Hypoxia-inducible factor-1 {alpha} expression predicts superior survival in patients with diffuse large B-cell lymphoma treated with R-CHOP.

PURPOSE Hypoxia-inducible factor (HIF) controls the expression of genes in response to hypoxia, as well as a wide range of other cellular processes. We previously showed constitutive stabilization of HIF-1alpha in the majority of patients with diffuse large B-cell lymphoma (DLBCL). To our knowledge, the prognostic significance of HIF in lymphoma has never been investigated. PATIENTS AND METHODS We studied the immunohistochemical protein expression of HIF-1alpha on tissue microarrays from 153 patients with DLBCL treated in sequential cohorts with cyclophosphamide, doxorubicin, oncovin, and prednisone (CHOP) or rituximab-CHOP (R-CHOP) from 1999 to 2002. Results were correlated with patient outcome. Results Median follow-up for all patients was 80 months. Among all patients, HIF-1alpha was expressed in 62% of germinal center and 59% of non-germinal center patients. With HIF-1alpha analyzed as a dependent variable, there were no survival differences in CHOP-treated patients. In the R-CHOP group, however, HIF-1alpha protein expression correlated with significantly improved progression-free survival (PFS) and overall survival (OS). Five-year PFS for HIF-1alpha-positive patients was 71% v 43% for HIF-1alpha-negative patients (P = .0187), whereas 5-year OS was 75% and 54%, respectively (P = .025). In multivariate analysis with International Prognostic Index criteria, HIF-1alpha remained a significant predictor for PFS (P = .026) and OS (P = .043). Compared with other biomarkers, HIF-1alpha correlated only with BCL6 (P = .004). In terms of gene expression, we found several common gene associations of HIF-1alpha and the stromal-1 signature with genes predominantly involved in regulation of the extracellular matrix (eg, BGN, COL1A2, COL5A1, and PLOD2). CONCLUSION The expression of HIF-1alpha protein is an important independent favorable prognostic factor for survival in patients with DLBCL treated with R-CHOP.

Nuclear factor-kappaB-mediated transforming growth factor-beta-induced expression of vimentin is an independent predictor of biochemical recurrence after radical prostatectomy.

PURPOSE: Transforming growth factor-beta (TGF-beta)-mediated epithelial-to-mesenchymal transition (EMT) has been shown to occur in some cancers; however, the pathway remains controversial and varies with different cancers. In addition, the mechanisms by which TGF-beta and the EMT contribute to prostate cancer recurrence are largely unknown. In this study, we elucidated TGF-beta-mediated EMT as a predictor of disease recurrence after therapy for prostate cancer, which has not been reported before. EXPERIMENTAL DESIGN: We analyzed TGF-beta-induced EMT using nuclear factor-kappaB (NF-kappaB) as an intermediate mediator in prostate cancer cell lines. A total of 287 radical prostatectomy specimens were evaluated using immunohistochemistry in a high-throughput tissue microarray analysis. Levels of TGF-beta signaling components and EMT-related factors were analyzed using specific antibodies. Results were expressed as the percentage of cancer cells that stained positive for a given antibody and were correlated with disease recurrence rates at a mean of 7 years following radical prostatectomy. RESULTS: In prostate cancer cell lines, TGF-beta-induced EMT was mediated by NF-kappaB signaling. Blockade of NF-kappaB or TGF-beta signaling resulted in abrogation of vimentin expression and inhibition of the invasive capability of these cells. There was high risk of biochemical recurrence associated with tumors that displayed high levels of expression of TGF-beta1, vimentin, and NF-kappaB and low level of cytokeratin 18. This was particularly true for vimentin, which is independent of patients' Gleason score. CONCLUSIONS: The detection of NF-kappaB-mediated TGF-beta-induced EMT in primary tumors predicts disease recurrence in prostate cancer patients following radical prostatectomy. The changes in TGF-beta signaling and EMT-related factors provide novel molecular markers that may predict prostate cancer outcomes following treatment.

Recommendations for collection and handling of specimens from group breast cancer clinical trials.

Recommendations for specimen collection and handling have been developed for adoption across breast cancer clinical trials conducted by the Breast International Group (BIG)-sponsored Groups and the National Cancer Institute (NCI)-sponsored North American Cooperative Groups. These recommendations are meant to promote identifiable standards for specimen collection and handling within and across breast cancer trials, such that the variability in collection/handling practices that currently exists is minimized and specimen condition and quality are enhanced, thereby maximizing results from specimen-based diagnostic testing and research. Three working groups were formed from the Cooperative Group Banking Committee, BIG groups, and North American breast cancer cooperative groups to identify standards for collection and handling of (1) formalin-fixed, paraffin-embedded (FFPE) tissue; (2) blood and its components; and (3) fresh/frozen tissue from breast cancer trials. The working groups collected standard operating procedures from multiple group specimen banks, administered a survey on banking practices to those banks, and engaged in a series of discussions from 2005 to 2007. Their contributions were synthesized into this document, which focuses primarily on collection and handling of specimens to the point of shipment to the central bank, although also offers some guidance to central banks. Major recommendations include submission of an FFPE block, whole blood, and serial serum or plasma from breast cancer clinical trials, and use of one fixative and buffer type (10% neutral phosphate-buffered formalin, pH 7) for FFPE tissue across trials. Recommendations for proper handling and shipping were developed for blood, serum, plasma, FFPE, and fresh/frozen tissue.

Hypoxia inducible factor-alpha activation in lymphoma and relationship to the thioredoxin family.

Hypoxia inducible factors (HIFs) activate oncogenic pathways, while thioredoxins (Trx), including Trx1 and Trx reductases-1 and -2 (TrxR1 and TrxR2), promote HIF-alpha stabilization. In immunoblotting studies in lymphoma cell lines we found that Raji and SUDHL4 cells exhibited normoxic HIF-2alpha protein stabilization. Five cell lines showed increased TrxR1 expression, while only Namalwa, HF1 and SUDHL4 had Trx1 and TrxR2 activation. Tissue microarrays in diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL) identified different HIF expression among histological subgroups (e.g. 44% DLBCL vs. 11% of FL cases with moderate-to-high expression of HIF-1alpha and HIF-2alpha, P = 0.0017). These data demonstrate that HIF and the thioredoxin family are abnormally activated in lymphoma.

Somatic acquisition of TGFBR1*6A by epithelial and stromal cells during head and neck and colon cancer development.

TGFBR1*6A is a common hypomorphic variant of the type I transforming growth factor (TGF)-beta receptor (TGFBR1), which transduces TGF-beta growth inhibitory signals less effectively than TGFBR1. Recent studies suggest that TGFBR1*6A confers a selective growth advantage to both normal appearing and cancerous epithelial cells in the presence of TGF-beta. We have previously shown that TGFBR1*6A is somatically acquired in head and neck and colon cancer (10). Using microdissected tissues, we show that TGFBR1*6A is somatically acquired by stromal and epithelial cells adjacent to colorectal and head and neck tumors. Somatic acquisition of the TGFBR1*6A allele is not accompanied by acquisition of other tumor-specific mutations. Furthermore, lymphocytes located within the stroma or the normal appearing epithelium do not have evidence of TGFBR1*6A acquisition. The highest TGFBR1*6A/TGFBR1 allelic ratio is observed at the tumor's edge, and traces of TGFBR1*6A are detected as far as 2 cm away from the tumor, which is suggestive of centrifugal spread of cells that harbor TGFBR1*6A. Assessment of CDH1 and CDH2 expression does not indicate epithelial-mesenchymal transformation. The results suggest that TGFBR1*6A somatic acquisition is a critical event in the early stages of cancer development that is associated with field cancerization. They also represent the first human report of somatically acquired altered stromal TGF-beta signaling during oncogenesis and the first report of a concordant mutation in the stromal and epithelial compartments in colon cancer.

Combined analysis of molecular and clinical predictors of gefitinib activity in advanced non-small cell lung cancer: epidermal growth factor receptor mutations do not tell the whole story.

BACKGROUND: Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors have been introduced in the standard therapy of non-small-cell lung cancer (NSCLC), but they benefit a minority of patients. The study of molecular markers may identify the subset of patients who are the most appropriate to treat with these agents. METHODS: We analyzed 43 patients with advanced NSCLC who were treated with gefitinib, an oral EGFR tyrosine kinase inhibitor, were included in analysis. We evaluated EGFR in tumor tissue by using immunohistochemistry and fluorescence in situ hybridization. We also studied downstream molecules (AKT, ERK, p38 MAPK) and their activation status and the presence of EGFR mutations in tumor tissue in exons 18-21. RESULTS: Three patients had tumors with EGFR mutations, all of which had EGFR gene amplification with a ratio of 2 or greater (p= 0.001). There was no correlation between EGFR protein expression and gene amplification. Six patients (14%) achieved an objective response and nine (21%) had stable disease; the median survival was 162 days. EGFR mutations, high levels of AKT protein expression, rash of any grade, and no history of smoking were predictive of disease control (objective response plus stable disease). Only 3 of 15 patients (20%) with disease control had an EGFR mutation. On multivariate analysis, rash and AKT were independent predictors of disease control. Patients with rash survived longer than patients without rash. CONCLUSIONS: EGFR mutation-positive tumors are present in a small fraction of patients who achieve disease control with gefitinib. Other molecular markers, such as AKT, need to be further evaluated. Clinical parameters remain major determinants of gefitinib activity in NSCLC.

Phase II trial of arsenic trioxide in relapsed and refractory acute myeloid leukemia, secondary leukemia and/or newly diagnosed patients at least 65 years old.

The prognosis for patients with relapsed/refractory AML, secondary leukemia and AML in older adults is extremely poor. An appealing alternative approach to intensive cytotoxic chemotherapy is to induce apoptosis with a novel agent. There is in vitro evidence that arsenic trioxide (ATO) has anti-proliferative and pro-apoptotic effects on myeloid leukemia cell lines. To evaluate efficacy and toxicities of ATO, we conducted a phase II trial including subjects with relapsed/refractory or secondary AML or age > or = 65 years with de novo disease. Eleven subjects were entered with a median age of 77 years (56-90) and a median total dose of ATO of 415.55 mg (91.5-793) with a daily dose of 0.25 mg/kg. Median survival following the first dose of ATO was 2.25 months (0.4-19). Myelosuppression was the major adverse effect, most likely due to disease progression rather than drug-related. All subjects had progressive disease. There was no direct treatment-related mortality. Based on this study, we do not recommend single agent ATO as a treatment option for AML.