The Kv4.3-encoded current (I:(Kv4.3)) has been identified as the major component of the voltage-dependent Ca(2+)-independent transient outward current (I:(to1)) in human and canine ventricular cells. Experimental evidence supports a correlation between I:(to1) density and prominence of the phase 1 notch; however, the role of I:(to1) in modulating action potential duration (APD) remains unclear. To help resolve this role, Markov state models of the human and canine Kv4.3- and Kv1.4-encoded currents at 35 degrees C are developed on the basis of experimental measurements. A model of canine I:(to1) is formulated as the combination of these Kv4.3 and Kv1.4 currents and is incorporated into an existing canine ventricular myocyte model. Simulations demonstrate strong coupling between L-type Ca(2+) current and I:(Kv4.3) and predict a bimodal relationship between I:(Kv4.3) density and APD whereby perturbations in I:(Kv4.3) density may produce either prolongation or shortening of APD, depending on baseline I:(to1) current level.

Immune checkpoint therapy has revolutionized cancer treatment by blocking inhibitory pathways in T cells that limits the an effective anti-tumor immune response. Therapeutics targeting CTLA-4 and PD1/PDL1 have progressed to first line therapy in multiple tumor types with some patients exhibiting tumor regression or remission. However, the majority of patients do not benefit from checkpoint therapy emphasizing the need for alternative therapeutic options. Lymphocyte Activation Gene 3 (LAG3) or CD223 is expressed on multiple cell types including CD4 + and CD8 + T cells, and T regs , and is required for optimal T cell regulation and homeostasis. Persistent antigen-stimulation in cancer or chronic infection leads to chronic LAG3 expression, promoting T cell exhaustion. Targeting LAG3 along with PD1 facilitates T cell reinvigoration. A substantial amount of pre-clinical data and mechanistic analysis has led to LAG3 being the third checkpoint to be targeted in the clinic with nearly a dozen therapeutics under investigation. In this review, we will discuss the structure, function and role of LAG3 in murine and human models of disease, including autoimmune and inflammatory diseases, chronic viral and parasitic infections, and cancer, emphasizing new advances in the development of LAG3-targeting immunotherapies for cancer that are currently in clinical trials.

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Immunotherapy using autologous T-cells has emerged to be a powerful treatment option for patients with metastatic melanoma. These include the adoptive transfer of autologous tumor-infiltrating lymphocytes (TIL), T-cells transduced with high-affinity T-cell receptors (TCR) against major melanosomal tumor antigens, and T cells transduced with chimeric antigen receptors (CAR) composed of hybrid immunoglobulin light chains with endo-domains of T-cell signaling molecules. Among these and other options for T-cell therapy, TIL together with high-dose IL-2 has had the longest clinical history with multiple clinical trials in centers across the world consistently demonstrating durable clinical response rates near 50% or more. A distinct advantage of TIL therapy making it still the T-cell therapy of choice is the broad nature of the T-cell recognition against both defined as well as un-defined tumors antigens against all possible MHC, rather than the single specificity and limited MHC coverage of the newer TCR and CAR transduction technologies. In the past decade, significant inroads have been made in defining the phenotypes of T cells in TIL mediating tumor regression. CD8+ T cells are emerging to be critical, although the exact subset of CD8+ T cells exhibiting the highest clinical activity in terms of memory and effector markers is still controversial. We present a model in which both effector-memory and more differentiated effector T cells ultimately may need to cooperate to mediate long-term tumor control in responding patients. Although TIL therapy has shown great potential to treat metastatic melanoma, a number of issues have emerged that need to be addressed to bring it more into the mainstream of melanoma care. First, we have a reached the point where a pivotal phase II or phase III trials are needed in an attempt to gain regulatory approval of TIL as standard-of-care. Second, improvements in how we expand TIL for therapy are needed, that minimize the time the T-cells are in culture and improve the memory and effector characteristics of the T cells for longer persistence and enhanced anti-tumor activity in vivo. Third, there is a critical need to identify surrogate and predictive biomarkers in order to better select suitable patients for TIL therapy in order to improve response rate and duration. Overall, the outlook for TIL therapy for melanoma is very bright. We predict that TIL will indeed emerge to become an approved treatment in the upcoming years through pivotal clinical trials. Moreover, new approaches combining TIL with targeted signaling pathway drugs, such as mutant B-RAF inhibitors, and synergistic immunomodulatory interventions enhancing T-cell costimulation and preventing negative regulation, should further increase therapeutic efficacy and durable complete response rates.

Adoptive T-cell therapy (ACT) using expanded tumor-infiltrating lymphocytes (TIL) with high-dose IL-2 is a promising form of immunotherapy for Stage IV melanoma having clinical response rates of 50% or more. One of the major problems preventing further success of this therapy is that the current protocols used to highly expand TIL for infusion drive CD8+ T cells to differentiate into effector cells losing key co-stimulatory molecules such as CD28 and CD27. This has been associated with a lack of persistence in vivo for reasons not entirely clear. In this study, we demonstrate that while human melanoma CD8+ TIL lost CD27 and CD28 expression during the rapid expansion for ACT, they gained expression of the alternative co-stimulatory molecule CD137/4-1BB, and to a lesser extent CD134/OX40. Post-REP TIL were found to be highly sensitive to activation-induced cell death (AICD) when re-activated through the TCR with low levels of OKT3 antibody. However, co-ligation of 4-1BB using two different agonistic anti-4-1BB antibodies potently prevented AICD of post-REP CD8+ TIL, including those specific for MART-1, and facilitated even further cell expansion. This was correlated with increased levels of bcl-2 and bcl-xL together with decreased bim expression. 4-1BB-co-stimulated post-REP TIL also expressed increased levels of the cytolytic granule proteins and exhibited enhanced CTL activity against melanoma cells. Lastly, post-REP CD8+ TIL were protected from cell death by anti-4-1BB ligation when exposed to HLA-matched melanoma cells. Our results indicate that 4-1BB co-stimulation may significantly improve TIL survival during melanoma ACT and boost anti-tumor cytolytic activity.

Background-This study sought to define the technique and results of magnetic resonance imaging (MRI) of pulmonary vein (PV) anatomy before and after catheter ablation of atrial fibrillation (AF). Methods and Results-Twenty-eight patients with AF underwent ablation. Patients underwent gadolinium-enhanced MRI before and 6 weeks after their procedures. A control group of 27 patients also underwent MRI. Variant PV anatomy was observed in 38% of patients. AF patients had larger PV diameters than control subjects, but no difference was observed in the size of the PV ostia among AF patients. The PV ostia were oblong in shape with an anteroposterior dimension less than the superoinferior dimension. The left PVs had a longer "neck" than the right PVs. A detectable PV narrowing was observed in 24% of veins. The severity of stenosis was severe in 1 vein (1.4%), moderate in 1 vein (1.4%), and mild in 15 veins (21.1%). All patients were asymptomatic, and none required treatment. Conclusions-This study demonstrates that AF patient have larger PVs than control subjects and demonstrates the value of MRI in facilitating AF ablation. The benefits of preprocedural MRI of PVs include the ability to evaluate the number, size, and shape of the PVs. MRI also provides an assessment of the severity of PV stenosis.

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Background-The transient outward potassium current (I to ) encoded by the Kv4 family of potassium channels is important in the repolarization of cardiac myocytes. KChIPs are a recently identified group of Ca 2ϩ -binding accessory subunits that modulate Kv4-encoded currents. KChIP2 is the only family member expressed in the heart. Methods and Results-We previously cloned 2 novel splice variants of KChIP2 from human heart, named KChIP2S and KChIP2T. The transmural distribution of KChIP2 mRNA and protein in human and canine left ventricle was examined using kinetic RT-PCR and Western blots in the same tissues. A steep gradient of mRNA with greater KChIP2 expression in the epicardium was observed. However, no gradient of immunoreactive protein was observed. Immunocytochemistry reveals KChIP2 expression in the t-tubules and the nucleus. The predominant effects of all 3 KChIP2 splice variants on hKv4.3-encoded current are to increase the density, slow the current decay in a Ca 2ϩ -dependent manner, and hasten recovery from inactivation in a splice variant-specific fashion. Conclusions-A family of KChIP2 proteins is expressed in human hearts that exhibits differential modulation of hKv4.3 current in a Ca 2ϩ -dependent fashion. The effect of KChIP2 on the biophysical properties of expressed Kv4.3 current and the absence of a gradient of protein across the ventricular wall suggest that KChIP2 is either not a requisite component of human or canine ventricular I to or that its functional effect is being affected or additionally modified by other factors present in myocardial cells.