Genomic characterization of HER2-positive breast cancer and response to neoadjuvant trastuzumab and chemotherapy-results from the ACOSOG Z1041 (Alliance) trial.

Background: HER2 (ERBB2) gene amplification and its corresponding overexpression are present in 15-30% of invasive breast cancers. While HER2-targeted agents are effective treatments, resistance remains a major cause of death. The American College of Surgeons Oncology Group Z1041 trial (NCT00513292) was designed to compare the pathologic complete response (pCR) rate of distinct regimens of neoadjuvant chemotherapy and trastuzumab, but ultimately identified no difference. Patients and methods: In supplement to tissues from 37 Z1041 cases, 11 similarly treated cases were obtained from a single institution study (NCT00353483). We have extracted genomic DNA from both pre-treatment tumor biopsies and blood of these 48 cases, and performed whole genome (WGS) and exome sequencing. Coincident with these efforts, we have generated RNA-seq profiles from 42 of the tumor biopsies. Among patients in this cohort, 24 (50%) achieved a pCR. Results: We have characterized the genomic landscape of HER2-positive breast cancer and investigated associations between genomic features and pCR. Cases assigned to the HER2-enriched subtype by RNA-seq analysis were more likely to achieve a pCR compared to the luminal, basal-like, or normal-like subtypes (19/27 versus 3/15; P = 0.0032). Mutational events led to the generation of putatively active neoantigens, but were overall not associated with pCR. ERBB2 and GRB7 were the genes most commonly observed in fusion events, and genomic copy number analysis of the ERBB2 locus indicated that cases with either no observable or low-level ERBB2 amplification were less likely to achieve a pCR (7/8 versus 17/40; P = 0.048). Moreover, among cases that achieved a pCR, tumors consistently expressed immune signatures that may contribute to therapeutic response. Conclusion: The identification of these features suggests that it may be possible to predict, at the time of diagnosis, those HER2-positive breast cancer patients who will not respond to treatment with chemotherapy and trastuzumab. ClinicalTrials.gov identifiers: NCT00513292, NCT00353483.

Epigenetic activation of the prostaglandin receptor EP4 promotes resistance to endocrine therapy for breast cancer.

Approximately 75% of breast cancers express estrogen receptor α (ERα) and depend on estrogen signals for continued growth. Aromatase inhibitors (AIs) prevent estrogen production and inhibit ER signaling, resulting in decreased cancer recurrence and mortality. Advanced tumors treated with AIs almost always develop resistance to these drugs via the upregulation of alternative growth signals. The mechanisms that drive this resistance-especially epigenetic events that alter gene expression-are, however, not well understood. Genome-wide DNA methylation and expression analysis of cell line models of acquired AI resistance indicated that prostaglandin E2 receptor 4 (PTGER4) is upregulated after demethylation in resistant cells. Knockdown and inhibitor studies demonstrate that PTGER4 is essential for estrogen-independent growth. Our exploratory analysis of downstream signaling indicates that PTGER4 likely promotes AI resistance via ligand-independent activation of the ERα-cofactor CARM1. We believe that we have discovered a novel epigenetic mechanism for altering cell signaling and acquiring endocrine therapy resistance. Our findings indicate that PTGER4 is a potential drug target in AI-resistant cancers. In addition, the epigenetic component of PTGER4 regulation suggests that further study of PTGER4 may yield valuable insights into how DNA methylation-targeted diagnoses and treatments can improve AI-resistant breast cancer treatment.

The Z-cad dual fluorescent sensor detects dynamic changes between the epithelial and mesenchymal cellular states.

BACKGROUND: The epithelial to mesenchymal transition (EMT) has been implicated in metastasis and therapy resistance of carcinomas and can endow cancer cells with cancer stem cell (CSC) properties. The ability to detect cancer cells that are undergoing or have completed EMT has typically relied on the expression of cell surface antigens that correlate with an EMT/CSC phenotype. Alternatively these cells may be permanently marked through Cre-mediated recombination or through immunostaining of fixed cells. The EMT process is dynamic, and these existing methods cannot reveal such changes within live cells. The development of fluorescent sensors that mirror the dynamic EMT state by following the expression of bona fide EMT regulators in live cells would provide a valuable new tool for characterizing EMT. In addition, these sensors will allow direct observation of cellular plasticity with respect to the epithelial/mesenchymal state to enable more effective studies of EMT in cancer and development. RESULTS: We generated a lentiviral-based, dual fluorescent reporter system, designated as the Z-cad dual sensor, comprising destabilized green fluorescent protein containing the ZEB1 3' UTR and red fluorescent protein driven by the E-cadherin (CDH1) promoter. Using this sensor, we robustly detected EMT and mesenchymal to epithelial transition (MET) in breast cancer cells by flow cytometry and fluorescence microscopy. Importantly, we observed dynamic changes in cellular populations undergoing MET. Additionally, we used the Z-cad sensor to identify and isolate minor subpopulations of cells displaying mesenchymal properties within a population comprising predominately epithelial-like cells. The Z-cad dual sensor identified cells with CSC-like properties more effectively than either the ZEB1 3' UTR or E-cadherin sensor alone. CONCLUSIONS: The Z-cad dual sensor effectively reports the activities of two factors critical in determining the epithelial/mesenchymal state of carcinoma cells. The ability of this stably integrating dual sensor system to detect dynamic fluctuations between these two states through live cell imaging offers a significant improvement over existing methods and helps facilitate the study of EMT/MET plasticity in response to different stimuli and in cancer pathogenesis. Finally, the versatile Z-cad sensor can be adapted to a variety of in vitro or in vivo systems to elucidate whether EMT/MET contributes to normal and disease phenotypes.

Predicting response and survival in chemotherapy-treated triple-negative breast cancer.

BACKGROUND: In this study, we evaluated the ability of gene expression profiles to predict chemotherapy response and survival in triple-negative breast cancer (TNBC). METHODS: Gene expression and clinical-pathological data were evaluated in five independent cohorts, including three randomised clinical trials for a total of 1055 patients with TNBC, basal-like disease (BLBC) or both. Previously defined intrinsic molecular subtype and a proliferation signature were determined and tested. Each signature was tested using multivariable logistic regression models (for pCR (pathological complete response)) and Cox models (for survival). Within TNBC, interactions between each signature and the basal-like subtype (vs other subtypes) for predicting either pCR or survival were investigated. RESULTS: Within TNBC, all intrinsic subtypes were identified but BLBC predominated (55-81%). Significant associations between genomic signatures and response and survival after chemotherapy were only identified within BLBC and not within TNBC as a whole. In particular, high expression of a previously identified proliferation signature, or low expression of the luminal A signature, was found independently associated with pCR and improved survival following chemotherapy across different cohorts. Significant interaction tests were only obtained between each signature and the BLBC subtype for prediction of chemotherapy response or survival. CONCLUSIONS: The proliferation signature predicts response and improved survival after chemotherapy, but only within BLBC. This highlights the clinical implications of TNBC heterogeneity, and suggests that future clinical trials focused on this phenotypic subtype should consider stratifying patients as having BLBC or not.

Concordance among gene expression-based predictors for ER-positive breast cancer treated with adjuvant tamoxifen.

BACKGROUND: ER-positive (ER+) breast cancer includes all of the intrinsic molecular subtypes, although the luminal A and B subtypes predominate. In this study, we evaluated the ability of six clinically relevant genomic signatures to predict relapse in patients with ER+ tumors treated with adjuvant tamoxifen only. METHODS: Four microarray datasets were combined and research-based versions of PAM50 intrinsic subtyping and risk of relapse (PAM50-ROR) score, 21-gene recurrence score (OncotypeDX), Mammaprint, Rotterdam 76 gene, index of sensitivity to endocrine therapy (SET) and an estrogen-induced gene set were evaluated. Distant relapse-free survival (DRFS) was estimated by Kaplan-Meier and log-rank tests, and multivariable analyses were done using Cox regression analysis. Harrell's C-index was also used to estimate performance. RESULTS: All signatures were prognostic in patients with ER+ node-negative tumors, whereas most were prognostic in ER+ node-positive disease. Among the signatures evaluated, PAM50-ROR, OncotypeDX, Mammaprint and SET were consistently found to be independent predictors of relapse. A combination of all signatures significantly increased the performance prediction. Importantly, low-risk tumors (>90% DRFS at 8.5 years) were identified by the majority of signatures only within node-negative disease, and these tumors were mostly luminal A (78%-100%). CONCLUSIONS: Most established genomic signatures were successful in outcome predictions in ER+ breast cancer and provided statistically independent information. From a clinical perspective, multiple signatures combined together most accurately predicted outcome, but a common finding was that each signature identified a subset of luminal A patients with node-negative disease who might be considered suitable candidates for adjuvant endocrine therapy alone.

Neoadjuvant endocrine therapy in primary breast cancer: indications and use as a research tool.

Neoadjuvant endocrine therapy has been increasingly employed in clinical practice to improve surgical options for postmenopausal women with bulky hormone receptor-positive breast cancer. Recent studies indicate that tumour response in this setting may predict long-term outcome of patients on adjuvant endocrine therapy, which argues for its broader application in treating hormone receptor-positive disease. From the research perspective, neoadjuvant endocrine therapy provides a unique opportunity for studies of endocrine responsiveness and the development of novel therapeutic agents.

Chronic kidney disease after hematopoietic cell transplantation: a systematic review.

Advances in hematopoietic cell transplantation (HCT) have broadened its indications for use and resulted in more long-term HCT survivors. Some survivors develop chronic kidney disease (CKD); however, the incidence and risk factors are unclear. We performed a systematic review of studies identified from databases (MEDLINE, EMBASE, Science Citation Index), conference abstracts and reference lists from selected manuscripts. From 927 manuscripts, 28 patient cohorts were identified in which 9317 adults and children underwent HCT and 7317 (79%) survived to at least 100 days, permitting inclusion of 5337 (73% of survivors) in quantitative analyses. Although definitions and measurements varied widely, approximately 16.6% of HCT patients developed CKD and estimated glomerular filtration rate (eGFR in mL/min/1.73 m(2)) decreased by 24.5 after 24 months. This decrease was greater amongst patients undergoing allogeneic HCT (DeltaeGFR = -40.0 versus -18.6 for autologous transplants). Several commonly reported risk factors for CKD were investigated, including acute renal failure, total body irradiation, graft versus host disease and long-term cyclosporine use. In conclusion, CKD following HCT is likely to be common; however, prospective studies with uniform definitions of CKD and risk factors are needed to confirm these findings and better define the underlying mechanisms to promote therapies that prevent this complication.

First-line endocrine treatment of breast cancer: aromatase inhibitor or antioestrogen?

Until recently, endocrine therapy for breast cancer was relatively simple. If the tumour expressed hormone receptors, regardless of stage and age, tamoxifen was indicated. While this largely remains the case for premenopausal women, clinical trials in postmenopausal women have broadened our choice to include one of three selective aromatase inhibitors (AIs), the nonsteroidal agents anastrozole or letrozole and the steroidal agent exemestane. Comparative data concerning the efficacy, toxicity, tolerability and cost of AI vs tamoxifen continues to evolve with over 40 000 women slated to be involved in clinical trials. Currently, tamoxifen remains an appropriate choice for adjuvant treatment, and will remain so unless a clear survival advantage emerges for adjuvant AI therapy. However, anastrozole is widely seen as a useful alternative, with particular merit for patients prone to the development of serious tamoxifen side effects. For endocrine therapy naïve advanced disease, several trials have provided evidence that a nonsteroidal AI has replaced tamoxifen as optimal treatment. In the neoadjuvant setting, letrozole was also more effective than tamoxifen, both in terms of response rates and the incidence of breast-conserving surgery, and so AI therefore also dominates this evolving indication. The ongoing adjuvant clinical trials ask all the relevant questions regarding tamoxifen and AI in combination, sequence and duration, except for 5 years of an AI vs a longer period. For both the advanced and early-stage disease, resistance remains the key obstacle to overcome, and trials that combine endocrine agents with signal transduction inhibitors such as HER1 and HER2 kinase inhibitors, farnesyl transferase inhibitors, mTOR inhibitors as well as COX2 inhibitors are being developed in a concerted attempt to address this problem.

Letrozole is more effective neoadjuvant endocrine therapy than tamoxifen for ErbB-1- and/or ErbB-2-positive, estrogen receptor-positive primary breast cancer: evidence from a phase III randomized trial.

PURPOSE: Expression of ErbB-1 and ErbB-2 (epidermal growth factor receptor and HER2/neu) in breast cancer may cause tamoxifen resistance, but not all studies concur. Additionally, the relationship between ErbB-1 and ErbB-2 expression and response to selective aromatase inhibitors is unknown. A neoadjuvant study for primary breast cancer that randomized treatment between letrozole and tamoxifen provided a context within which these issues could be addressed prospectively. PATIENTS AND METHODS: Postmenopausal patients with estrogen- and/or progesterone receptor-positive (ER+ and/or PgR+) primary breast cancer ineligible for breast-conserving surgery were randomly assigned to 4 months of neoadjuvant letrozole 2.5 mg daily or tamoxifen 20 mg daily in a double-blinded study. Immunohistochemistry (IHC) for ER and PgR was conducted on pretreatment biopsies and assessed by the Allred score. ErbB-1 and ErbB-2 IHC were assessed by intensity and completeness of membranous staining according to published criteria. RESULTS: For study biopsy-confirmed ER+ and/or PgR+ cases that received letrozole, 60% responded and 48% underwent successful breast-conserving surgery. The response to tamoxifen was inferior (41%, P =.004), and fewer patients underwent breast conservation (36%, P =.036). Differences in response rates between letrozole and tamoxifen were most marked for tumors that were positive for ErbB-1 and/or ErbB-2 and ER (88% v 21%, P =.0004). CONCLUSION: ER+, ErbB-1+, and/or ErbB-2+ primary breast cancer responded well to letrozole, but responses to tamoxifen were infrequent. This suggests that ErbB-1 and ErbB-2 signaling through ER is ligand-dependent and that the growth-promoting effects of these receptor tyrosine kinases on ER+ breast cancer can be inhibited by potent estrogen deprivation therapy.

Effect of anticoagulants and storage temperatures on stability of plasma and serum hormones.

OBJECTIVES: To determine the effect of different anticoagulants and storage conditions on the stability of hormones in plasma and serum. DESIGN AND METHODS: Human blood samples were collected from volunteers into EDTA, lithium heparin, sodium fluoride/potassium oxalate, or tubes without anticoagulant, plasma and serum left at -20 degrees C, 4 degrees C or 30 degrees C for 24 and 120 hours then assayed for ACTH, aldosterone, alpha-subunit, AVP, CRH, C-peptide, estradiol, FSH, glucagon, GH, IGF-1, IGFBP-3, insulin, leptin, LH, PPP, PTH, prolactin and VIP, or at room temperature for 0 to 72 hours (BNP, NT-BNP)(n = 6 per condition). RESULTS: The anticoagulant altered the measured concentrations for 9 hormones when compared to EDTA. All hormones except ACTH were stable for > 120 hours in EDTA or fluoride at 4 degrees C, but only 13 hormones were stable in all anticoagulants. At 30 degrees C, 8 hormones were stable for > 120 hours in EDTA, and 3 hormones in all anticoagulants. BNP and NT-BNP were stable for < 24 hours when stored in EDTA or heparin at room temperature. DISCUSSION: Storage of samples in EDTA plasma at 4 degrees C is suitable for most hormones (except ACTH) for up to 120 hours.

Genes that co-cluster with estrogen receptor alpha in microarray analysis of breast biopsies.

The estrogen receptor plays a critical role in the pathogenesis and clinical behavior of breast cancer. To better understand the molecular basis of estrogen-dependent forms of this disease we studied gene expression profiles from 53 primary breast cancer biopsies. Gene expression data for more than 7000 genes were generated from each tumor sample with oligo microarrays. A standard correlation-clustering algorithm identified 18 genes that co-clustered with estrogen receptor alpha. Eleven of these genes had previously been associated with estrogen regulation or breast tumorigenesis including trefoil factor 1 and estrogen regulated LIV-1. Additional study of these 18 genes may further delineate the role of estrogen receptor in breast cancer, generate new predictive biomarkers for response to endocrine therapies and identify novel therapeutic targets.

Neoadjuvant endocrine therapy for breast cancer: medical perspectives.

The indolent nature of estrogen-dependent breast cancer is the most important obstacle for development of new adjuvant endocrine treatments. Clinical trials require thousands of study participants and at least a decade of clinical investigation. How can we be sure that a new endocrine agent warrants this extraordinary level of investment? Traditionally, we have relied on advanced breast cancer trials to determine which drugs are suitable for adjuvant studies. However, with endocrine agents the high incidence of resistance in metastatic breast cancer may mask important advances in efficacy. Recent clinical results with the aromatase inhibitor letrozole suggest that neoadjuvant endocrine therapy is a highly informative additional approach to consider when planning adjuvant studies. In this report, new neoadjuvant endocrine therapy study designs are discussed that address the following issues: (a) the scientific opportunities afforded by gene microarray studies and other genetic technologies to investigate the molecular basis of estrogen-dependent breast cancer; (b) studies that address critical drug development questions as a prelude to adjuvant studies; and (c) the conduct of randomized trials that compare neoadjuvant chemotherapy with neoadjuvant aromatase inhibitor therapy to establish a place for neoadjuvant endocrine therapy in routine clinical practice.

Preoperative endocrine therapy for older women with breast cancer: renewed interest in an old idea.

BACKGROUND: Tamoxifen as sole therapy (primary tamoxifen therapy) was investigated in the 1970s and 1980s as an alternative to surgery for older patients with breast cancer. While the majority of primary breast tumors responded to tamoxifen, long-term local disease control was poor. The use of primary tamoxifen therapy is therefore restricted to frail, elderly, and infirm patients who cannot tolerate surgery. In contrast, short-term preoperative endocrine therapy to downstage estrogen receptor-positive (ER+) tumors is under increasing scrutiny as a nontoxic neoadjuvant approach for older women. METHODS: The literature on primary tamoxifen therapy and preoperative endocrine therapy was reviewed to construct an opinion piece on the feasibility and safety of preoperative endocrine therapy. RESULTS: A review of nine phase II trials and a meta-analysis of two randomized trials suggest that the initial response rates to preoperative endocrine therapy will exceed 50% to 60% for patients with ER+ disease. A short delay in surgery to administer 3 to 4 months of preoperative endocrine therapy is unlikely to compromise long-term outcomes. CONCLUSIONS: Preoperative endocrine therapy is a logical approach for older patients with ER+ disease as a well-tolerated means to increase the rate of breast-conserving surgery. Several clinical trials comparing tamoxifen with selective aromatase inhibitors in the preoperative setting have been conducted, and the results are expected soon. These studies will determine if a large multicenter national trial of preoperative endocrine therapy should be conducted.

Mannose 6-phosphate/insulin-like growth factor 2 receptor, a bona fide tumor suppressor gene or just a promising candidate?

The mannose 6-phosphate/insulin-like growth factor 2 receptor (M6P/IGF2R) is considered a "candidate" tumor suppressor gene. This hypothesis has been provoked by the identification of loss of heterozygosity (LOH) at the M6P/IGF2R locus on chromosome 6q26 in breast and liver cancer, accompanied by point mutations in the remaining allele. Somatic mutations in coding region microsatellites have also been described in replication error positive (RER+) tumors of the gastrointestinal tract, endometrium and brain. These genetic data are compelling, but a tumor suppressor gene candidate has to meet functional as well as genetic criteria. This review weighs the evidence and discusses the observations that are necessary to promote M6P/IGF2R from candidate to bona fide tumor suppressor gene.

Antitumor polycyclic acridines. 7. Synthesis and biological properties of DNA affinic tetra- and pentacyclic acridines.

New synthetic routes to a series of tetra- and pentacyclic acridines related in structure to marine natural products are reported. The novel water-soluble agent dihydroindolizino[7,6,5-kl]acridinium chloride 14 has inhibitory activity in a panel of non-small-cell lung and breast tumor cell lines exceeding that of m-AMSA. The salt inhibited the release of minicircle products of kDNA confirming that disorganization of topoisomerase II partly underlies the activity of the compound. COMPARE analysis of the NCI mean graph profile of compound 14 at the GI(50) level corroborates this conclusion with Pearson correlation coefficients (>0.6) to clinical agents of the topoisomerase II class: however, this correlation was not seen at the LC(50) level. The inhibitory action of 14 on Saccharomyces cerevisiae transfected with human topoisomerase II isoforms showed a 3-fold selectivity against the IIalpha isoform over the IIbeta isoform. Unlike m-AMSA, 14 is not susceptible to P-glycoprotein-mediated drug efflux and retains activity in lung cells with derived resistance to the topoisomerase II inhibitor etoposide.

A pilot trial assessing the efficacy of paroxetine hydrochloride (Paxil) in controlling hot flashes in breast cancer survivors.

BACKGROUND: Many breast cancer survivors suffer debilitating hot flashes. Estrogen, the drug of choice in perimenopausal women, is generally not recommenced to breast cancer survivors. Nonhormonal treatments are mostly disappointing. Anecdotal reports in our institution suggested that the selective serotonin-reuptake inhibitor, paroxetine hydrochloride, might be efficacious in alleviating hot flashes. PATIENTS AND METHODS: Thirty women with prior breast cancer who were suffering at least two hot flashes a day entered a single institution pilot trial to evaluate paroxetine's efficacy in reducing the frequency and severity of hot flashes. After completing daily diaries for one week on no therapy, the women received open-label paroxetine, 10 mg daily for one week, followed by four weeks of paroxetine, 20 mg daily. The women completed hot-flash daily diaries throughout the study period, and a health-related symptom-assessment questionnaire and a quality-of-life rating scale in the first and sixth week of the study. RESULTS: Twenty-seven women completed the six-week study period. The mean reduction of hot flash frequency was 67% (95% confidence interval (95% CI): 56%-79%). The mean reduction in hot flash severity score was 75% (95% CI: 66%-85%). There was a statistically significant improvement in depression, sleep, anxiety, and quality of life scores. Furthermore, 25 (83%) of the study participants chose to continue paroxetine therapy at the end of study. The most common adverse effect was somnolence, resulting in drug discontinuation in two women, and dose reduction in two women. One woman discontinued drug due to anxiety. CONCLUSIONS: Paroxetine hydrochloride is a promising new treatment for hot flashes in breast cancer survivors, and warrants further evaluation in a double-blind randomized placebo-controlled trial.

Activation of the transforming growth factor beta signaling pathway and induction of cytostasis and apoptosis in mammary carcinomas treated with the anticancer agent perillyl alcohol.

The mechanisms of action of the anticancer agent perillyl alcohol (POH), presently in Phase II clinical trials, were investigated in advanced rat mammary carcinomas. Gross and ultrastructural morphology of POH-mediated tumor regression indicated that apoptosis accounted for the marked reduction in the epithelial compartment. Characterization of cell growth and death indices revealed that apoptosis was induced within 48 h of chemotherapy, before the induction of cytostasis. RNA expression studies, based on a multiplexed-nuclease protection assay, demonstrated that cell cycle- and apoptosis-related genes were differentially expressed within 48 h of POH treatment; p21(Cip1/WAF1), bax, bad, and annexin I were induced; cyclin E and cyclin-dependent kinase 2 were repressed; and bcl-2 and p53 were unchanged. Next, a potential role for transforming growth factor beta (TGF-beta) signaling in POH-mediated carcinoma regression was explored. RNA expression studies, again based on a multiplexed-nuclease protection assay, showed that TGF-beta-related genes were induced and temporally regulated during POH treatment: (a) c-jun and c-fos were transiently induced within 12 h of chemotherapy; (b) TGF-beta1 was induced within 24 h of chemotherapy; (c) the mannose 6-phosphate/insulin-like growth factor II receptor and the TGF-beta type I and II receptors were induced within 48 h of chemotherapy; and (d) smad3 was induced during active carcinoma regression. In situ protein expression studies, based on fluorescence-immunohistochemistry in concert with confocal microscopy, confirmed up-regulation and demonstrated colocalization of TGF-beta1, the mannose 6-phosphate/insulin-like growth factor II receptor, the TGF-beta type I and II receptors, and Smad2/Smad3 in epithelial cells. Nuclear localization of Smad2/Smad3 indicated that the TGF-beta signaling pathway was activated in regressing carcinomas. Subpopulations of Smad2/Smad3-positive and apoptotic nuclei colocalized, indicating a role for Smads in apoptosis. Thus, Smads may serve as a potential biomarker for anticancer activity. Importantly, none of the POH-mediated anticancer activities were observed in normal mammary gland.

The RIIbeta regulatory subunit of protein kinase A binds to cAMP response element: an alternative cAMP signaling pathway.

cAMP, through the activation of cAMP-dependent protein kinase (PKA), is involved in transcriptional regulation. In eukaryotic cells, cAMP is not considered to alter the binding affinity of CREB/ATF to cAMP-responsive element (CRE) but to induce serine phosphorylation and consequent increase in transcriptional activity. In contrast, in prokaryotic cells, cAMP enhances the DNA binding of the catabolite repressor protein to regulate the transcription of several operons. The structural similarity of the cAMP binding sites in catabolite repressor protein and regulatory subunit of PKA type II (RII) suggested the possibility of a similar role for RII in eukaryotic gene regulation. Herein we report that RIIbeta subunit of PKA is a transcription factor capable of interacting physically and functionally with a CRE. In contrast to CREB/ATF, the binding of RIIbeta to a CRE was enhanced by cAMP, and in addition, RIIbeta exhibited transcriptional activity as a Gal4-RIIbeta fusion protein. These experiments identify RIIbeta as a component of an alternative pathway for regulation of CRE-directed transcription in eukaryotic cells.

The mannose 6-phosphate/insulin-like growth factor 2 receptor (M6P/IGF2R), a putative breast tumor suppressor gene.

Loss of heterozygosity (LOH) at the mannose 6-phosphate/insulin-like growth factor 2 receptor gene locus (M6P/IGF2R) on 6q26-27 has recently been demonstrated in approximately 30% of both invasive and in situ breast cancers. LOH was coupled with somatic point mutations in the remaining allele in several instances, leading to the proposition that M6P/IGF2R is a tumor suppressor gene. Somatic mutations in M6P/IGF2R have also been described in hepatoma and gastrointestinal cancers with the replication error positive (RER+) phenotype. These data indicate that M6P/IGF2R loss of function mutations may be involved in the pathogenesis of a wide spectrum of malignancies. Extensive data on the normal function of the M6P/IGF2R suggest that loss of M6P/IGF2R activity may contribute to multiple aspects of tumor pathophysiology, including deregulated growth, apoptosis, angiogenesis and invasion.