Identifying Indications for Neoadjuvant Therapy in Cholangiocarcinoma.

The recent Hot Topics section focuses on survival rates for patients with cholangiocarcinoma.

Activated T cells secrete an alternatively spliced form of common γ-chain that inhibits cytokine signaling and exacerbates inflammation.

The common γ-chain (γc) plays a central role in signaling by IL-2 and other γc-dependent cytokines. Here we report that activated T cells produce an alternatively spliced form of γc mRNA that results in protein expression and secretion of the γc extracellular domain. The soluble form of γc (sγc) is present in serum and directly binds to IL-2Rß and IL-7Rα proteins on T cells to inhibit cytokine signaling and promote inflammation. sγc suppressed IL-7 signaling to impair naive T cell survival during homeostasis and exacerbated Th17-cell-mediated inflammation by inhibiting IL-2 signaling upon T cell activation. Reciprocally, the severity of Th17-cell-mediated inflammatory diseases was markedly diminished in mice lacking sγc. Thus, sγc expression is a naturally occurring immunomodulator that regulates γc cytokine signaling and controls T cell activation and differentiation.

The Abundance and Availability of Cytokine Receptor IL-2Rß (CD122) Constrain the Lymphopenia-Induced Homeostatic Proliferation of Naive CD4 T Cells.

Lymphopenia-induced homeostatic proliferation (LIP) is a critical mechanism for restoring T cell immunity upon lymphodepleting insults or infections. LIP is primarily driven by homeostatic cytokines, such as IL-7 and IL-15, but not all T cells respond with the same efficiency to homeostatic proliferative cues. Although CD8 T cells vigorously proliferate under lymphopenic conditions, naive CD4 T cells are substantially impaired in their response to homeostatic cytokines, and they fail to fully expand. In this study, we show that the availability of IL-2Rß (CD122), which is a receptor subunit shared by IL-2 and IL-15, affects both the cytokine responsiveness and the LIP of naive CD4 T cells in the mouse. The enumeration of surface IL-2Rß molecules on murine naive CD4 and naive CD8 T cells revealed a 5-fold difference in IL-2Rß abundance. Notably, it was the limited availability of IL-2Rß that impaired CD4 T cell responsiveness to IL-15 and suppressed their LIP. As such, forced IL-2Rß expression on CD4 T cells by transgenesis bestowed IL-15 responsiveness onto naive CD4 T cells, which thus acquired the ability to undergo robust LIP. Collectively, these results identify IL-2Rß availability as a new regulatory mechanism to control cytokine responsiveness and the homeostatic proliferation of murine CD4 T cells.

The molecular basis and cellular effects of distinct CD103 expression on CD4 and CD8 T cells.

Integrin CD103 mediates the adhesion and tissue retention of T cells by binding to E-cadherin which is abundant on epithelial cells. Notably, CD103 is highly expressed on CD8 T cells but conspicuously absent on most CD4 T cells. The mechanism controlling such lineage-specific expression of CD103 remains unclear. Using a series of genetically engineered mouse models, here, we demonstrate that the regulatory mechanism of CD103 expression is distinct between CD4 and CD8 T cells, and that the transcription factor Runx3 plays an important but not an essential role in this process. We further found that the availability of integrin ß7 which heterodimerizes with CD103 was necessary but also constrained the surface expression of CD103. Notably, the forced surface expression of CD103 did not significantly alter the thymic development of conventional T cells but severely impaired the generation of MHC-II-restricted TCR transgenic T cells, revealing previously unappreciated aspects of CD103 in the selection and maturation of CD4 T cells. Unlike its effect on CD4 T cell development, however, CD103 overexpression did not significantly affect CD4 T cells in peripheral tissues. Moreover, the frequency and number of CD4 T cells in the small intestine epithelium did not increase even though E-cadherin is highly expressed in this tissue. Collectively, these results suggest that most mature CD4 T cells are refractory to the effects of CD103 expression, and that they presumably utilize CD103-independent pathways to control their tissue retention and residency.

SOCS3 is a suppressor of γc cytokine signaling and constrains generation of murine Foxp3+ regulatory T cells.

SOCS3 is a cytosolic inhibitor of cytokine signaling that suppresses the activation of cytokine receptor-associated JAK kinases. Mechanistically, SOCS3 is recruited to a site in the cytokine receptors known as the SOCS3-interaction motif, and then binds JAK molecules to inhibit their kinase activity. The SOCS3-interaction motif is found in receptors of the gp130 cytokine family but mostly absent from other cytokine receptors, including γc. Thus, SOCS3 has been considered a selective suppressor of gp130 family cytokines, but not γc cytokines. Considering that γc signaling induces SOCS3 expression in T cells, here we revisited the role of SOCS3 on γc signaling. Using SOCS3 transgenic mice, we found that increased abundance of SOCS3 not only suppressed signaling of the gp130 family cytokine IL-6, but also signaling of the γc family cytokine IL-7. Consequently, SOCS3 transgenic mice were impaired in IL-7-dependent T cell development in the thymus and the homeostasis of mature T cells in peripheral tissues. Moreover, enforced SOCS3 expression interfered with the generation of Foxp3+ regulatory T cells that requires signaling by the γc family cytokine IL-2. Collectively, we report an underappreciated role for SOCS3 in suppressing γc cytokine signaling, effectively expanding its scope of target cytokines in T cell immunity.

Ikaros is required to survive positive selection and to maintain clonal diversity during T-cell development in the thymus.

The zinc-finger protein Ikaros is a key player in T-cell development and a potent tumor suppressor in thymocytes. To understand the molecular basis of its function, we disabled Ikaros activity in vivo using a dominant negative Ikaros transgene (DN-IkTg). In DN-IkTg mice, T-cell development was severely suppressed, and positively selected thymocytes clonally expanded, resulting in a small thymus with a heavily skewed T-cell receptor (TCR) repertoire. Notably, DN-IkTg induced vigorous proliferation concomitant to downregulation of antiapoptotic factor expression such as Bcl2. Ikaros activity was required during positive selection, and specifically at the CD4(+)CD8(lo) intermediate stage of thymocyte differentiation, where it prevented persistent TCR signals from inducing aberrant proliferation and expansion. In particular, DN-IkTg induced the accumulation of CD4 single-positive (SP) thymocytes with a developmentally transitional phenotype, and it imposed a developmental arrest accompanied by massive apoptosis. Thus, we identified an in vivo requirement for Ikaros function, which is to suppress the proliferative potential of persistent TCR signals and to promote the survival and differentiation of positively selected thymocytes.

Pim1 permits generation and survival of CD4+ T cells in the absence of γc cytokine receptor signaling.

γ-Chain (γc) cytokine receptor signaling is required for the development of all lymphocytes. Why γc signaling plays such an essential role is not fully understood, but induction of the serine/threonine kinase Pim1 is considered a major downstream event of γc as Pim1 prevents apoptosis and increases metabolic activity. Consequently, we asked whether Pim1 overexpression would suffice to restore lymphocyte development in γc-deficient mice. By analyzing Pim1-transgenic γc-deficient mice (Pim1(Tg) γc(KO) ), we show that Pim1 promoted T-cell development and survival in the absence of γc. Interestingly, such effects were largely limited to CD4(+) lineage αß T cells as CD4(+) T-cell numbers improved to near normal levels but CD8(+) T cells remained severely lymphopenic. Notably, Pim1 over-expression failed to promote development and survival of any T-lineage cells other than αß T cells, as we observed complete lack of γδ, NKT, FoxP3(+) T regulatory cells and TCR-ß(+) CD8αα IELs in Pim1(Tg) γc(KO) mice. Collectively, these results uncover distinct requirements for γc signaling between CD4(+) αß T cells and all other T-lineage cells, and they identify Pim1 as a novel effector molecule sufficient to drive CD4(+) αß T-cell development and survival in the absence of γc cytokine receptor signaling.

An in vivo IL-7 requirement for peripheral Foxp3+ regulatory T cell homeostasis.

All T cells are dependent on IL-7 for their development and for homeostasis. Foxp3(+) regulatory T cells (Tregs) are unique among T cells in that they are dependent on IL-2. Whether such IL-2 dependency is distinct from or in addition to an IL-7 requirement has been a confounding issue, particularly because of the absence of an adequate experimental system to address this question. In this study, we present a novel in vivo mouse model where IL-2 expression is intact but IL-7 expression was geographically limited to the thymus. Consequently, IL-7 is not available in peripheral tissues. Such mice were generated by introducing a thymocyte-specific IL-7 transgene onto an IL-7 null background. In these mice, T cell development in the thymus, including Foxp3(+) Treg numbers, was completely restored, which correlates with the thymus-specific expression of transgenic IL-7. In peripheral cells, however, IL-7 expression was terminated, which resulted in a general paucity of T cells and a dramatic reduction of Foxp3(+) Treg numbers. Loss of Tregs was further accompanied by a significant reduction in Foxp3(+) expression levels. These data suggest that peripheral IL-7 is not only necessary for Treg survival but also for upregulating Foxp3 expression. Collectively, we assessed the effect of a selective peripheral IL-7 deficiency in the presence of a fully functional thymus, and we document a critical requirement for in vivo IL-7 in T cell maintenance and specifically in Foxp3(+) cell homeostasis.

CD8 lineage-specific regulation of interleukin-7 receptor expression by the transcriptional repressor Gfi1.

Interleukin-7 receptor α (IL-7Rα) is essential for T cell survival and differentiation. Glucocorticoids are potent enhancers of IL-7Rα expression with diverse roles in T cell biology. Here we identify the transcriptional repressor, growth factor independent-1 (Gfi1), as a novel intermediary in glucocorticoid-induced IL-7Rα up-regulation. We found Gfi1 to be a major inhibitory target of dexamethasone by microarray expression profiling of 3B4.15 T-hybridoma cells. Concordantly, retroviral transduction of Gfi1 significantly blunted IL-7Rα up-regulation by dexamethasone. To further assess the role of Gfi1 in vivo, we generated bacterial artificial chromosome (BAC) transgenic mice, in which a modified Il7r locus expresses GFP to report Il7r gene transcription. By introducing this BAC reporter transgene into either Gfi1-deficient or Gfi1-transgenic mice, we document in vivo that IL-7Rα transcription is up-regulated in the absence of Gfi1 and down-regulated when Gfi1 is overexpressed. Strikingly, the in vivo regulatory role of Gfi1 was specific for CD8(+), and not CD4(+) T cells or immature thymocytes. These results identify Gfi1 as a specific transcriptional repressor of the Il7r gene in CD8 T lymphocytes in vivo.

Interleukin-6 expands homeostatic space for peripheral T cells.

T cell homeostasis and survival is dependent on interleukin-7 (IL-7). Immune activation, however, downregulates IL-7 receptor expression on T cells so that T cell survival during activation must be maintained independently of IL-7. The pro-inflammatory cytokine IL-6 shares common signaling pathways with IL-7 and can promote T cell survival in vitro. But whether IL-6 promotes T cell survival and homeostasis in vivo is not clear. Notably, IL-6 overexpression results in massive plasmacytosis and autoimmunity so that an IL-6 effect on in vivo T cell survival has remained untested. To overcome this limitation, here we generated IL-6 transgenic mice on an immunoglobulin heavy chain (IgH) deficient background which rendered them B cell deficient. Notably, such IgH(KO)IL6(Tg) mice were free of any signs of inflammation or autoimmunity and remained healthy throughout the course of analysis. In these mice, we found that IL-6 overexpression significantly increased peripheral T cell numbers, but importantly without increasing thymopoiesis. Moreover, IL-6 signaled T cells maintained their naïve phenotype and did not express activation/memory markers, suggesting that increased T cell numbers were due to increased T cell survival and not because of expansion of activated T cells. Mechanistically, we found that IL-6 signaling induced expression of pro-survival factors Mcl-1 and Pim-1/-2 but not Bcl-2. Thus, IL-6 is a T cell homeostatic cytokine that expands T cell space and can maintain the naïve T cell pool.

Survival in patients with neuroendocrine tumors of the colon, rectum and small intestine.

BACKGROUND: Neuroendocrine neoplasms (NENs) of the colon, rectum and small intestine (SI) are increasing in incidence and prevalence. We evaluated the 5-year overall survival (OS), and cancer-specific survival (CSS). METHODS: The Surveillance, Epidemiology, and End Results (SEER) 18 registry from 2000 to 2017 was accessed to identify patients with colonic, rectal, and SI NENs. RESULTS: 46,665 patients were diagnosed with NENs of the colon (n = 10,518, 22.5%), rectum (18,063, 38.7%), and SI (18,084, 38.8%). By tumor site alone, patients with well-differentiated neuroendocrine tumors (NETs) of the rectum had improved 5-year OS (HR 0.72, 95% CI 0.68-0.77, p < 0.001). However, patients with rectal poorly-differentiated neuroendocrine carcinomas (NECs) who underwent oncologic resection had lower 5-year OS (35.1%) compared to colon (41.9%), and SI (72.5%). CONCLUSIONS: Surgical resection may improve 5-year OS for NECs of the SI and colon, except in the rectum where survival was reduced. More frequent surveillance and timely initiation of systemic therapy should be considered for rectal NECs.

The Cytokine Receptor IL-7Rα Impairs IL-2 Receptor Signaling and Constrains the In Vitro Differentiation of Foxp3+ Treg Cells.

IL-7 receptor signaling is essential for the generation and maintenance of conventional T cells. Immunosuppressive Foxp3+ Treg cells, however, express uniquely low amounts of the IL-7-proprietary IL-7Rα so that they are impaired in IL-7 signaling. Because Treg cells depend on IL-2, the loss of IL-7Rα has been considered irrelevant for Treg cells. In contrast, here, we report that IL-7Rα downregulation is necessary to maximize IL-2R signaling. Although IL-7Rα overexpression promoted IL-7 signaling, unexpectedly, IL-2 signaling was suppressed in the same cells. Mechanistically, we found that γc, which is a receptor subunit shared by IL-7R and IL-2R, directly binds and pre-associates with IL-7Rα, thus limiting its availability for IL-2R binding. Consequently, overexpression of signaling-deficient, tailless IL-7Rα proteins inhibited IL-2R signaling, demonstrating that IL-7Rα sequesters γc and suppresses IL-2R signaling by extracellular interactions. Collectively, these results reveal a previously unappreciated regulatory mechanism of IL-2 receptor signaling that is governed by IL-7Rα abundance.

Multiple Endocrine Neoplasia Type 1: A Case Report With Review of Imaging Findings.

BACKGROUND: Multiple endocrine neoplasia type 1 (MEN1) is a rare, autosomal dominant inherited syndrome caused by mutations in the MEN1 tumor suppressor gene. The diagnosis is defined clinically by the presence of 2 or more primary MEN1 tumors (parathyroid, anterior pituitary, and pancreatic islet). We describe the case of a patient who presented with classic history and imaging findings for MEN1. CASE REPORT: A male in his early thirties with a history of hyperparathyroidism and a transsphenoidal prolactinoma resection presented years later with abdominal symptoms concerning for Zollinger-Ellison syndrome: worsening epigastric abdominal pain, nausea, vomiting, and diarrhea. Contrast-enhanced computed tomography (CT) of the abdomen revealed hyperenhancing pancreatic lesions and duodenal inflammation, suggesting pancreatic neuroendocrine tumor (gastrinoma) with secondary duodenitis. Bilateral indeterminate hypoattenuating adrenal nodules were also seen on contrast-enhanced CT, and follow-up magnetic resonance imaging confirmed benign adrenal adenomas. Furthermore, thyroid ultrasound and sestamibi scintigraphy revealed a parathyroid adenoma. With confirmatory imaging findings, history, and presenting symptoms, the patient was clinically diagnosed with MEN1 syndrome and underwent surgical and medical management. CONCLUSION: This case exhibits the classic history with corresponding imaging findings of MEN1 syndrome, including pancreatic neuroendocrine tumors, parathyroid adenoma, and adrenal adenomas. High clinical suspicion for MEN1 should lead to endocrinology evaluation with appropriate laboratory workup and targeted imaging evaluation of the typical endocrine organs as described for this patient.

Overcoming resistance to targeted therapy with immunotherapy and combination therapy for metastatic melanoma.

Resistance to targeted therapy is an ongoing problem for the successful treatment of Stage IV metastatic melanoma. For many patients, the use of targeted therapies, such as BRAF kinase inhibitors, were initially promising yet resistance inevitably occurred. Even after combining BRAF kinase inhibitors with MEK pathway inhibitors to offset re-activation of the MAP kinase pathway, resistance is still documented. Similarly, outcomes with immune checkpoint inhibitors as monotherapy were optimistic for some patients without relapse or progression, yet the majority of patients undergoing monotherapy have progressive disease. Will immunotherapy and combination therapy trials overcome resistance in metastatic melanoma? In an effort to treat resistant disease, new clinical trials evaluating the combination of immunotherapy with other therapies, such as kinase inhibitors, adoptive cell therapy, chimeric CD40 ligand to boost costimulation, or a tumor-specific oncolytic virus enhancing granulocyte macrophage colony-stimulating factor (GM-CSF) expression, are currently underway. Updated studies on the mechanisms of resistance, immune escape and options to reinvigorate immune cells support the continued discovery of new and improved forms of therapy.