Tuba Esfandyari

Wireless pH‐motility capsule for colonic transit: prospective comparison with radiopaque markers in chronic constipation

Background  Colon transit (CT) measurements are used in the management of significant constipation. The radiopaque marker (ROM) method provides limited information.

A phase 2a, randomized, double‐blind 28‐day study of TZP‐102 a ghrelin receptor agonist for diabetic gastroparesis

Background  Gastroparesis causes significant morbidity and treatment options are limited. TZP‐102 a novel, macrocyclic, selective, oral ghrelin receptor agonist, was evaluated in a randomized, double‐blind, placebo‐controlled trial in patients with diabetic gastroparesis.

Phase 2b, randomized, double-blind 12-week studies of TZP-102, a ghrelin receptor agonist for diabetic gastroparesis.

TZP‐102, a potent, oral, ghrelin receptor agonist, improved diabetic gastroparesis symptoms in Phase 2a.

Ral overactivation in malignant peripheral nerve sheath tumors.

ABSTRACT Ras leads an important signaling pathway that is deregulated in neurofibromatosis type 1 and malignant peripheral nerve sheath tumor (MPNST). In this study, we show that overactivation of Ras and many of its downstream effectors occurred in only a fraction of MPNST cell lines. RalA, however, was overactivated in all MPNST cells and tumor samples compared to nontransformed Schwann cells. Silencing Ral or inhibiting it with a dominant-negative Ral (Ral S28N) caused a significant reduction in proliferation, invasiveness, and in vivo tumorigenicity of MPNST cells. Silencing Ral also reduced the expression of epithelial mesenchymal transition markers. Expression of the NF1-GTPase-related domain (NF1-GRD) diminished the levels of Ral activation, implicating a role for neurofibromin in regulating RalA activation. NF1-GRD treatment caused a significant decrease in proliferation, invasiveness, and cell cycle progression, but cell death increased. We propose Ral overactivation as a novel cell signaling abnormality in MPNST that leads to important biological outcomes with translational ramifications.

Utilizing Ras Signaling Pathway to Direct Selective Replication of Herpes Simplex Virus-1

Re-engineering the tropism of viruses is an attractive translational strategy for targeting cancer cells. The Ras signal transduction pathway is a central hub for a variety of pro-oncogenic events with a fundamental role in normal and neoplastic physiology. In this work we were interested in linking Ras activation to HSV-1 replication in a direct manner in order to generate a novel oncolytic herpes virus which can target cancer cells. To establish such link, we developed a mutant HSV-1 in which the expression of ICP4 (infected cell protein-4, a viral protein necessary for replication) is controlled by activation of ELK, a transcription factor down-stream of the Ras pathway and mainly activated by ERK (extracellular signal-regulated kinase, an important Ras effector pathway). This mutant HSV-1 was named as Signal-Smart 1 (SS1). A series of prostate cells were infected with the SS1 virus. Cells with elevated levels of ELK activation were preferentially infected by the SS1 virus, as demonstrated by increased levels of viral progeny, herpetic glycoprotein C and overall SS1 viral protein production. Upon exposure to SS1, the proliferation, invasiveness and colony formation capabilities of prostate cancer cells with increased ELK activation were significantly decreased (p<0.05), while the rate of apoptosis/necrosis in these cells was increased. Additionally, high Ras signaling cells infected with SS1 showed a prominent arrest in the G1 phase of the cell cycle as compared to cells exposed to parental HSV-1. The results of this study reveal the potential for re-modeling the host-herpes interaction to specifically interfere with the life of cancer cells with increased Ras signaling. SS1 also serves as a “prototype” for development of a family of signal-smart viruses which can target cancer cells on the basis of their signaling portfolio.

Inhibition of Mesothelin as a Novel Strategy for Targeting Cancer Cells

Mesothelin, a differentiation antigen present in a series of malignancies such as mesothelioma, ovarian, lung and pancreatic cancer, has been studied as a marker for diagnosis and a target for immunotherapy. We, however, were interested in evaluating the effects of direct targeting of Mesothelin on the viability of cancer cells as the first step towards developing a novel therapeutic strategy. We report here that gene specific silencing for Mesothelin by distinct methods (siRNA and microRNA) decreased viability of cancer cells from different origins such as mesothelioma (H2373), ovarian cancer (Skov3 and Ovcar-5) and pancreatic cancer (Miapaca2 and Panc-1). Additionally, the invasiveness of cancer cells was also significantly decreased upon such treatment. We then investigated pro-oncogenic signaling characteristics of cells upon mesothelin-silencing which revealed a significant decrease in phospho-ERK1 and PI3K/AKT activity. The molecular mechanism of reduced invasiveness was connected to the reduced expression of β-Catenin, an important marker of EMT (epithelial-mesenchymal transition). Ero1, a protein involved in clearing unfolded proteins and a member of the ER-Stress (endoplasmic reticulum-stress) pathway was also markedly reduced. Furthermore, Mesothelin silencing caused a significant increase in fraction of cancer cells in S-phase. In next step, treatment of ovarian cancer cells (OVca429) with a lentivirus expressing anti-mesothelin microRNA resulted in significant loss of viability, invasiveness, and morphological alterations. Therefore, we propose the inhibition of Mesothelin as a potential novel strategy for targeting human malignancies.

Inhibition of RalA signaling pathway in treatment of non-small cell lung cancer

Non-small cell lung cancer (NSCLC) is the most common type of lung cancer and relatively resistant to chemotherapy. The most prevalent molecular abnormality in NSCLC is the overactivation of K-Ras proto-oncogene; therefore, elucidating down-stream Ras signaling in NSCLC is significantly important in developing novel therapies against this malignancy. Our work indicates that RalA, an important effector of Ras, is activated in NSCLC cell lines. While RalA was also overactivated in fetal human broncho-epithelial cells, RalBP1 (Ral binding protein-1), an important down-stream effector of RalA, was expressed at higher levels in cancer cell lines. Aurora kinase-A (AKA), an upstream activator of RalA, was also found to be active only in malignant cells. The outcome of inhibition of RalA (by gene specific silencing using a lentivirus) on the malignant phenotype of A549 cells was also studied. While proliferation and invasiveness of A549 cells were reduced upon silencing RalA, apoptosis and necrosis were elevated in such conditions. Additionally, the in vivo tumorigenesis of A549 cells was reduced upon partial inhibition of RalA and AKA using pharmacological inhibitors. Finally, we were interested in evaluating the level of active RalA in the fraction of NSCLC cells expressing cancer stem cell markers. For this purpose cells with increased expression of CD44 were separated from A549 cells and compared with cells with low level of expression of this marker and an unsorted population. A significant enhancement of RalA activation in high CD44+ cells was found as potential evidence for involvement of RalA signaling in initiation of the neoplastic procedure and an important contributor for tumor maintenance in NSCLC. Further studies can reveal therapeutic, preventive and diagnostic value of RalA pathway in this deadly disease.

RalA signaling pathway as a therapeutic target in hepatocellular carcinoma (HCC).

Ral (Ras like) leads an important proto‐oncogenic signaling pathway down‐stream of Ras. In this work, RalA was found to be significantly overactivated in hepatocellular carcinoma (HCC) cells and tissues as compared to non‐malignant samples. Other elements of RalA pathway such as RalBP1 and RalGDS were also expressed at higher levels in malignant samples. Inhibition of RalA by gene‐specific silencing caused a robust decrease in the viability and invasiveness of HCC cells. Additionally, the use of geranyl–geranyl transferase inhibitor (GGTI, an inhibitor of Ral activation) and Aurora kinase inhibitor II resulted in a significant decrease in the proliferation of HCC cells. Furthermore, RalA activation was found to be at a higher level of activation in HCC stem cells that express CD133. Transgenic mouse model for HCC (FXR‐Knockout) also revealed an elevated level of RalA‐GTP in the liver tumors as compared to background animals. Finally, subcutaneous mouse model for HCC confirmed effectiveness of inhibition of aurora kinase/RalA pathway in reducing the tumorigenesis of HCC cells in vivo. In conclusion, RalA overactivation is an important determinant of malignant phenotype in differentiated and stem cells of HCC and can be considered as a target for therapeutic intervention.

Transcription factors down-stream of Ras as molecular indicators for targeting malignancies with oncolytic herpes virus

Overactivation in Ras signaling has been under intensive study as the molecular basis for development of cancer. Such overactivation can occur in the presence or absence of mutations in Ras gene resulting in activation of a series of down‐stream effectors such as transcription factors. Different studies have shown the activation of Ras down‐stream effectors in non‐Hodgkin lymphoma (NHL) although mutations in Ras are not prevalent in this malignancy. Since overactivation in Ras signaling also increases permissiveness of cancer cells to infection by oncolytic versions of herpes simplex virus (e.g. R3616), we were interested in evaluating the value of transcription factors down‐stream of Ras as molecular indicators for permissiveness to herpes therapy. In order to accomplish this, and also to assess the permissiveness of lymphoma cells to infection with R3616, we used NHL cell lines Daudi, Jurkat, NC37, Raji, Ramos and ST486. Once the levels of phosphorylation (activation) of extracellular‐signal regulated kinase (ERK, a Ras effector pathway) and its down‐stream transcription factor ELK were evaluated, Raji and NC37 showed a significant increase in the phosphorylation levels of both molecules while ATF2 (another transcription factor down‐stream of p38‐kinase pathway) seemed to be activated in all studied cells. Raji and NC37 cells were also most permissive cells to infection with R3616 while their permissiveness was decreased upon treatment of cells with an inhibitor of ELK‐DNA binding portraying ERK/ELK as a suitable predictive indicator for selection of cancer cells with increased sensitivity to R3616. This study, therefore, for the first time documents permissiveness of lymphoma cells to oncolytic herpes viruses and introduces ELK as a suitable factor for predicting tumor susceptibility to these novel anticancer agents.

The utility of lower endoscopic ultrasound-guided fine needle aspiration for the diagnosis of benign and malignant pelvic diseases.

Background: The utility of endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) for the diagnosis of pelvic masses has been suggested but limited data are available in the literature regarding its diagnostic accuracy. Goals: To report our institutional experience with EUS-FNA for the diagnosis of a variety of pelvic diseases. Methods: Patients who were referred for the evaluation of pelvic lesions using lower EUS-FNA were included in this retrospective analysis if they had available surgical pathology (obtained after EUS) which was considered the gold standard against which the EUS-FNA findings would have been compared. The diagnostic accuracy of EUS-FNA for pelvic masses was analyzed and any early or late complications after the procedure were reported. A pelvic mass was defined in the study as any mass seen with an imaging modality in the pelvic area including those involving the colonic wall. Results: Twenty patients had EUS-FNA followed by surgery for whom FNA cytology and surgical pathology findings were available. EUS-FNA reached the correct diagnosis in 19 out of 20 patients, whereas for the missing 1 malignant lymph node wherein FNA revealed benign cytology, surgical specimen confirmed metastatic colon cancer. The sensitivity and specificity of EUS-FNA were 90% and 100%, respectively, with positive and negative predictive values of 100% and 90%, respectively. No early or late complications were encountered with this procedure for the sampling of cystic and noncystic masses. Conclusions: EUS-FNA has excellent diagnostic accuracy for pelvic masses. It represents a safe procedure with excellent yield and thus may be used as a first line modality for the evaluation and diagnosis of pelvic masses within its reach.