The future of affordable cancer immunotherapy.

The treatment of cancer was revolutionized within the last two decades by utilizing the mechanism of the immune system against malignant tissue in so-called cancer immunotherapy. Two main developments boosted cancer immunotherapy: 1) the use of checkpoint inhibitors, which are characterized by a relatively high response rate mainly in solid tumors; however, at the cost of serious side effects, and 2) the use of chimeric antigen receptor (CAR)-T cells, which were shown to be very efficient in the treatment of hematologic malignancies, but failed to show high clinical effectiveness in solid tumors until now. In addition, active immunization against individual tumors is emerging, and the first products have reached clinical approval.

Preventing skin toxicities induced by EGFR inhibitors by topically blocking drug-receptor interactions.

Epidermal growth factor receptor (EGFR) inhibitors are used to treat many advanced-stage epithelial cancers but induce severe skin toxicities in most treated patients. These side effects lead to a deterioration in the quality of life of the patients and compromise the anticancer treatment. Current treatment strategies for these skin toxicities focus on symptom reduction rather than preventing the initial trigger that causes the toxicity.

The role of immune checkpoint receptors in the malignant phenotype of cutaneous T cell lymphoma.

Immune checkpoint receptors (ICR) modulate the immune response and are critical hubs for immunotherapy. However, data on their role in T lymphoid malignancies, such as cutaneous T cell lymphoma (CTCL), is sparse. We aimed to explore the role of ICR in the malignant features of transformed T lymphocytes and evaluate the effect of ICR-targeting monoclonal antibodies, often used as immunotherapy for solid tumors.

Alternative Splicing of the Inhibitory Immune Checkpoint Receptor SLAMF6 Generates a Dominant Positive Form, Boosting T-cell Effector Functions.

SLAMF6 is a homotypic receptor of the Ig-superfamily associated with progenitor-exhausted T cells. Here we show that in humans, SLAMF6 has three splice isoforms involving its V-domain. Although the canonical receptor inhibited T-cell activation through SAP recruitment, the short isoform SLAMF6 had a strong agonistic effect.

SLAMF6​ deficiency augments tumor killing and skews toward an effector phenotype revealing it as a novel T cell checkpoint.

SLAMF6 is a homotypic receptor of the Ig-superfamily whose exact role in immune modulation has remained elusive. Its constitutive expression on resting and activated T cells precludes it from being a exhaustion marker. By breeding Pmel-1 mice with SLAMF6 -/- mice, we generated donors for T cells lacking SLAMF6 and expressing a transgenic TCR for gp100-melanoma antigen.

Soluble SLAMF6 Receptor Induces Strong CD8 T-cell Effector Function and Improves Anti-Melanoma Activity .

SLAMF6, a member of the SLAM (signaling lymphocyte activation molecules) family, is a homotypic-binding immune receptor expressed on NK, T, and B lymphocytes. Phosphorylation variance between T-cell subclones prompted us to explore its role in anti melanoma immunity. Using a 203-amino acid sequence of the human SLAMF6 (seSLAMF6) ectodomain, we found that seSLAMF6 reduced activation-induced cell death and had an antiapoptotic effect on tumor-infiltrating lymphocytes.

Phenotype-based variation as a biomarker of sensitivity to molecularly targeted therapy in melanoma.

Transcriptomic phenotypes defined for melanoma have been reported to correlate with sensitivity to various drugs. In this study, we aimed to define a minimal signature that could be used to distinguish melanoma sub-types , and to determine suitable drugs by which these sub-types can be targeted. By using primary melanoma cell lines, as well as commercially available melanoma cell lines, we find that the evaluation of MLANA and INHBA expression is as capable as one based on a combined analysis performed with genes for stemness, EMT and invasion/proliferation, in identifying melanoma subtypes that differ in their sensitivity to molecularly targeted drugs.

Immune Monitoring of Patients Treated With a Whole-Cell Melanoma Vaccine Engineered to Express 4-1BBL.

CD8 lymphocytes are mandatory mediators of tumor regression. To enhance their specific antitumor activity, we aimed to improve a melanoma cell-based vaccine by transfecting it with 4-1BB ligand, a costimulatory and immune modulatory molecule. Thirty-four American Joint Committee on Cancer (AJCC) stage IIB-IV patients were vaccinated with a melanoma antigen-rich cell line engineered to express HLA-A2 and 4-1BBL (M20/A2/BBL).

Adjuvant Autologous Melanoma Vaccine for Macroscopic Stage III Disease: Survival, Biomarkers, and Improved Response to CTLA-4 Blockade.

Background. There is not yet an agreed adjuvant treatment for melanoma patients with American Joint Committee on Cancer stages III B and C. We report administration of an autologous melanoma vaccine to prevent disease recurrence.

Messenger RNA encoding constitutively active Toll-like receptor 4 enhances effector functions of human T cells.

Adoptive T cell therapy of cancer employs a large number of ex-vivo-propagated T cells which recognize their targets either by virtue of their endogenous T cell receptor (TCR) or via genetic reprogramming. However, both cell-extrinsic and intrinsic mechanisms often diminish the in-vivo potency of these therapeutic T cells, limiting their clinical efficacy and broader use. Direct activation of human T cells by Toll-like receptor (TLR) ligands induces T cell survival and proliferation, boosts the production of proinflammatory cytokines and augments resistance to regulatory T cell (Treg) suppression.

Human T cell crosstalk is induced by tumor membrane transfer.

Trogocytosis is a contact-dependent unidirectional transfer of membrane fragments between immune effector cells and their targets, initially detected in T cells following interaction with professional antigen presenting cells (APC). Previously, we have demonstrated that trogocytosis also takes place between melanoma-specific cytotoxic T lymphocytes (CTLs) and their cognate tumors. In the present study, we took this finding a step further, focusing on the ability of melanoma membrane-imprinted CD8+ T cells to act as APCs (CD8+ T-APCs).

Imprinting of lymphocytes with melanoma antigens acquired by trogocytosis facilitates identification of tumor-reactive T cells.

Trogocytosis is a contact-dependent intercellular transfer of membrane fragments and associated molecules from APCs to effector lymphocytes. We previously demonstrated that trogocytosis also occurs between tumor target and cognate melanoma Ag-specific cytotoxic T cells (CTL). In this study, we show that, following trogocytosis, immune effector cells acquire molecular components of the tumor, including surface Ags, which are detectable by specific mAbs.

Trogocytosis is a gateway to characterize functional diversity in melanoma-specific CD8+ T cell clones.

Trogocytosis, the transfer of membrane patches from target to immune effector cells, is a signature of tumor-T cell interaction. In this study, we used the trogocytosis phenomenon to study functional diversity within tumor-specific T cell clones with identical TCR specificity. MART-1(26-35)-specific CD8 T cell clones, which differed in their trogocytosis capacity (low [2D11], intermediate [2G1], high [2E2]), were generated from melanoma patients.

HLA-B35 correlates with a favorable outcome following adjuvant administration of an HLA-matched allogeneic melanoma vaccine.

This work presents survival data of 42 melanoma patients at high risk for disease recurrence who received an allogeneic melanoma vaccine composed of three cell lines, each matching at least one allele of the recipient's human leukocyte antigen (HLA)-A and -B loci. The 5-year overall survival (OS) rate and disease-free survival (DFS) compared favorably with the standard interferon-α regimen. Interestingly, patients bearing HLA-B35 had significantly better OS and DFS (OS of 100% and DFS of 90% for HLA-B35 vs 56% and 23%, for the non-B35 patients).

Transcutaneous immunization with hydrophilic recombinant gp100 protein induces antigen-specific cellular immune response.

The objective of this study was to evaluate the potential of transcutaneous immunization with tumor antigen to induce cell-mediated immunity. For this purpose, hydrophilic recombinant gp100 protein (HR-gp100) was topically applied on human intact skin in vitro, and used as a vaccine in a mouse model. We demonstrate that HR-gp100 permeates into human skin, and is processed and presented by human dendritic cells.

Apheresis induces oxidative stress in blood cells.

Oxidative stress mediates damage to various cells and is thought to be involved in various pathologies, including hereditary and acquired hemolytic anemias. It is induced by a multitude of physiological and environmental factors, including extracorporeal manipulation of blood. As a result, hemodialysis induces oxidative damage to red blood cells, thereby increasing their susceptibility to hemolysis and shortening their life span.

Autologous melanoma vaccine induces antitumor and self-reactive immune responses that affect patient survival and depend on MHC class II expression on vaccine cells.

Purpose: Autologous melanoma cells display a broad variety of tumor antigens and were used for treatment of American Joint Committee on Cancer stages III and IV melanoma as an adjuvant or active therapy. Survival data and immune response were evaluated in vaccinated patients.

Experimental Design: Forty-seven patients received 2,4-dinitrophenyl-conjugated autologous melanoma vaccine as an adjuvant (23 patients) or therapy (24 patients).

Capture of tumor cell membranes by trogocytosis facilitates detection and isolation of tumor-specific functional CTLs.

The success of adoptive cell transfer in the treatment of metastatic cancer in humans is dependent on the selection of highly active tumor-specific cytotoxic T cells. We report here that CTLs capture membrane fragments from their targets while exerting cytotoxic activity and thus gain a detectable functional signature by which they can be identified. Fluorochrome labeling or biotinylation was used to tag tumor cells.

A melanoma multiepitope polypeptide induces specific CD8+ T-cell response.

Strategies using epitope-based vaccination are being considered for melanoma immunotherapy, in an attempt to overcome failure of other modalities. In the present study, we designed and produced a multiepitope polypeptide for melanoma (MEP-mel), which contains three repeats of four antigenic epitopes (gp100: 209-217 (210M); gp100: 280-288 (288V); Mart1: 26-35 (27L); tyrosinase: 368-376 (370D). The peptides were attached to each other by linkers containing sequences recognized by the proteasome, to improve protein cleavage and antigen presentation.

Immunological activation following transcutaneous delivery of HR-gp100 protein.

Transcutaneous immunization aims at taking advantage of the skin's immune system for the purpose of immunoprotection. In the present study, we evaluated the potential of topical delivery of a recombinant melanoma protein, HR-gp100, derived from a shortened sequence of the native gp100 gene. The protein was applied on the skin, with and without the addition of two forms of heat labile enterotoxin (nLT and LTB).

Heat labile enterotoxin of E. coli: a potential adjuvant for transcutaneous cancer immunotherapy.

Escherichia coli heat labile enterotoxin (LT) has been shown to penetrate intact skin and to activate adaptive immunity. A nontoxic mutant, nLT, and its B subunit (LTB), have been evaluated separately for their potential use as a tool for transcutaneous delivery of antigens for cancer immunotherapy. We have shown that FITC-labeled nLT is taken up by human dendritic cells (hDC) in vitro and in mouse skin, and induces maturation and activation of hDC in vitro.

Recombinant hydrophilic human gp100: uptake by dendritic cells and stimulation of autologous CD8+ lymphocytes from melanoma patients.

Native gp100, a glycoprotein highly expressed in the majority of melanomas, contains several immunogenic peptides that are recognized by cytotoxic lymphocytes (CTLs) in the context of major histocompatibility complex (MHC) class I molecules. The objective of this study was to evaluate the ability of dendritic cells (DCs) from melanoma patients to take up gp100 protein and stimulate specific autologous CTL. The gp100 used in this study was a recombinant molecule with diminished hydrophobicity, HR-gp100, produced in Escherichia coli bacteria and in Pichia pastoris yeast.

Production and purification of melanoma gp100 antigen and polyclonal antibodies.

gp100 is a melanoma-associated antigen found to carry immunogenic epitopes that can induce a CTL response against tumor cells. Production and purification of large quantities of this polypeptide may be important in the context of diagnosis and vaccinating against melanoma. To overcome the hydrophobic nature of gp100, we cloned and expressed only a part of the protein, and obtained a hydrophilic recombinant polypeptide (HR-gp100) that contained most of the immunogenic peptides.

Interleukin-2 improves tumour response to DNP-modified autologous vaccine for the treatment of metastatic malignant melanoma.

This paper is a report of response rate (RR) and survival of 34 metastatic melanoma patients who received a dinitrophenyl (DNP)-modified autologous melanoma cell vaccine. In all, 27 patients started the vaccine as a primary treatment for metastatic melanoma and seven started it as an adjuvant, with no evidence of disease at the time, but had developed new metastases. Interleukin-2 (IL-2) was administered in 24 out of the 34 patients: 19 who progressed on vaccine alone and five who had the combination from start.

Topical treatment of persistent cutaneous leishmaniasis with ethanolic lipid amphotericin B.

A 1.5-year-old infant was referred because of spreading skin lesions diagnosed earlier as cutaneous leishmaniasis that did not respond to repeated courses with paromomycin-containing ointment. After consideration of the alternative therapeutic options, the infant was treated with a topical colloidal solution of amphotericin B for 3 weeks, This mode of therapy resulted in resolution of the skin lesions.