Chemokines, along with their receptors, exert critical roles in tumor development and progression. In prostate cancer (PCa), interleukin-8 (IL-8/CXCL8) was shown to enhance angiogenesis, proliferation, and metastasis. CXCL8 activates two receptors, CXCR1 and CXCR2. While CXCR2 expression was shown to promote PCa growth and metastasis, the role of CXCR1 remains unclear.
In this study, we stably expressed CXCR1 and, as control, CXCR2 in the androgen-dependent PCa cell line MDA-PCa-2b to evaluate the effect of CXCR1 in tumor development.
MDA-PCa-2b-CXCR1 cells showed decreased cell migration, protein kinase-B (AKT) activation, prostate-specific antigen (PSA) expression, cell proliferation, and tumor development in nude mice, relative to MDA-PCa-2b-Vec and MDA-PCa-2b-CXCR2 cells. MDA-PCa-2b-CXCR1 cells also displayed a significant transition to mesenchymal phenotypes as characterized by decreased E-cadherin expression and a corresponding increased level of N-cadherin and vimentin expression. RNA-seq and Western blot analysis revealed a significant increase in the tumor suppressor integral membrane protein 2A (ITM2A) expression in MDA-PCa-2b-CXCR1 compared to control cells. In prostate adenocarcinoma tissue, ITM2A expression was also shown to be downregulated relative to a normal prostate. Interestingly, the overexpression of ITM2A in MDA-PCa-2b cells (MDA-PCa-2b-ITM2A-GFP) inhibited tumor growth similar to that of MDA-PCa-2b-CXCR1.
Taken together, the data suggest that CXCR1 expression in MDA-PCa-2b cells may upregulate ITM2A to abrogate tumor development.

Cancer is associated with a reduction in immature and mature circulating dendritic cells (DCs), and with an impaired migratory capacity, compared with healthy donors. Therefore, modern approaches to the in vitro generation of DCs loaded with tumor antigens and their use for inducing antitumor immune responses in vivo are being investigated. The purpose of the present study was to investigate the phenotypic and functional characteristics of peripheral blood DC subsets in patients with non-small cell lung cancer (NSCLC), and the development of an antitumor cytotoxic response by mononuclear cells (MNCs) from patients using in vitro generated antigen-primed DCs. Heparinized peripheral venous blood samples were obtained from 10 healthy donors and 20 patients with a histologically verified diagnosis of NSCLC. The ability of antigen-activated DCs to stimulate the activity of MNCs against autologous tumor cells was evaluated using a cytotoxic test. Peripheral blood DC subsets from patients with NSCLC were identified to be decreased and to exhibit an impaired ability to mature, compared with healthy donors. Furthermore, DCs generated from MNCs from patients with NSCLC were able to stimulate a specific cytotoxic response when loaded with autologous tumor lysates or RNA and matured, in vitro. A perforin and granzyme B-dependent mode of cytotoxicity was primarily induced. The ability of DCs loaded with tumor antigens to increase the cytotoxic activity of MNCs against NSCLC cells in vitro indicates the effective induction and co-stimulation of T lymphocytes by the generated DCs.

DotScan antibody microarrays were initially developed for the extensive surface profiling of live leukemia and lymphoma cells. DotScan's diagnostic capability was validated with an extensive clinical trial using mononuclear cells from the blood or bone marrow of leukemia or lymphoma patients. DotScan has also been used for the profiling of surface proteins on peripheral blood mononuclear cells (PBMC) from patients with HIV, liver disease, and stable and progressive B-cell chronic lymphocytic leukemia (CLL). Fluorescence multiplexing allowed the simultaneous profiling of cancer cells and leukocytes from disaggregated colorectal and melanoma tumor biopsies after capture on DotScan. In this chapter, we have used DotScan for the surface profiling of extracellular vesicles (EV) recovered from conditioned growth medium of cancer cell lines and the blood of patients with CLL. The detection of captured EV was performed by enhanced chemiluminescence (ECL) using biotinylated antibodies that recognized antigens expressed on the surface of the EV subset of interest. DotScan was also used to profile EV from the blood of healthy individuals and the ascites fluid of ovarian cancer patients. DotScan binding patterns of EV from human plasma and other body fluids may yield diagnostic or prognostic signatures for monitoring the incidence, treatment, and progression of cancers.

Despite activity as single agent cancer therapies, Rapamycin (rapa) and its rapalogs may have their greatest effects when combined with other therapeutic modalities. In addition to direct antitumor activity, rapalogs reverse multiple tumor-intrinsic immune evasion mechanisms. These should facilitate tumor-specific T cell activity, but since rapa directly inhibits effector T cells, this potential immune enhancement is lost. We hypothesized that if T cells were rendered resistant to rapa they could capitalize on its downregulation of tumor immune evasion. We therefore modified T cells with a rapa-resistant mutant of mTor, mTorRR, and directed them to B lymphomas by coexpressing a chimeric antigen receptor (CAR) for CD19 (CAR.CD19-28ζ). T cells expressing transgenic mTorRR from a piggyBac transposon maintain mTor signaling, proliferate in the presence of rapa and retain their cytotoxic function and ability to secrete interferon-γ (IFNγ) after stimulation, effector functions that were inhibited by rapa in control T cells. In combination, rapa and rapa-resistant-CAR.CD19-28ζ-expressing T cells produced greater antitumor activity against Burkitt's lymphoma and pre-B ALL cell lines in vitro than CAR.CD19-28ζ T cells or rapa alone. In conclusion, the combination of rapa and rapa-resistant, CAR.CD19-28ζ-expressing T cells may provide a novel therapy for the treatment of B cell malignancies and other cancers.

The aim of the present study has been to establish serum-free culture conditions for ex vivo expansion and differentiation of human CD34(+) cells into erythroid lineage and to study the chromatin structure, gene expression, and transcription factor recruitment at the alpha-globin locus in the developing erythron.
A basal Iscove's modified Dulbecco's medium cell culture medium with 1% bovine serum albumin as a serum replacement and a combination of cytokines and growth factors was used for expansion and differentiation of the CD34(+) cells. Expression patterns of the alpha- and beta-like genes at various stages of erythropoiesis was studied by reverse transcriptase quantitative polymerase chain reaction analysis, profile of key erythroid transcription factors was investigated by Western blotting, and the chromatin structure and transcription factor recruitment at the alpha-globin locus was investigated by chromatin immunoprecipitation quantitative polymerase chain reaction analysis.
Human CD34(+) cells in the serum-free medium undergo near synchronous erythroid differentiation to yield large amount of cells at different differentiation stages. We observe distinct patterns of the histone modifications and transcription factor binding at the alpha-globin locus during erythroid differentiation of CD34(+) cells. Nuclear factor erythroid-derived 2 (NF-E2) was present at upstream activator sites even before addition of erythropoietin (EPO), while bound GATA-1 was only detectable after EPO treatment. After 7 days of EPO treatment, H3K4Me2 modification uniformly increases throughout the alpha-globin locus. Acetylation at H3K9 and binding of Pol II, NF-E2, and GATA-1 were restricted to certain hypersensitive sites of the enhancer and theta gene, and were conspicuously low at the alpha-like globin promoters. Rearrangement of the insulator binding factor CTCF took place at and around the alpha-globin locus as CD34(+) cells differentiated into erythroid pathway.
Our results indicate that remodeling of the upstream elements may be the primary event in activation of alpha-globin gene expression. Activation of alpha-globin genes upon EPO treatment involves initial binding of Pol II, downregulation of pre-existing factors like NF-E2, removal of CTCF from the locus, then rebinding of CTCF in an altered pattern, and concurrent or subsequent binding of transcription factors like GATA-1.