Anti-OX40L alone or in combination with anti-CD40L and CTLA4Ig does not inhibit the humoral and cellular response to a major grass pollen allergen.

BACKGROUND: IgE-mediated allergy is a common disease characterized by a harmful immune response towards otherwise harmless environmental antigens. Induction of specific immunological non-responsiveness towards allergens would be a desirable goal. Blockade of costimulatory pathways is a promising strategy to modulate the immune response in an antigen-specific manner. Recently, OX40 (CD134) was identified as a costimulatory receptor important in Th2-mediated immune responses. Moreover, synergy between OX40 blockade and 'conventional' costimulation blockade (anti-CD40L, CTLA4Ig) was observed in models of alloimmunity. OBJECTIVE: We investigated the potential of interfering with OX40 alone or in combination with CD40/CD28 signals to influence the allergic immune response. METHODS: The OX40 pathway was investigated in an established murine model of IgE-mediated allergy where BALB/c mice are repeatedly immunized with the clinically relevant grass pollen allergen Phl p 5. Groups were treated with combinations of anti-OX40L, CTLA4Ig and anti-CD40L. In selected mice, Tregs were depleted with anti-CD25. RESULTS: Blockade of OX40L alone at the time of first or second immunization did not modulate the allergic response on the humoral or effector cell levels but slightly on T cell responses. Administration of a combination of anti-CD40L/CTLA4Ig delayed the allergic immune response, but antibody production could not be inhibited after repeated immunization even though the allergen-specific T cell response was suppressed in the long run. Notably, additional blockade of OX40L had no detectable supplementary effect. Immunomodulation partly involved regulatory T cells as depletion of CD25(+) cells led to restored T cell proliferation. CONCLUSIONS AND CLINICAL RELEVANCE: Collectively, our data provide evidence that the allergic immune response towards Phl p 5 is independent of OX40L, although reduction on T cell responses and slightly on the asthmatic phenotype was detectable. Besides, no relevant synergistic effect of OX40L blockade in addition to CD40L/CD28 blockade could be detected. Thus, the therapeutic potential of OX40L blockade for IgE-mediated allergy appears to be ineffective in this setting.

Rapamycin and CTLA4Ig synergize to induce stable mixed chimerism without the need for CD40 blockade.

The mixed chimerism approach achieves donor-specific tolerance in organ transplantation, but clinical use is inhibited by the toxicities of current bone marrow (BM) transplantation (BMT) protocols. Blocking the CD40:CD154 pathway with anti-CD154 monoclonal antibodies (mAbs) is exceptionally potent in inducing mixed chimerism, but these mAbs are clinically not available. Defining the roles of donor and recipient CD40 in a murine allogeneic BMT model, we show that CD4 or CD8 activation through an intact direct or CD4 T cell activation through the indirect pathway is sufficient to trigger BM rejection despite CTLA4Ig treatment. In the absence of CD4 T cells, CD8 T cell activation via the direct pathway, in contrast, leads to a state of split tolerance. Interruption of the CD40 signals in both the direct and indirect pathway of allorecognition or lack of recipient CD154 is required for the induction of chimerism and tolerance. We developed a novel BMT protocol that induces mixed chimerism and donor-specific tolerance to fully mismatched cardiac allografts relying on CD28 costimulation blockade and mTOR inhibition without targeting the CD40 pathway. Notably, MHC-mismatched/minor antigen-matched skin grafts survive indefinitely whereas fully mismatched grafts are rejected, suggesting that non-MHC antigens cause graft rejection and split tolerance.

Variations in genes involved in dormancy associated with outcome in patients with resected colorectal liver metastases.

BACKGROUND: Tumor dormancy has been described as a state of hibernation. Dormancy can be switched to proliferation by different pathways, which may play a critical role in tumor recurrence. In this study, we investigated genetic variations within genes involved in tumor dormancy and their association with recurrence and outcome in patients with colorectal liver metastases (CLM) who underwent neoadjuvant bevacizumab-based chemotherapy. PATIENTS AND METHODS: Genomic DNA was extracted from resected CLM (FFPE) from 149 patients. Single-nucleotide polymorphisms (SNPs) in 14 genes associated with dormancy were analyzed by direct Sanger DNA sequencing and evaluated for response, recurrence-free survival (RFS), overall survival (OS) and recurrence patterns. RESULTS: NME1 rs34214448 C>A was significantly associated with RFS in univariable analysis (P = 0.039) and with intrahepatic recurrence (P = 0.014). NOTCH3 rs1044009 T>C and CD44 rs8193 C>T showed a significant difference in 3-year OS rates (P = 0.004 and P = 0.042, respectively). With respect to radiological response, CD44 rs8193 C>T variant genotypes were associated with a significantly higher response rate (P = 0.033). Recursive partitioning analyses revealed that Dll4 rs12441495 C>G, NME1 rs34214448 C>A and NOTCH3 rs1044009 T>C were the dominant SNPs predicting histological response, RFS and OS, respectively. CONCLUSION: Our data suggest that gene variations within genes involved in tumor dormancy pathways are associated with response and outcome in patients with resected CLM. These data may lead to new and more effective treatment strategies targeting tumor dormancy.

Variations in genes involved in immune response checkpoints and association with outcomes in patients with resected colorectal liver metastases.

In patients with colorectal liver metastases (CLM), liver resection offers the possibility of cure and long-term survival. The liver is a highly immunogenic organ harboring ~80% of the body's tissue macrophages. Emerging data demonstrate a critical role of the immune response for cancer treatment. We investigated variations within genes involved in immune response checkpoints and their association with outcomes in patients with CLM who underwent neoadjuvant chemotherapy including bevacizumab and liver resection. Single-nucleotide polymorphisms (SNPs) in nine genes (CCL2, CCR2, LAG3, NT5E, PDCD1, CD274, IDO1, CTLA4 and CD24) were analyzed in genomic DNA from 149 patients with resected bevacizumab-pretreated CLM by direct Sanger DNA sequencing, and correlated with response, recurrence-free survival (RFS), overall survival (OS), probability of cure and recurrence patterns. IDO1 (indoleamine 2, 3-dioxygenase) rs3739319 G>A and CD24 rs8734 G>A showed a significant difference in 3-year OS rates. In addition, IDO1 rs3739319 G>A was significantly associated with extrahepatic recurrence. Recursive partitioning analyses revealed that IDO1 rs3739319 G>A was the dominant SNP predicting RFS and OS. Our data suggest that variants within genes involved in immune response checkpoints are associated with outcomes in patients with resected CLM and might lead to improved treatment strategies modulating anti-tumor immune response by targeting novel immune checkpoints.

Prophylactic and therapeutic vaccination with carrier-bound Bet v 1 peptides lacking allergen-specific T cell epitopes reduces Bet v 1-specific T cell responses via blocking antibodies in a murine model for birch pollen allergy.

BACKGROUND: Vaccines consisting of allergen-derived peptides lacking IgE reactivity and allergen-specific T cell epitopes bound to allergen-unrelated carrier molecules have been suggested as candidates for allergen-specific immunotherapy. OBJECTIVE: To study whether prophylactic and therapeutic vaccination with carrier-bound peptides from the major birch pollen allergen Bet v 1 lacking allergen-specific T cell epitopes has influence on Bet v 1-specific T cell responses. METHODS: Three Bet v 1-derived peptides, devoid of Bet v 1-specific T cell epitopes, were coupled to KLH and adsorbed to aluminium hydroxide to obtain a Bet v 1-specific allergy vaccine. Groups of BALB/c mice were immunized with the peptide vaccine before or after sensitization to Bet v 1. Bet v 1- and peptide-specific antibody responses were analysed by ELISA. T cell and cytokine responses to Bet v 1, KLH, and the peptides were studied in proliferation assays. The effects of peptide-specific and allergen-specific antibodies on T cell responses and allergic lung inflammation were studied using specific antibodies. RESULTS: Prophylactic and therapeutic vaccination with carrier-bound Bet v 1 peptides induced a Bet v 1-specific IgG antibody response without priming/boosting of Bet v 1-specific T cells. Prophylactic and therapeutic vaccination of mice with the peptide vaccine induced Bet v 1-specific antibodies which suppressed Bet v 1-specific T cell responses and allergic lung inflammation. CONCLUSION AND CLINICAL RELEVANCE: Vaccination with carrier-bound allergen-derived peptides lacking allergen-specific T cell epitopes induces allergen-specific IgG antibodies which suppress allergen-specific T cell responses and allergic lung inflammation.

[Critical assessment of a new endoscopic anatomic concept for the so-called cardia in the sense of the notions of Parmenides and Martin Heidegger]. / Kritische Betrachtung neuer Konzepte zur endoskopischen Anatomie des ösphagogastralen Übergangs im Sinne des Denkens von Parmenides und Martin Heidegger.

Current endoscopic anatomy interposes the gastric cardia between the tubular oesophagus and the proximal stomach. In contrast to that, recent evidence unfolds a different view. Using "PubMed" and "Scopus" searches, we examined if the novel understanding regarding the cardia goes in line with the concept of unfolding, as described by Heidegger based on the ancient didactic poetry of Parmenides. What has been taken as gastric cardia in fact represents reflux-damaged, dilated, columnar lined oesophagus (CLO): dilated distal oesophagus (DDO). Due to its macroscopic gastric appearance it cannot be discriminated from the stomach by endoscopy. Differentiation between DDE and proximal stomach requires the histopathology of measured multi-level biopsies obtained from the DDO and the proximal stomach. Cardaic, onxytocardiac mucosa and intestinal metaplasia (IM; Barrett's oesophagus) define CLO and thus the oesophageal location, while oxyntic mucosa (OM) of the proximal stomach verifies a gastric biopsy location. Endoscopically visible CLO and DDO define the morphological manifestation of reflux: the squamo-oxyntic gap (SOG). Biopsies obtained from the level of the diaphragmatic impressions allow differentiation between an enlarged hiatus with normal anatomic content (CLO; oesophagus) vs. hernia with abnormal content (OM; stomach). Non-dysplastic Barrett's oesophagus exists in 10 %-17 % of asymptomatic and in 20 %-100 % (with increasing CLO length) of reflux symptom-positive individuals (annual cancer risk: 0.2 %-0.7 %). These data justify biopsy of an endoscopically normal appearing squamocolumnar junction for the exclusion of Barrett's oesophagus and cancer risk. In the absence of contraindications, cancer risk-based therapy of dysplastic Barrett's oesophagus includes radiofrequency ablation (RFA) ± endoscopic resection. The perception of the cardia as reflux damaged DDO mirrors the concept of unfolding, as described by the interpretation of the didactic poem of Parmenides by Heidegger. Our data recommend to omit the term "cardia" and allocate morphology either to the oesophagus (CLO, DDO) or to the proximal stomach or indicate that allocation is impossible (i. e.. tumour-induced). Future studies will have to test the value of this novel concept for diagnosis, treatment of gastro-oesophageal reflux disease and cancer prevention.

P16INK4A immunohistochemistry for detection of human papilloma virus-associated penile squamous cell carcinoma is superior to in-situ hybridization.

We evaluated p16INK4A as a reliable option to detect human papilloma virus (HPV) DNA in penile tumor specimens. Formalin-fixed paraffin embedded samples of 26 patients with penile cancer and another 18 cases with non-tumorigenic lesions were stained by three different widely used commercially available chromogenic in-situ hybridization assays high-risk HPV CISH Y1443 (Genpoint, DAKO), pan HPV CISH Y1404 (Genpoint, DAKO), INFORM HPV III (Ventana, Tucson, Arizona) and p16INK4A immunohistochemistry, then compared to the known gold standard polymerase chain reaction detecting HPV 16, 18, 31, and 33. Immunoreactivity for p16INK4A was evaluated by using a 4-tiered (0, 1, 2, and 3) pattern based system. 19 cases were positive for p16INK4A, 13 of which showed a continuous transepithelial staining (pattern 3). Pan HPV ISH showed positivity in 9 cases, high-risk HPV ISH in 7 cases and INFORM HPVIII ISH in 7 cases. p16INK4A IHC pattern 3 versus pattern 0, 1 and 2 exhibited a specificity and positive predictive value of 100 percent, with a sensitivity and negative predictive value of 72 and 62 percent, respectively, which was much better than all HPV in-situ hybridization methods referred to polymerase chain reaction. p16INK4A seems to be a superior marker for the detection of HPV-associated penile squamous cell carcinoma compared to CISH tests, but is not recommend for the detection of non-tumorigenic lesions, where PCR should be used for the initial assessment.

HCRP1 expression status is a significant prognostic marker in oral and oropharyngeal cancer.

OBJECTIVE: The hepatocellular carcinoma-related protein 1 (HCRP1) is a key factor in the degradation of the epidermal growth factor receptor. In this study, we assessed the prognostic significance of HCRP1 expression in patients with oral and oropharyngeal squamous cell carcinoma (OOSCC). METHODS: HCRP1 expression was determined by immunohistochemistry on tissue biopsy sections of 111 patients with locally advanced OOSCC undergoing neoadjuvant chemoradiotherapy followed by surgery. The Kaplan-Meier method and Cox regression models were used for survival analyses. RESULTS: Low HCRP1 expression was associated with poor recurrence-free survival (P = 0.046) and overall survival (P = 0.03). Multivariate analysis revealed that low HCRP1 expression remained an independent risk factor for relapse (HR 2.98, 95% CI 1.19-7.49, P = 0.02) and death (HR 3.04, 95% CI 1.19-7.79, P = 0.02). CONCLUSION: Low HCRP1 expression was found to be of adverse prognostic significance in patients with OOSCC who received preoperative chemoradiotherapy.

KRAS status and outcome of liver resection after neoadjuvant chemotherapy including bevacizumab.

BACKGROUND: The prognostic value of KRAS mutation in patients with colorectal cancer liver metastases (CLM) receiving neoadjuvant chemotherapy including bevacizumab before liver resection is unclear. METHODS: The KRAS and BRAF status of resected CLM was assessed in prospectively studied patients. Mutations were correlated with recurrence-free and overall survival. Only patients with remaining vital tumour cells in the resected specimen and those without disease progression were analysed; those with progressive disease did not undergo resection. RESULTS: A total of 60 patients were enrolled. Fifteen (25 per cent) had a KRAS mutation, but none of the 60 patients had a BRAF mutation. The radiological response to neoadjuvant chemotherapy including bevacizumab, assessed according to the Response Evaluation Criteria In Solid Tumours, was partial in 52 patients (87 per cent) and the remaining eight had stable disease. The partial response rate was similar in patients with a KRAS mutation and those with the wild-type gene (12 of 15 versus 40 of 45 patients; P = 0·400). KRAS mutation had a negative prognostic effect on recurrence-free survival (hazard ratio (HR) 2·48, 95 per cent confidence interval 1·26 to 4·89; P = 0·009) and overall survival (HR 3·51, 1·30 to 9·45; P = 0·013). CONCLUSION: This study provided further evidence for the prognostic importance of KRAS status in terms of recurrence-free and overall survival. Neoadjuvant chemotherapy including bevacizumab elicited a response, irrespective of KRAS status, in this selected group of patients with CLM.

Prevalence and clinical significance of GB virus type C/hepatitis G virus coinfection in patients with chronic hepatitis C undergoing antiviral therapy.

Coinfection with GBV-C/HGV in patients with chronic hepatitis C (CHC) may influence clinical course and response rates of antiviral therapy. Aim of the study was to investigate the prevalence of GBV-C/HGV/HCV coinfection and its influence on outcome of interferon/ribavirin combination therapy. Three hundred and four patients with CHC [m/f = 211/93, age: 42 (18-65)] were investigated. HGV RNA detection was performed by polymerase chain reaction prior to and 6 months after the end of antiviral therapy. HGV/HCV coinfection could be identified in 37/304 (12.2%) patients with intravenous drug abuse as the most common source of infection (N = 21, (56.8%)). The predominant HCV genotype in coinfected individuals was HCV-3a (HCV-3a: 51.4%, HCV-1: 37.8%, HCV-4: 10.8%). HGV coinfection was more prevalent in patients infected with HCV-3 compared to HCV-1 or HCV-4 [19/45 (42.2%) vs. 14/185 (7.6%) vs. 4/52 (7.7%), P < 0.01]. Patients with HGV/HCV coinfection were younger [35 (18-56) vs. 43 (19-65), years; P < 0.01], and advanced fibrosis (F3-F4) was less frequent (22.2% vs. 42.9%, P < 0.05). A sustained virological response was achieved more frequently in HGV/HCV coinfected patients [26/37 (70.3%)] than in monoinfected patients [120/267 (44.9%), P < 0.01]. HGV RNA was undetectable in 65.7% of the coinfected patients at the end of follow-up. Intravenous drug abuse seems to be a major risk factor for HGV coinfection in patients with chronic hepatitis C. Coinfection with HGV does not worsen the clinical course of chronic hepatitis C or diminish response of HCV to antiviral therapy. Interferon/ribavirin combination therapy also clears HGV infection in a high proportion of cases.