Efficacy and Safety of Intensity-Modulated Radiotherapy Following Transarterial Chemoembolization in Patients With Unresectable Hepatocellular Carcinoma.

Three-dimensional conformal radiotherapy in combination with transarterial chemoembolization (TACE) has been beneficial in patients with unresectable hepatocellular carcinoma (HCC). There have been few clinical reports on the use of intensity-modulated radiotherapy (IMRT) in combination with TACE for these patients. The purpose of this study was to assess the efficacy and toxicity of IMRT following TACE in unresectable HCC.The medical records of consecutive patients with unresectable HCC, who underwent IMRT following TACE from January 2009 to June 2014, were retrospectively reviewed in order to assess the overall survival (OS), progression-free survival (PFS), tumor response, and treatment-associated toxicity.A total of 64 lesions in 54 patients were included in the analysis. IMRT was delivered at a median dose of 50 Gy (range 44-70 Gy) at 1.8 to 2.0 Gy per fraction. The overall response rate was achieved in 64.8% of patients with complete response in 20.4% of patients at 3 months after completion of IMRT. The median OS was 20.2 months (95% CI = 8.6-31.9), and the actuarial 1-, 2-, and 3-year OS rates were 84.6%, 49.7%, and 36.7%, respectively. The median PFS was 10.5 months (95% CI = 7.3-13.7) and the 1-, 2-, and 3-year PFS rates were 44.2%, 23.4%, and 14.6%, respectively. The responders had a significantly higher OS rate than the nonresponders (3-year OS 48.0% vs 14.4%, P = 0.001). During and the first month following IMRT, 10 (18.5%) patients developed grade 3 hematological toxicity, and 3 (5.6%) developed grade 3 hepatic toxicity. No patient experienced grade 4 or 5 toxicity. Radiation-induced liver disease was not observed.Our findings suggest that IMRT following TACE could be a favorable treatment option for both its safety profile and clinical benefit in patients with unresectable HCC.

Targeting CD44 augments the efficacy of Tregs in autoimmune diabetes.

Curing type 1 diabetes (T1D) will require lasting control of the autoimmune response that destroys insulin-producing islet ß-cells. Re-establishing tolerance by restoring/replacing Tregs has significant potential for treatment of T1D but will require strategies to augment and maintain their efficacy. We previously showed that polyclonal in vitro-induced Tregs can reverse recent onset of T1D in ∼ 50% of NOD mice. Here we report that treatment of newly hyperglycemic animals with a short course of anti-CD44 at the time of Treg transfer improved diabetes reversal to >90%. Anti-CD44 treatment alone delayed diabetes onset and increased the frequencies of pancreatic CD4(+) T cells producing IL-2 or TGF-ß, cytokines that support Treg function and survival, without altering production of IFN-γ. These anti-CD44 effects on endogenous T cells were also observed in the context of polyclonal Treg transfer, and the combination treatment also reduced pancreatic infiltrates. The results provide compelling evidence that approaches to modulate the pancreatic milieu to support Treg function and counteract inflammation in the pancreas can greatly enhance the efficacy of adoptively transferred Tregs, and suggest that approaches achieving these outcomes hold promise for long-term control of autoimmunity in T1D.

Islet antigen-specific Th17 cells can induce TNF-α-dependent autoimmune diabetes.

Type 1 diabetes (T1D) results from autoimmune destruction of pancreatic ß-cells. Although Th1 cells are key orchestrators of T1D, the function(s) of the more recently identified Th17 subset are unclear due to inherent plasticity. In this study, we analyzed Th17 cells for stability and diabetogenicity in NOD mice. We found that like Th1 cells, Th17 are a distinct population throughout the prediabetic phase. At diabetes onset, there were marked increases in IL-17-producing Th17 cells and IFN-γ-producing Th1 cells in the pancreas as well as in the serum levels of these cytokines, indicating that these proinflammatory mediators serve as biomarkers of advanced autoimmunity. Although naturally occurring Th17 cells in diabetic mice did not contribute to diabetes development in transfer models, islet-specific Th17 cells were diabetogenic independently of IL-17 and displayed inflammation-induced Th17-to-Th1 reprogramming that could be elicited by Th1 cells. However, an inability to generate Th1 cells because of Stat4, Ifngr, and Ifng deficiencies did not prevent diabetes. Instead, TNF-α could mediate diabetes in response to either Th17 cells or Th1 cells. The results identify a previously unknown mechanism by which Th17 cells can contribute to T1D. Our studies also suggest that when developing interventions for T1D, it will be potentially advantageous to focus on mechanisms common to effector T cells rather than on the signature cytokines of various subsets.

Harnessing memory adaptive regulatory T cells to control autoimmunity in type 1 diabetes.

Type 1 diabetes (T1D) results from autoimmune destruction of insulin-producing ß-cells in the pancreatic islets. There is an immediate need to restore both ß-cell function and immune tolerance to control disease progression and ultimately cure T1D. Currently, there is no effective treatment strategy to restore glucose regulation in patients with T1D. FoxP3-expressing CD4(+) regulatory T cells (Tregs) are potential candidates to control autoimmunity because they play a central role in maintaining self-tolerance. However, deficiencies in either naturally occurring Tregs (nTregs) themselves and/or their ability to control pathogenic effector T cells have been associated with T1D. Here, we hypothesize that nTregs can be replaced by FoxP3(+) adaptive Tregs (aTregs), which are uniquely equipped to combat autoreactivity in T1D. Unlike nTregs, aTregs are stable and provide long-lived protection. In this review, we summarize the current understanding of aTregs and their potential for use as an immunological intervention to treat T1D.

IL-7 uniquely maintains FoxP3(+) adaptive Treg cells that reverse diabetes in NOD mice via integrin-ß7-dependent localization.

Type 1 diabetes (T1D) develops as a consequence of a progressive autoimmune response that destroys insulin-producing ß-cells in pancreatic islets. Because of their role(s) in controlling immune responses, considerable effort has been directed toward resolving whether regulatory T cells (Tregs) offer a clinical treatment to restore tolerance in T1D. We previously reported that in vitro-induced adaptive Treg cells (aTregs) can reverse T1D and persist as protective memory cells in the NOD mouse model. In the current study, we investigated mechanisms that regulate aTregs. We found that these FoxP3(+) aTregs expressed high levels of the IL-7 receptor, IL-7Rα, without the high affinity receptor for IL-2, CD25, which is found on natural Treg cells (nTregs). IL-7Rα expression was mirrored by the dependency of aTregs on IL-7 for persistence. IL-10 and TGF-ß, effector cytokines of aTregs, were not essential for their maintenance at the level of systemic antibody blocking. Nevertheless, IL-10 modulated cytokine production by aTregs and TGF-ß was critical for protection. aTregs were found to infiltrate islets and the expression of integrin-ß7 was required for their localization in the pancreas. Furthermore, blocking aTreg entry into the pancreas prevented their control of diabetogenic effector T cells, implying the need for local control of the autoimmune response. The distinct homeostatic regulation of aTregs independently of a response to IL-2, which is defective in T1D patients, suggests that these cells represent a translatable candidate to control the autoimmune response.

CD44 regulates survival and memory development in Th1 cells.

Optimal immunity to microorganisms depends upon the regulated death of clonally expanded effector cells and the survival of a cohort of cells that become memory cells. After activation of naive T cells, CD44, a widely expressed receptor for extracellular matrix components, is upregulated. High expression of CD44 remains on memory cells and despite its wide usage as a "memory marker," its function is unknown. Here we report that CD44 was essential for the generation of memory T helper 1 (Th1) cells by promoting effector cell survival. This dependency was not found in Th2, Th17, or CD8(+) T cells despite similar expression of CD44 and the absence of splice variants in all subsets. CD44 limited Fas-mediated death in Th1 cells and its ligation engaged the phosphoinositide 3-kinase-Akt kinase signaling pathway that regulates cell survival. The difference in CD44-regulated apoptosis resistance in T cell subpopulations has important implications in a broad spectrum of diseases.

Multifaceted regulation of T cells by CD44.

CD44 is a widely-expressed adhesion receptor that is associated with diverse biological processes involving migrating cells, including inflammation, angiogenesis, bone metabolism and wound healing. In the immune system, CD44 is upregulated after activation of naive T lymphocytes during their responses against invading microbes. Once an infection is cleared, elevated levels of CD44 remain on the surface of memory T cells that mediate protection against re-infection. While this has led to the use of highly sustained CD44 expression on T cells as an indicator of a previous immune response, the relevance to T-cell responses or homeostasis has been largely unexplored. Our recent studies demonstrate that CD44 selectively regulates the survival of the Th1 subset of CD4 T cells, but not other T-cell subpopulations. These findings, together with studies of CD44 in other cell types, suggest that differences in the engagement of signaling mechanisms are likely to underlie differential regulation of T-cell responses and underscore the importance of this adhesion receptor to immune cell regulation and protection against viruses and intracellular bacteria.

Quantitative analysis of T cell homeostatic proliferation.

T cell homeostatic proliferation occurs on transfer of T cells into lymphopenic recipients; transferred cells undergo several rounds of division in the absence of specific antigen stimulation. For a quantitative analysis of this phenomenon, we applied a mathematical method to describe proliferating T cells to match peak distributions from actual CFSE dilution data. For in vitro stimulation of T cells with anti-CD3/anti-CD28, our simulation confirmed a high proportion of cells entering cell cycle with a low proportion undergoing apoptosis. When applied to homeostatic proliferation, it described striking differences in CD4 and CD8 T cell proliferation rates, and accurately predicted that successive divisions were accompanied by higher rates of apoptosis, limiting the accumulation of proliferating cells. Thus, the presence of multiple CFSE dilution peaks cannot be considered equivalent to lymphocyte expansion. Finally, genetic effects were identified that may help explain links between homeostatic proliferation and autoimmunity.

Itk is not essential for CD28 signaling in naive T cells.

Itk, a member of the Tec family of tyrosine kinases, is critical for TCR signaling, leading to the activation of phospholipase C gamma1. Early biochemical studies performed in tumor cell lines also implicated Itk in CD28 signaling. These data were complemented by functional studies on primary Itk-/- T cells that suggested a negative role for Itk in CD28 signaling. In this report, we describe a thorough analysis of CD28-mediated responses in T cells lacking Itk. Using purified naive CD4+ T cells from Itk-/- mice, we examine a range of responses dependent on CD28 costimulation. We also analyze Akt and glycogen synthase kinase-3beta phosphorylation in response to stimulation of CD28 alone. Overall, these experiments demonstrate that CD28 signaling, as well as CD28-mediated costimulation of TCR signaling, function efficiently in the absence of Itk. These findings indicate that Itk is not essential for CD28 signaling in primary naive CD4+ T cells.