selected results were confirmed in cells of different types derived from individual rats. CONCLUSIONS: CDCs exhibited a balanced profile of paracrine factor production and, among various comparator cell types/subpopulations, provided the greatest functional benefit in experimental myocardial infarction.

Dual in vivo quantification of integrin-targeted and protease-activated agents in cancer using fluorescence molecular tomography (FMT).

PURPOSE: Integrins, especially α(v)ß(3) and α(v)ß(5), are upregulated in tumor cells and activated endothelial cells and as such, serve as cancer biomarkers. We developed a novel near-infrared-labeled optical agent for the in vivo detection and quantification of α(v)ß(3)/α(v)ß(5). PROCEDURES: A small peptidomimetic α(v)ß(3) antagonist was synthesized, coupled to a near-infrared fluorescent (NIRF) dye, and tested for binding specificity using integrin-overexpressing cells, inhibition of vitronectin-mediated cell attachment, binding to tumor and endothelial cells in vitro, and competition studies. Pharmacokinetics, biodistribution, specificity of tumor targeting, and the effect of an antiangiogenic treatment were assessed in vivo. RESULTS: The integrin NIRF agent showed strong selectivity towards α(v)ß(3/)α(v)ß(5) in vitro and predominant tumor distribution in vivo, allowing noninvasive and real-time quantification of integrin signal in tumors. Antiangiogenic treatment significantly inhibited integrin signal in vivo but had no effect on a cathepsin-cleavable NIR agent. Simultaneous imaging revealed different patterns of distribution reflecting the underlying differences in integrin and cathepsin biology during tumor progression. CONCLUSIONS: NIRF-labeled integrin antagonists allow noninvasive molecular fluorescent imaging and quantification of tumors in vivo, improving and providing more refined approaches for cancer detection and treatment monitoring.