Inhibitory receptors on immune cells are pivotal regulators of immune escape in cancer. Among these inhibitory receptors, CTLA-4 (targeted clinically by ipilimumab) serves as a dominant off-switch while other receptors such as PD-1 and LAG-3 seem to serve more subtle rheostat functions. However, the extent of synergy and cooperative interactions between inhibitory pathways in cancer remain largely unexplored. Here, we reveal extensive coexpression of PD-1 and LAG-3 on tumor-infiltrating CD4⁺ and CD8⁺ T cells in three distinct transplantable tumors. Dual anti–LAG-3/anti–PD-1 antibody treatment cured most mice of established tumors that were largely resistant to single antibody treatment. Despite minimal immunopathologic sequelae in PD-1 and LAG-3 single knockout mice, dual knockout mice abrogated self-tolerance with resultant autoimmune infiltrates in multiple organs, leading to eventual lethality. However, Lag3^−/−Pdcd1^−/− mice showed markedly increased survival from and clearance of multiple transplantable tumors. Together, these results define a strong synergy between the PD-1 and LAG-3 inhibitory pathways in tolerance to both self and tumor antigens. In addition, they argue strongly that dual blockade of these molecules represents a promising combinatorial strategy for cancer. Cancer Res; 72(4); 917–27. ©2011 AACR.

Global 5-hydroxymethylcytosine content is significantly reduced in tissue stem/progenitor cell compartments and in human cancers

What's known on the subject? and What does the study add? Steroid hormone receptor signals have been implicated in bladder tumourigenesis and tumour progression. The expression of androgen and/or oestrogen receptors has been assessed in bladder cancer, leading to conflicting data of expression levels and their relationship to histopathological characteristics of the tumours. We simultaneously analyze three receptors in non‐neoplastic bladder tissues as well as in primary and metastatic bladder tumour specimens. Our data demonstrate that the expression status correlates with tumour grades/stages and patients’ outcomes. OBJECTIVE: To assess the expression of the androgen receptor (AR) and oestrogen receptors (ERs) in bladder tumours because recent studies have shown conflicting results and the prognostic significance of their expression remains unclear. PATIENTS AND METHODS: We investigated the expression of AR, ERα and ERβ in 188 bladder tumour specimens, as well as matched 141 non‐neoplastic bladder and 14 lymph node metastasis tissues, by immunohistochemistry. We then evaluated the relationships between their expression and the clinicopathological features available for the present patient cohort. RESULTS: AR/ERα/ERβ was positive in 80%/50%/89% of benign urothelium, 50%/67%/41% of benign stroma, 42%/27%/49% of primary tumours and 71%/64%/71% of metastatic tumours. Significantly lower expression of AR/ERα was found in high‐grade tumours (36%/23%) and tumours invading muscularis propria (33%/19%) compared to low‐grade tumours (55%; P = 0.0232/38%; P = 0.0483) and tumours not invading muscularis propria (51%; P = 0.0181/35%; P = 0.0139), respectively. Significantly higher expression of ERβ was found in high‐grade tumours (58%) and tumours invading muscularis propria (67%) compared to low‐grade tumours (29%; P = 0.0002) and tumours not invading muscularis propria (34%; P < 0.0001), respectively. Kaplan–Meier and log‐rank tests further showed that positivity of ERβ (but not AR or ERα) was associated with the recurrence of low‐grade tumours ( P = 0.0072); the progression of low‐grade tumours ( P = 0.0005), high‐grade tumours not invading muscularis propria ( P = 0.0020) and tumours invading muscularis propria ( P = 0.0010); or disease‐specific mortality in patients with tumours invading muscularis propria ( P = 0.0073). CONCLUSIONS: Compared to benign bladders, a significant decrease in the expression of AR, ERα or ERβ in bladder cancer was seen. Loss of AR or ERα was strongly associated with higher grade/more invasive tumours, whereas ERβ expression was increased in high‐grade/invasive tumours and predicted a worse prognosis.

Background TMPRSS2-ERG fusions have been identified in about one-half of all prostatic adenocarcinomas (PCas). Fluorescence in situ hybridization (FISH) and reverse transcription polymerase chain reaction have been the most commonly used techniques in this setting. The aim of this study was to evaluate the utility of ERG immunoexpression as a surrogate for TMPRSS2-ERG fusion in a large series of PCa cases. Materials and Methods Four hundred twenty-seven radical retropubic prostatectomy tissue samples were used to construct 10 tissue microarrays (TMAs). FISH analysis was previously conducted using dual-color interphase break-apart probes for the 5′ and 3′ regions of the ERG gene. ERG expression was evaluated using a commercial rabbit anti-ERG monoclonal antibody (clone EPR3864; Epitomics, Burlingame, CA). Each TMA spot was independently assessed, and any nuclear staining positivity was considered as indicative of ERG expression. Results TMPRSS2-ERG fusions were detected by FISH in 195 (45.7%) of the PCa cases. ERG immunoexpression was found in 192 (45.0%) of the PCa cases and in none of the nontumoral tissue samples. Mean ERG H-scores were significantly higher in tumors harboring FISH-detected TMPRSS2-ERG fusions (P<0.00001), and there was a strong association between ERG immunohistochemical expression and the TMPRSS2-ERG status defined by FISH (P<0.00001), with a sensitivity of 86% (95% CI, 80%-90%) and a specificity of 89% (95% CI, 84%-93%). Receiver-operating characteristic curve analysis showed that ERG immunoexpression had a high accuracy for identifying TMPRSS2-ERG fusions detected by FISH, with an area under the curve of 0.87 (95% CI, 0.84%-0.91; P<0.00001). Conclusions We found that ERG immunohistochemical expression has a high accuracy for defining the TMPRSS-ERG fusion status. ERG immunohistochemistry may offer an accurate, simpler, and less costly alternative for evaluation of ERG fusion status in PCa than FISH.