Antreas Hindoyan

Integrative Survival-Based Molecular Profiling of Human Pancreatic Cancer

Purpose: To carry out an integrative profile of human pancreatic ductal adenocarcinoma (PDAC) to identify prognosis-significant genes and their related pathways. Experimental Design: A concordant survival-based whole genome in silico array analysis of DNA copy number, and mRNA and miRNA expression in 25 early-stage PDAC was carried out. A novel composite score simultaneously integrated gene expression with regulatory mechanisms to identify the signature genes with the most levels of prognosis-significant evidence. The predominant signaling pathways were determined via a pathway-based approach. Independent patient cohorts (n = 148 and 42) were then used as in vitro validation of the array findings. Results: The composite score identified 171 genes in which expressions were able to define two prognosis subgroups (P = 3.8e-5). Eighty-eight percent (151 of 171) of the genes were regulated by prognosis-significant miRNAs. The phosphoinositide 3-kinase/AKT pathway and SRC signaling were densely populated by prognosis-significant genes and driven by genomic amplification of SRC and miRNA regulation of p85α and CBL. On tissue microarray validation (n = 148), p85α protein expression was associated with improved survival for all patients (P = 0.02), and activated P-SRC (Y418) was associated shorter survival for patients with low-grade histology tumors (P = 0.04). Interacting P-SRC and p85α revealed that they define two distinct PDAC patient subgroups (P = 0.0066). Furthering the importance of these pathways, CBL protein expression was associated with improved survival (P = 0.03) on a separate cohort (n = 42). Conclusions: These pathways and related genes may represent putative clinical biomarkers and possible targets of individualized therapy in the distinct patient subgroups they define. Clin Cancer Res; 18(5); 1352–63. ©2012 AACR.

Identification of CD166 as a Surface Marker for Enriching Prostate Stem/Progenitor and Cancer Initiating Cells

New therapies for late stage and castration resistant prostate cancer (CRPC) depend on defining unique properties and pathways of cell sub-populations capable of sustaining the net growth of the cancer. One of the best enrichment schemes for isolating the putative stem/progenitor cell from the murine prostate gland is Lin-;Sca1+;CD49fhi (LSChi), which results in a more than 10-fold enrichment for in vitro sphere-forming activity. We have shown previously that the LSChi subpopulation is both necessary and sufficient for cancer initiation in the Pten-null prostate cancer model. To further improve this enrichment scheme, we searched for cell surface molecules upregulated upon castration of murine prostate and identified CD166 as a candidate gene. CD166 encodes a cell surface molecule that can further enrich sphere-forming activity of WT LSChi and Pten null LSChi. Importantly, CD166 could enrich sphere-forming ability of benign primary human prostate cells in vitro and induce the formation of tubule-like structures in vivo. CD166 expression is upregulated in human prostate cancers, especially CRPC samples. Although genetic deletion of murine CD166 in the Pten null prostate cancer model does not interfere with sphere formation or block prostate cancer progression and CRPC development, the presence of CD166 on prostate stem/progenitors and castration resistant sub-populations suggest that it is a cell surface molecule with the potential for targeted delivery of human prostate cancer therapeutics.

Expression Of GRP78, Master Regulator Of The Unfolded Protein Response, Increases Chemoresistance In Pancreatic Ductal Adenocarcinoma

The prognosis for patients with pancreatic ductal adenocarcinoma (PDAC) is dismal. Although gemcitabine (GEM) is the standard chemotherapeutic agent for adjuvant therapy of resectable PDAC, recurrent disease is observed in an alarming number of GEM-treated patients. Regardless of the adjuvant therapy, the vast majority of patients treated with chemotherapy after surgical resection show tumor recurrence. A better understanding of the molecular mechanisms that contribute to chemoresistance would aid the development of more effective treatment strategies. GRP78 is an endoplasmic reticulum (ER) chaperone protein that primarily resides in the lumen of the ER and is the master regulator of the unfolded protein response (UPR). Here, we report that expression of GRP78 is significantly higher in GEM-resistant PDAC compared to GEM-sensitive PDAC patient samples. We show that GRP78 induces chemoresistance in PDAC cells. Our results also show that knockdown of GRP78 reduces chemoresistance in PDAC. Finally, we found that IT-139, a ruthenium-based anticancer drug, can overcome GRP78-mediated chemoresistance. In vitro, IT-139 restores sensitivity to cytotoxic drugs in drug-resistant PDAC cells and induces twice as much cell death in combination treatment compared with GEM alone. In vivo, a single weekly IT-139 treatment in combination with GEM caused a 35% increase in median survival and a 25% increase in overall survival compared to GEM alone. Collectively, our data show that GRP78 expression promotes chemoresistance in PDAC and therapeutic strategies, blocking the activity of GRP78 increases the efficacy of currently available therapies. Mol Cancer Ther; 15(5); 1043–52. ©2016 AACR.

Anomalous coronary arteries: cardiovascular computed tomographic angiography for surgical decisions and planning.

Cardiovascular computed tomographic angiography (CCTA) provides an understanding of the three-dimensional (3D) coronary artery anatomy in relation to cardiovascular thoracic structures important to the surgical management of anomalous coronary arteries (ACAs). Although some ACA variants are not clinically significant, others can lead to ischemia/infarction, related acute ventricular dysfunction, ventricular arrhythmias, and sudden cardiac death. The CCTA is important to surgical decision making, as it provides noninvasive visualization of the coronary arteries with (1) assessment of origin, course, and termination of coronary artery anomalies in the context of 3D thoracic anatomy, (2) characterization of anatomy helpful for differentiation of benign versus hemodynamically significant variants, (3) identification of other cardiothoracic anomalies, and (4) detection of coronary artery disease. High-risk ACA anatomy in the appropriate clinical setting can require surgical intervention with decisions including minimally invasive versus open sternotomy approach, correction via reimplantation of a coronary artery, alteration of the ACA course without reimplantation, or bypass of an ACA. Given the rarity of ACA, there is limited data in the literature, and significant controversy related to the management issues. The management of ACA requires comprehensive clinical history, thorough assessment of cardiac function, and detailed anatomic imaging. Future studies will need to address the long-term outcome based on detailed assessment of original anatomy and surgical approach.

Escherichia coli HU protein has a role in the repair of abasic sites in DNA.

HU is one of the most abundant DNA binding proteins in Escherichia coli. We find that it binds strongly to DNA containing an abasic (AP) site or tetrahydrofuran (THF) (apparent Kd ≈50 nM). It also possesses an AP lyase activity that cleaves at a deoxyribose but not at a THF residue. The binding and cleavage of an AP site was observed only with the HUαβ heterodimer. Site-specific mutations at K3 and R61 residues led to a change in substrate binding and cleavage. Both K3A(α)K3A(β) and R61A(α)R61A(β) mutant HU showed significant reduction in binding to DNA containing AP site; however, only R61A(α)R61A(β) mutant protein exhibited significant loss in AP lyase activity. Both K3A(α)K3A(β) and R61K(α)R61K(β) showed slight reduction in AP lyase activities. The function of HU protein as an AP lyase was confirmed by the ability of hupA or hupB mutations to further reduce the viability of an E. coli dut(Ts) xth mutant, which generates lethal AP sites at 37°C; the hupA and hupB derivatives, respectively, had a 6-fold and a 150-fold lower survival at 37°C than did the parental strain. These data suggest, therefore, that HU protein plays a significant role in the repair of AP sites in E. coli.

Cardiovascular Magnetic Resonance Imaging: Overview of Clinical Applications in the Context of Cardiovascular CT

CMR can evaluate myocardial contractility, volumetry, strain, flow, perfusion, viability, and vascular anatomy without ionizing radiation or iodinated contrast agents. Multiple pulse sequences are acquired in different orientations to the heart and relevant vasculature, some of which require gadolinium-based contrast agents. A strength of CMR is the ability to determine tissue characteristics including edema, hemorrhage, iron content, inflammation, and diffuse and focal fibrosis useful for the diagnosis of cardiomyopathic processes, pericardial disease, and cardiac masses. CMR assessment of cardiovascular structure, function, hemodynamics, extracardiac vasculature and thoracic structure, makes it a useful adjunct to echocardiography in patients with congenital heart disease and valvular disease. Although imaging of the coronary arteries is feasible with CMR, CCTA is the gold standard for noninvasive coronary angiography providing detailed visualization of the entire coronary artery tree. Decisions related to performing CMR versus CCTA require information patient profile and strengths and limitations of each modality in relation to the posed clinical question.

Abstract B87: Determing molecular and cellular mechanisms of gemcitabine response and relapse using human PDAC xenograft models.

Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer deaths in the U.S., with an overall median survival of less than one year. Of patients who are candidates for curative-intent surgical resection, median survival increases to almost two years. Moreover, addition of adjuvant gemcitabine treatment nearly doubles disease-free survival, with a 5-year survival of 23%. Despite this survival benefit, a majority of patients are refractory to treatment, relapse quickly, and succumb to disease. In the field, it has been hypothesized that two mechanisms may determine PDAC tumor cell chemoresistance to gemcitabine. First, expression of the dCK metabolic axis with its associated transporters and enzymes can greatly influence gemcitabine toxicity and pharmacodynamics. Second, certain tumor populations with cancer stem cell-like activity may be chemoresistant due to altered genetic or epigenetic mechanisms. We decided to test these hypotheses and determine the mechanisms governing gemcitibine resistance using patient-derived xenograft models we have generated. We have treated seven human xenograft lines in vivo with gemcitabine and found that two tumor lines completely responded throughout the treatment course, whereas five tumor lines relapsed immediately after drug withdrawal. We first checked dCK levels and activity, as well as other genes previously reported to be associated with gemcitabine import and metabolism, but found no correlation. We next examined cancer stem cell content with established cell surface markers, and found no significant differences among xenograft lines before or after treatment. Furthermore, we utilized a pulse-chase lineage tracing approach to track repopulating cells in vivo in relapsing tumors after treatment, and found that many tumor cells are capable of proliferation, not being confined to a specific tumor subpopulation. Together, our preliminary study suggests neither the dCK axis nor subpopulations within the PDAC mass are responsible for gemcitabine relapse. We are currently performing gene expression microarray analysis on these tumors aiming at identifying pathway alterations segregating response and relapse groups, which may provide novel molecular insights governing gemcitabine sensitivity. Citation Format: Antreas A. Hindoyan, Linh Tran, Sindhuja Godavarthi, Daniel Braas, Heather Christofk, Timothy Donahue, Hong Wu. Determing molecular and cellular mechanisms of gemcitabine response and relapse using human PDAC xenograft models. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Progress and Challenges; Jun 18-21, 2012; Lake Tahoe, NV. Philadelphia (PA): AACR; Cancer Res 2012;72(12 Suppl):Abstract nr B87.

Impact of Relaxation Training on Patient-Perceived Measures of Anxiety, Pain, and Outcomes after Interventional Electrophysiology Procedures

Background: Electrophysiology procedures vary in invasiveness, duration, and anesthesia utilized. While complications are low and efficacy high, cases are elective and patient experiences related to anxiety, pain, and perceived outcomes are not well studied. We sought to determine if a 30‐minute audio compact disc (CD) that teaches relaxation techniques and wellness perception prior to an elective procedure impacts validated measures of anxiety, pain, and procedural outcomes.

Cardiovascular Magnetic Resonance Imaging: Overview of Clinical Applications

Technologic advances in cardiovascular magnetic resonance imaging (CMR) and cardiovascular computed tomography angiography (CCTA) allow these modalities to comprehensively visualize cardiovascular structures and function. The decision to perform CMR vs. CCTA requires knowledge of the individual strengths and limitations of these imaging techniques, the specific details of a patient’s medical history, and the clinical questions which need to be answered. In many clinical scenarios, echocardiography is performed as an initial study to assess cardiovascular substrates, with CMR or CCTA performed when further cardiovascular characterization is necessary and a noninvasive approach is preferable. Given the rapid evolution of these technologies, appropriateness criteria have been developed for specific cardiovascular indications [1].

Orientation and Approach to Cardiovascular Images

Mastery of conventional anatomy and physiology is essential as a template for understanding of cardiovascular variation and pathology. A multitude of analysis and reconstruction tools including orthogonal, double oblique, and curved multiplanar reformatted 2-D, and volume-rendered 3-D views can be utilized for comprehensive diagnosis of cardiovascular pathology. The goal of this chapter is orientation to review and analysis of cardiovascular structure from CCTA image sets using a systematic approach.