Orchestration and Prognostic Significance of Immune Checkpoints in the Microenvironment of Primary and Metastatic Renal Cell Cancer.

PURPOSE: Clear cell renal cell carcinoma (ccRCC) has shown durable responses to checkpoint blockade therapies. However, important gaps persist in the understanding of its immune microenvironment. This study aims to investigate the expression and prognostic significance of immune checkpoints in primary and metastatic ccRCC, in relation with mature dendritic cells (DC) and T-cell densities. EXPERIMENTAL DESIGN: We investigated the infiltration and the localization of CD8(+) T cells and mature DC, and the expression of immune checkpoints (PD-1, LAG-3, PD-L1, and PD-L2) in relation with prognosis, in 135 primary ccRCC tumors and 51 ccRCC lung metastases. RNA expression data for 496 primary ccRCC samples were used as confirmatory cohort. RESULTS: We identify two groups of tumors with extensive CD8(+) T-cell infiltrates. One group, characterized by high expression of immune checkpoints in the absence of fully functional mature DC, is associated with increased risk of disease progression. The second group, characterized by low expression of immune checkpoints and localization of mature DC in peritumoral immune aggregates (tertiary lymphoid structures), is associated with good prognosis. CONCLUSIONS: The expression of the immune checkpoints and the localization of DC in the tumor microenvironment modulate the clinical impact of CD8(+) T cells in ccRCC.

Phase I study of the Hedgehog pathway inhibitor IPI-926 in adult patients with solid tumors.

PURPOSE: To conduct a first-in-human phase I study to determine the dose-limiting toxicities (DLT), characterize the pharmacokinetic profile, and document the antitumor activity of IPI-926, a new chemical entity that inhibits the Hedgehog pathway (HhP). EXPERIMENTAL DESIGN: Patients with solid tumors refractory to standard therapy were given IPI-926 once daily (QD) by mouth in 28-day cycles. The starting dose was 20 mg, and an accelerated titration schedule was used until standard 3 + 3 dose-escalation cohorts were implemented. Pharmacokinetics were evaluated on day -7 and day 22 of cycle 1. RESULTS: Ninety-four patients (32F, 62M; ages, 39-87) received doses ranging from 20 to 210 mg QD. Dose levels up to and including 160 mg administered QD were well tolerated. Toxicities consisted of reversible elevations in aspartate aminotransferase (AST), alanine aminotransferase (ALT) and bilirubin, fatigue, nausea, alopecia, and muscle spasms. IPI-926 was not associated with hematologic toxicity. IPI-926 pharmacokinetics were characterized by a slow absorption (T(max) = 2-8 hours) and a terminal half-life (t(1/2)) between 20 and 40 hours, supporting QD dosing. Of those HhP inhibitor-naïve patients with basal cell carcinoma (BCC) who received more than one dose of IPI-926 and had a follow-up clinical or Response Evaluation Criteria in Solid Tumors (RECIST) assessment, nearly a third (8 of 28 patients) showed a response to IPI-926 at doses ≥130 mg. CONCLUSIONS: IPI-926 was well tolerated up to 160 mg QD within 28-day cycles, which was established as the recommended phase II dose and schedule for this agent. Single-agent activity of IPI-926 was observed in HhP inhibitor-naïve patients with BCC.

Differential role of estrogen receptor beta in early versus metastatic non-small cell lung cancer.

Although women have an increased susceptibility to lung cancer, they also have a favorable clinical outcome. This may in part be due to female specific genetic and hormonal factors. In the present study, expression of ER-beta was investigated by immunohistochemistry using tissue samples from two cohorts: non-small cell lung cancer (NSCLC) diagnosed in 1999 in Manitoba and advanced NSCLC patients from the NCIC-CTG BR.18 trial. In the Manitoba cohort assessable tissue samples available in 79 patients (32 females and 47 males) and the majority (75%) had early stage disease. Fifty-one percent of patients expressed high levels of ER-beta (defined by ≥60, the median immunohistochemistry score) and its expression was comparable in males and females. The 3-year overall survival of the group was 53% and males had significantly worse survival compared to females (HR=2.37, 95%CI 1.15­4.91, P=0.02). Higher ER-beta 1 expression was associated with better survival in both univariate (HR=0.41, 95%CI 0.21­0.80, P=0.009) and in multivariate (HR=0.37, 95%CI 0.18­0.77, P=0.008) analysis. In the NCIC-CTG cohort that were more often later stage, assessable tissue samples from 48 cases were available however higher ER beta 1 expression correlated with poorer survival (HR= 1.94, 95%CI 1.01­3.75 P=0.047). These results suggest a differential impact of ER-beta 1 expression on clinical outcome by disease stage, that needs to be explored further and may explain contradictory observations reported in the literature.

CARM1 is an important determinant of ERα-dependent breast cancer cell differentiation and proliferation in breast cancer cells.

Breast cancers with estrogen receptor α (ERα) expression are often more differentiated histologically than ERα-negative tumors, but the reasons for this difference are poorly understood. One possible explanation is that transcriptional cofactors associated with ERα determine the expression of genes which promote a more differentiated phenotype. In this study, we identify one such cofactor as coactivator-associated arginine methyltransferase 1 (CARM1), a unique coactivator of ERα that can simultaneously block cell proliferation and induce differentiation through global regulation of ERα-regulated genes. CARM1 was evidenced as an ERα coactivator in cell-based assays, gene expression microarrays, and mouse xenograft models. In human breast tumors, CARM1 expression positively correlated with ERα levels in ER-positive tumors but was inversely correlated with tumor grade. Our findings suggest that coexpression of CARM1 and ERα may provide a better biomarker of well-differentiated breast cancer. Furthermore, our findings define an important functional role of this histone arginine methyltransferase in reprogramming ERα-regulated cellular processes, implicating CARM1 as a putative epigenetic target in ER-positive breast cancers.

A phosphorylation code for oestrogen receptor-alpha predicts clinical outcome to endocrine therapy in breast cancer.

To determine the relationship of the multiple sites of oestrogen receptor alpha (ERalpha) phosphorylation to clinical outcome after tamoxifen therapy, sections from tissue microarrays representing over 300 ER+ breast cancers from patients who were treated with surgery+radiation and then tamoxifen were used for immunohistochemical determination of total ERalpha, p-S104/106-ERalpha, p-S118-ERalpha, p-S167-ERalpha, p-S282-ERalpha, p-S294-ERalpha, p-T311-ERalpha and p-S559-ERalpha. Relationships of phosphorylated ERalpha to overall and relapse-free survival (RFS; breast cancer death or recurrence) were tested using single (univariate) and multiple (multivariate) predictor statistical models. Large tumour size, node positivity, high grade, progesterone receptor (PR) negative status and low levels of p-S282-ERalpha were significantly associated with reduced overall survival (OS). Along with tumour size and node status, a novel phosphorylation score (P(7) score > or = 3), taking into account all seven p-ERalpha sites, was significantly associated with reduced OS in univariate and multivariate analyses (hazard ratio (HR)=2.24, 95% confidence interval (CI) 1.15-4.34, n=335; P=0.018). Along with tumour size, node status, grade and PR status, a high P(7) score (> or = 3) was significantly associated with reduced RFS in univariate and multivariate analyses (HR=1.71, 95% CI 1.03-2.86, n=332; P=0.039). Since ERalpha is the site at which integration of diverse signals occurs to regulate breast cancer growth and survival, the ERalpha phosphorylation score may be a surrogate marker of the balance between oestrogen-dependent and crosstalk-dependent receptor activity, and is potentially a prognostic marker of clinical outcome in a tamoxifen-treated population of patients.

Estrogen receptor alpha phosphorylated at tyrosine 537 is associated with poor clinical outcome in breast cancer patients treated with tamoxifen.

A phospho-tyrosine 537-ERα (p-Y537-ERα) antibody was validated for immunohistochemistry of formalin-fixed paraffin-embedded human breast cancer biopsies and used to interrogate multiple ER positive breast cancer cases present on tissue microarrays already constructed by the Manitoba Breast Tumor Bank. Nuclear p-Y537-ERα protein expression was positively associated with positive nodes (Spearman r = 0.20, P = 0.0002) and large tumor size (r = 0.13, P = 0.02). On univariate analysis, high levels of p-Y537 were associated with poor overall survival (HR = 1.65, 95% CI 1.08-2.52, P = 0.02) but not relapse free survival from breast cancer. The association with overall survival was not significant on multivariate analysis. These data support the relevance of phosphorylation at p-Y537-ERα in human breast cancers and add further support to the presence of a phosphorylation code for ERα in human breast cancer in vivo.

The relevance of phosphorylated forms of estrogen receptor in human breast cancer in vivo.

Estrogen receptor (ER)alpha activity is regulated by phosphorylation at several sites. Recently several antibodies specific for individual phosphorylated sites within ERalpha have became available. Validation and use of these antibodies suggests that several forms of phosphorylated ERalpha can be detected in multiple ER+ human breast tumor samples, thus providing relevance for investigating the regulation and function of phosphorylated ERalpha in human breast cancer. Generally, the phosphorylated ERalpha isoforms are associated with parameters that suggest that they are markers of an intact estrogen dependent signaling pathway and better clinical outcome with respect to tamoxifen therapy. Profiling of phosphorylated ERalpha may provide better biomarkers of endocrine therapy response over and above those currently available.

Targeting the EGFR pathway for cancer therapy.

Clinical studies have shown that HER-2/Neu is over-expressed in up to one-third of patients with a variety of cancers, including B-cell acute lymphoblastic leukemia (B-ALL), breast cancer and lung cancer, and that these patients are frequently resistant to conventional chemo-therapies. Additionally, in most patients with multiple myeloma, the malignant cells over-express a number of epidermal growth factor receptors (EGFR)s and their ligands, HB-EGF and amphiregulin, thus this growth-factor family may be an important aspect in the patho-biology of this disease. These and other, related findings have provided the rationale for the targeting of the components of the EGFR signaling pathways for cancer therapy. Below we discuss various aspects of EGFR-targeted therapies mainly in hematologic malignancies, lung cancer and breast cancer. Beside novel therapeutic approaches, we also discuss specific side effects associated with the therapeutic inhibition of components of the EGFR-pathways. Alongside small inhibitors, such as Lapatinib (Tykerb, GW572016), Gefitinib (Iressa, ZD1839), and Erlotinib (Tarceva, OSI-774), a significant part of the review is also dedicated to therapeutic antibodies (e.g.: Trastuzumab/Herceptin, Pertuzumab/Omnitarg/rhuMab-2C4, Cetuximab/Erbitux/IMC-C225, Panitumumab/Abenix/ABX-EGF, and also ZD6474). In addition, we summarize, both current therapy development driven by antibody-based targeting of the EGFR-dependent signaling pathways, and furthermore, we provide a background on the history and the development of therapeutic antibodies.