T-Cell manipulation strategies to prevent graft-versus-host disease in haploidentical stem cell transplantation

Allogeneic haematopoietic stem cell transplantation (HSCT) from an human leukocyte antigen (HLA)-identical donor can be curative for eligible patients with non-malignant and malignant haematological disorders. HSCT from alternative donor sources, such as HLA-mismatched haploidentical donors, is increasingly considered as a viable therapeutic option for patients lacking HLA-matched donors. Initial attempts at haploidentical HSCT were associated with vigorous bidirectional alloreactivity, leading to unacceptably high rates of graft rejection and graft-versus-host disease (GVHD). More recently, new approaches for mitigating harmful T-cell alloreactivity that mediates GVHD, while preserving the function of tumour-reactive natural killer (NK) cells and γδ T cells, have led to markedly improved clinical outcomes, and are successfully being implemented in the clinic. This article will provide an update on in vitro strategies and in vivo approaches aimed at preventing GVHD by selectively manipulating key components of the adaptive immune response, such as T-cell receptor (TCR)- αβ T cells and CD45RA-expressing naive T cells

Targeting Multiple-Myeloma-Induced Immune Dysfunction to Improve Immunotherapy Outcomes

Multiple myeloma (MM) is a plasma cell malignancy associated with high levels of monoclonal (M) protein in the blood and/or serum. MM can occur de novo or evolve from benign monoclonal gammopathy of undetermined significance (MGUS). Current translational research into MM focuses on the development of combination therapies directed against molecularly defined targets and that are aimed at achieving durable clinical responses. MM cells have a unique ability to evade immunosurveillance through several mechanisms including, among others, expansion of regulatory T cells (Treg), reduced T-cell cytotoxic activity and responsiveness to IL-2, defects in B-cell immunity, and induction of dendritic cell (DC) dysfunction. Immune defects could be a major cause of failure of the recent immunotherapy trials in MM. This article summarizes our current knowledge on the molecular determinants of immune evasion in patients with MM and highlights how these pathways can be targeted to improve patients' clinical outcome

Granulocyte-colony stimulating factor drives the in vitro differentiation of human dendritic cells that induce anergy in naïve T cells

G-CSF is a modulator of T-cell and DC functions. Previous reports show that monocytes from G-CSF-treated (post-G) healthy donors differentiate into tolerogenic DC in vitro in the presence of autologous serum, containing high levels of IL-10 and IFN-α, and in turn induce type 1 Treg (Tr1) cells. However, the direct effect of G-CSF on DC differentiation was not investigated. Here, we show that monocytes differentiated in the presence of exogenous G-CSF (G-DC) remain CD14+CD1a−, but acquire a DC-like morphology, express CD83 and CD86 and low levels of the tolerogenic markers Ig-like transcript (ILT)4 and HLA-G. G-DC spontaneously produce IL-10 and, upon stimulation, low levels of IL-12. G-DC display low stimulatory capacity and induce anergy in naïve T cells, but do not confer suppressive function. Therefore, in vitro differentiation of monocyte-derived DC in the presence of G-CSF can replicate some but not all features of post-G DC. These findings indicate that the tolerogenic properties of G-CSF do not exclusively reside in its direct effect on DC, which in turn induce T-cell anergy, but also in its ability to generate a tolerogenic milieu in vivo, which is necessary for Tr1 cell induction and cannot be replicated in vitro

Survival and bioactivities of selected probiotic lactobacilli in yogurt fermentation and cold storage: New insights for developing a bi-functional dairy food

In previous work , we demon strated that two probiotic strains, name ly Lactobacillus casei PRA205 and Lactobacillus rhamnosus PRA331, produce fermented milks with potent angiotensin-converting enzyme (ACE)-inhibitory and antioxidant activities. Here, we tested these strains for the survivability and the release of antihypertensive and antioxidant peptides in yogurt fermentation and cold storage. For these purposes three yogurt batches were compared: one prepared using yogurt starters alone (Lactobacillus delbrueckii subspecies bulgaricus 1932 and Streptococcus thermophilus 99), and the remaining two containing either PRA205 or PRA331 in addition to yogurt starters. Despite the lower viable counts at the fermentation end compared to PRA331, PRA205 overcame PRA331 in survivability during refrigerated storage for 28 days, leading to viable counts (>108 CFU/g) higher than the minimum therapeutic threshold (106 CFU/g). Analyses of in vitro ACE-inhibitory and antioxidant activities of peptide fractions revealed that yogurt supplemented with PRA205 displays higher amounts of antihypertensive and antioxidant peptides than that produced with PRA331 at the end of fermentation and over storage. Two ACE-inhibitory peptides, Valine-Proline-Proline (VPP) and Isoleucine-Proline-Proline (IPP), were identified and quantified. This study demonstrated that L. casei PRA205 could be used as adjunct culture for producing bi-functional yogurt enriched in bioactive peptides and in viable cells, which bring health benefits to the host as probiotics

Granulocyte transfusions in children and adults with hematological malignancies: benefits and controversies

Bacterial and fungal infections continue to pose a major clinical challenge in patients with prolonged severe neutropenia after chemotherapy or hematopoietic stem cell transplantation (HSCT). With the advent of granulocyte colony-stimulating factor (G-CSF) to mobilize neutrophils in healthy donors, granulocyte transfusions have been broadly used to prevent and/or treat life-threatening infections in patients with severe febrile neutropenia and/or neutrophil dysfunction. Although the results of randomized controlled trials are inconclusive, there are suggestions from pilot and retrospective studies that granulocyte transfusions may benefit selected categories of patients. We will critically appraise the evidence related to the use of therapeutic granulocyte transfusions in children and adults, highlighting current controversies in the field and discussing complementary approaches to modulate phagocyte function in the host

The Asp272-Glu282 Region of Platelet Glycoprotein Ib Interacts with the Heparin-binding Site of -Thrombin and Protects the Enzyme from the Heparin-catalyzed Inhibition by Antithrombin III

Platelet glycoprotein Ib (GpIb) mediates interaction with both von Willebrand factor and thrombin. Thrombin binds to GpIb via its heparin-binding site (HBS) (De Candia, E., De Cristofaro, R., De Marco, L., Mazzucato, M., Picozzi, M., and Landolfi, R. (1997) Thromb. Haemostasis 77, 735–740; De Cristofaro, R., De Candia, E., Croce, G., Morosetti, R., and Landolfi, R. (1998) Biochem. J. 332, 643–650). To identify the thrombin-binding domain on GpIbα, we examined the effect of GpIbα1–282, a GpIbα fragment released by the cobra venom mocarhagin on the heparin-catalyzed rate of thrombin inhibition by antithrombin III (AT). GpIbα1–282 inhibited the reaction in a dose-dependent and competitive fashion. In contrast, the GpIbα1–271 fragment, produced by exposing GpIbα1–282 to carboxypeptidase Y, had no effect on thrombin inhibition by the heparin-AT complex. Measurements of the apparent equilibrium constant of the GpIbα1–282 binding to thrombin as a function of different salts (NaCl and tetramethyl-ammonium chloride) concentration (0.1–0.2 M) indicated a large salt dependence (Γ± = −4.5), similar to that pertaining to the heparin binding to thrombin. The importance of thrombin HBS in its interaction with GpIbα was confirmed using DNA aptamers, which specifically bind to either HBS (HD22) or the fibrinogen recognition site of thrombin (HD1). HD22, but not HD1, inhibited thrombin binding to GpIbα1–282. Furthermore, the proteolytic derivative γT-thrombin, which lacks the fibrinogen recognition site, binds to GpIbα via its intact HBS in a reaction that is inhibited by HD22. Neither α- nor γT-thrombin bound to GpIbα1–271, suggesting that the Asp272–Glu282 region of GpIbα may act as a “heparin-like” ligand for the thrombin HBS, thereby inhibiting heparin binding to thrombin. It was also demonstrated that intact platelets may dose-dependently inhibit the heparin-catalyzed thrombin inhibition by AT at enzyme concentrations <5 nM. Altogether, these findings show that thrombin HBS binds to the region of GpIbα involving the Asp272–Glu282 segment, protecting the enzyme from the inactivation by the heparin-AT system

Reverting immune suppression to enhance cancer immunotherapy

Tumors employ strategies to escape immune control. The principle aim of most cancer immunotherapies is to restore effective immune surveillance. Among the different processes regulating immune escape, tumor microenvironment-associated soluble factors, and/or cell surface-bound molecules are mostly responsible for dysfunctional activity of tumor-specific CD8+T cells. These dynamic immunosuppressive networks prevent tumor rejection at several levels, limiting also the success of immunotherapies. Nevertheless, the recent clinical development of immune checkpoint inhibitors or of molecules modulating cellular targets and immunosuppressive enzymes highlights the great potential of approaches based on the selective disruption of immunosuppressive networks. Currently, the administration of different categories of immunotherapy in combination regimens is the ultimate modality for impacting the survival of cancer patients. With the advent of immune checkpoint inhibitors, designed to mount an effective antitumor immune response, profound changes occurred in cancer immunotherapy: from a global stimulation of the immune system to a specific targeting of an immune component. This review will specifically highlight the players, the mechanisms limiting an efficient antitumor response and the current immunotherapy modalities tailored to target immune suppressive pathways. We also discuss the ongoing challenges encountered by these strategies and provide suggestions for circumventing hurdles to new immunotherapeutic approaches, including the use of relevant biomarkers in the optimization of immunotherapy regimens and the identification of patients who can benefit from defined immune-based approaches