Inhibition of Interleukin-10 in the tumor microenvironment can restore mesothelin chimeric antigen receptor T cell activity in pancreatic cancer in vitro.

BACKGROUND: Pancreatic cancer cells are known to shield themselves from immunosurveillance by secreting immune inhibitory cytokines such as Interleukin-10. Using mesothelin, a differentiating antigen that is overexpressed in pancreatic cancer, we assessed the negative effect of the tumor microenvironment on chimeric antigen receptor T cell-based immunotherapy and its reversal via depletion of Interleukin-10. METHODS: T cells cultured in pancreatic cancer-cell-conditioned medium were transduced with lentiviruses encoding mesothelin-chimeric antigen receptor in the presence or absence of anti-Interleukin-10-blocking antibody. RESULTS: Coculture supernatants of conditioned medium displayed significant inhibition of interferon γ and granzyme B secretion, both of which are crucial for induction of target cell cytotoxicity. In contrast, this inhibition was restored toward baseline when conditioned medium was Interleukin-10- depleted (p < .05 for both interferon γ and granzyme B). In addition, we observed a significant decrease in mesothelin-chimeric antigen receptor T cell-induced cytotoxicity of BxPC-3 target cells in the presence of conditioned medium. Furthermore, we observed a partial blunting of this inhibition when Interleukin-10 was depleted from the conditioned medium. CONCLUSION: Substantial reversal of tumor-derived immunosuppression may be achieved by blocking Interleukin-10 in the local microenvironment, allowing for more effective cytotoxicity of mesothelin-engrafted chimeric antigen receptor T cells and enhancing the potential for clinical application.

Enhanced phosphorylation of p53 by microRNA-26a leading to growth inhibition of pancreatic cancer.

PURPOSE: MicroRNA (miR)-26a has been identified as a tumor suppressor in pancreatic cancer cells. Although wild-type p53 controls cell-cycle progression, its mutant form normally present in pancreatic cancer loses this capability. Phosphorylation is known to restore wild-type activity to mutant p53. We, therefore, examined whether miR-26a treatment can restore wild-type functions of mutant p53 via phosphorylation, resulting in inhibition of cell growth. METHODS: The human pancreatic cancer cell line BxPc-3 harboring mutant p53 was used for colony formation, cell-cycle, and Western blotting assays. Gene profile analysis was conducted after transfection with pre-miR-26a. RESULTS: miR-26a expression significantly decreased cell proliferation by 80% along with marked inhibition of colony formation and cell migration. Cell-cycle inhibition at the G0/G1 interface was observed along with enhanced drug retention and increased chemosensitivity to gemcitabine. Mutant p53 was phosphorylated rapidly at its Ser9 and Ser392 residues, but not at Ser15 or Ser20. Gene profile analysis of pre-miR-26a-transfected cells showed a significant increase in gene transcripts promoting apoptosis and p53 activation, with decreased levels of genes involved in cell-cycle progression. CONCLUSION: Delivery of miR-26a may represent a novel strategy for inhibiting pancreatic cancer growth, at least in part by enhancing phosphorylation of mutant p53 to restore its wild-type functions.

Radiofrequency ablation offers a reliable surgical modality for the treatment of Barrett's esophagus with a minimal learning curve.

Radiofrequency ablation (RFA) has gained popularity as treatment for Barrett's esophagus. Inclusive series of patients from initiation of our Barrett's Therapy Program were studied. Review of patients undergoing RFA for Barrett's was performed from September 2008 to May 2011. Patients' outcomes were recorded and analyzed using standard statistical methods. Seventy patients were treated. Average age was 61 (28-70); 80 per cent were male. Seventy-four per cent had dysplasia; 44 low-grade and eight high-grade. A total of 75.7 per cent of patients had long and 24.3 per cent had short segment Barrett's. Procedures per patient ranged from one to seven. Number of treatments in long- and short-segment groups were not different (P = 0.11). The maximum number of treatments in the short-segment group was five with a median of three (44.3%). For long segment, the maximum of RFA procedures was seven, with a median of three (30.8%). Average procedure time was 20.8 minutes for long and 17.9 minutes for short segment. Mean follow-up was 16.1 (2-38) months. Complete response was accomplished in 81 per cent. There were 93.3 per cent of complete responders in the short-segment group versus 75 per cent in the long (P = 0.24). Complications included dysphagia (1), transient chest and cervical pain (1), and abdominal pain (1). Comparing the first 25 per cent of the RFA procedures to the later 75 per cent or first 50 per cent to second 50 per cent, there was no difference in operative time or complications. Two patients recurred, both in the long-segment group. RFA is a safe and effective means to eradicate Barrett's. By measure of treatment time, complication rate, and efficacy of therapy, there is minimal or no "learning curve" for experienced endoscopists.