Background: Tumor immune tolerance can derive from the recruitment of suppressor cell populations, including myeloid-derived suppressor cells (MDSC). In cancer patients, MDSC accumulation correlates with increased tumor burden, but the mechanisms of MDSC induction remain poorly understood. Methods: This study examined the ability of human tumor cell lines to induce MDSC from healthy donor PBMC using in vitro co-culture methods. These human MDSC were then characterized for morphology, phenotype, gene expression, and function. Results: Of over 100 tumor cell lines examined, 45 generated canonical CD33⁺HLA-DR^lowLineage^- MDSC, with high frequency of induction by cervical, ovarian, colorectal, renal cell, and head and neck carcinoma cell lines. CD33⁺ MDSC could be induced by cancer cell lines from all tumor types with the notable exception of those derived from breast cancer (0/9, regardless of hormone and HER2 status). Upon further examination, these and others with infrequent CD33⁺ MDSC generation were found to induce a second subset characterized as CD11b⁺CD33^lowHLA-DR^lowLineage^-. Gene and protein expression, antibody neutralization, and cytokine-induction studies determined that the induction of CD33⁺ MDSC depended upon over-expression of IL-1β, IL-6, TNFα, VEGF, and GM-CSF, while CD11b⁺ MDSC induction correlated with over-expression of FLT3L and TGFβ. Morphologically, both CD33⁺ and CD11b⁺ MDSC subsets appeared as immature myeloid cells and had significantly up-regulated expression of iNOS, NADPH oxidase, and arginase-1 genes. Furthermore, increased expression of transcription factors HIF1α, STAT3, and C/EBPβ distinguished MDSC from normal counterparts. Conclusions: These studies demonstrate the universal nature of MDSC induction by human solid tumors and characterize two distinct MDSC subsets: CD33⁺HLA-DR^lowHIF1α⁺/STAT3⁺ and CD11b⁺HLA-DR^lowC/EBPβ⁺, which should enable the development of novel diagnostic and therapeutic reagents for cancer immunotherapy.

Purpose: To evaluate MHC class I expression on papillary thyroid cancer (PTC) and analyze changes in MHC expression and associated immune activation with current and experimental treatments for thyroid cancer using in vitro PTC cell lines. Experimental Design: MHC class I expression and assessment of tumor-infiltrating leukocyte populations were evaluated by immunohistochemistry. PTC cell lines were analyzed for HLA-ABC expression by flow cytometry following tyrosine kinase inhibitor, IFNα or IFNγ, or radiation treatment. Functional changes in antigenicity were assessed by coculture of allogeneic donor peripheral blood leukocytes (PBL) with pretreated or untreated PTC cell lines and measurement of T-cell activation and cytokine production. Results: Both MHC class I and β2-microglobulin expression was reduced or absent in 76% of PTC specimens and was associated with reduced tumor-infiltrating immune cells, including effector (CD3⁺, CD8⁺, CD16⁺) and suppressor (FoxP3⁺) populations. Treatment of PTC cell lines with the MEK1/2 inhibitor selumetinib or IFN increased HLA-ABC expression. This phenotypic change was associated with increased T-cell activation (%CD25⁺ of CD3⁺) and IL2 production by PBL cocultured with treated PTC cell lines. Additive effects were seen with combination selumetinib and IFN treatment. Conclusions: MHC class I expression loss is frequent in human PTC specimens and represents a significant mechanism of immune escape. Increased antigenicity following selumetinib and IFN treatment warrants further study for immunotherapy of progressive PTC. Clin Cancer Res; 20(23); 6034–44. ©2014 AACR.

The development of recombinant therapeutic proteins has been a major revolution in modern medicine. Therapeutic-based monoclonal antibodies (mAbs) are growing rapidly, providing a potential class of human pharmaceuticals that can improve the management of cancer, autoimmune diseases, and other conditions. Most mAbs are typically of the immunoglobulin G (IgG) subclass, and they are glycosylated at the conserved asparagine position 297 (Asn-297) in the CH2 domain of the Fc region. Post-translational modifications here account for the observed high heterogeneity of glycoforms that may or not impact the stability, pharmacokinetics (PK), efficacy, and immunogenicity of mAbs. These modifications are also critical for the Fc receptor binding, and consequently, key antibody effector functions including antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). Moreover, mAbs produced in non-human cells express oligosaccharides that are not normally found in serum IgGs might lead to immunogenicity issues when administered to patients. This review summarizes our understanding of the terminal sugar residues, such as mannose, sialic acids, fucose, or galactose, which influence therapeutic mAbs either positively or negatively in this regard. This review also discusses mannosylation, which has significant undesirable effects on the PK of glycoproteins, causing a decreased mAbs’ half-life. Moreover, terminal galactose residues can enhance CDC activities and Fc–C1q interactions, and core fucose can decrease ADCC and Fc–FcγRs binding. To optimize the therapeutic use of mAbs, glycoengineering strategies are used to reduce glyco-heterogeneity of mAbs, increase their safety profile, and improve the therapeutic efficacy of these important reagents.

Due to their high specificity, monoclonal antibodies have been widely investigated for their application in drug delivery to the central nervous system (CNS) for the treatment of neurological diseases such as stroke, Alzheimer’s, and Parkinson’s disease. Research in the past few decades has revealed that one of the biggest challenges in the development of antibodies for drug delivery to the CNS is the presence of blood–brain barrier (BBB), which acts to restrict drug delivery and contributes to the limited uptake (0.1–0.2% of injected dose) of circulating antibodies into the brain. This article reviews the various methods currently used for antibody delivery to the CNS at the preclinical stage of development and the underlying mechanisms of BBB penetration. It also describes efforts to improve or modulate the physicochemical and biochemical properties of antibodies (e.g., charge, Fc receptor binding affinity, and target affinity), to adapt their pharmacokinetics (PK), and to influence their distribution and disposition into the brain. Finally, a distinction is made between approaches that seek to modify BBB permeability and those that use a physiological approach or antibody engineering to increase uptake in the CNS. Although there are currently inherent difficulties in developing safe and efficacious antibodies that will cross the BBB, the future prospects of brain-targeted delivery of antibody-based agents are believed to be excellent.

BACKGROUND: Primary lymphomas of the breast are very rare (0.2‐1.5% of breast malignancies) and the vast majority (95%) are of B‐cell origin. Recently, 40 cases of clinically indolent anaplastic large‐cell kinase (ALK)‐negative, T‐cell, anaplastic, non‐Hodgkin lymphomas (T‐ALCL) have been reported worldwide.METHODS: A tumor biopsy specimen from a patient in this series was obtained for characterization. By using a human stromal feeder layer and IL‐2, a novel cell line, TLBR‐1, was established from this biopsy and investigated by using cytogenetics and various biomolecular methods.RESULTS: Immunoperoxidase staining of the tumor biopsy showed a CD30/CD8/CD4 coexpressing T‐cell population that was epithelial membrane antigen (EMA)⁺ and perforin⁺. Multiplex polymerase chain reaction (PCR) of TCRγ genes showed monoclonality that suggested a T‐cell origin, yet pan‐T markers CD2/5/7, anaplastic large‐cell kinase (ALK)‐1, pancytokeratins, CD20, CD56, and Epstein‐Barr virus (EBV) by in situ hybridization (ISH) were negative. TLBR‐1 is IL‐2 dependent, has a relatively long doubling time (55 hours), and displays different cellular shapes in culture. Cytogenetic analysis of tumor and TLBR‐1 cells confirmed a highly anaplastic cell population with a modal number of 47 chromosomes lacking t(2;5). PCR screens for EBV and human T‐lymphotropic virus types 1 and 2 (HTLV‐1/2) were negative. Fluorescence‐activated cell‐sorting (FACS) analysis showed strong positivity for CD4/8, CD30, CD71, and CD26 expression, and antigen presentation (HLA‐DR⁺CD80⁺CD86⁺), IL‐2 signaling (CD25⁺CD122⁺), and NK (CD56⁺) markers, and Western blots demonstrated strong Notch1 expression. Severe combined immunodeficiency (SCID) mouse TLBR‐1 heterotransplants recapitulated the histology and marker characteristics of the original tumor.CONCLUSIONS: TLBR‐1, a novel ALK‐negative, T‐cell, anaplastic, large‐cell lymphoma, closely resembles the original biopsy and represents an important tool for studying this newly recognized disease entity. Cancer 2011. © 2010 American Cancer Society.