Edward E. Whang

Necrotizing pancreatitis: Contemporary analysis of 99 consecutive cases

ObjectiveTo analyze the impact of a conservative strategy of management in patients with necrotizing pancreatitis, reserving intervention for patients with documented infection or the late complications of organized necrosis. Summary Background DataThe role of surgery in patients with sterile pancreatic necrosis remains controversial. Although a conservative approach is being increasingly used, few studies have evaluated this strategy when applied to the entire spectrum of patients with necrotizing pancreatitis. MethodsThe authors reviewed 1,110 consecutive patients with acute pancreatitis managed at Brigham and Women’s Hospital between January 1, 1995, and January 1, 2000, focusing on those with pancreatic necrosis documented by contrast-enhanced computed tomography. Fine-needle aspiration, the presence of extraintestinal gas on computed tomography, or both were used to identify infection. ResultsThere were 99 (9%) patients with necrotizing pancreatitis treated, with an overall death rate of 14%. In three patients with underlying medical problems, the decision was made initially not to intervene. Of the other 62 patients without documented infection, all but 3 were managed conservatively; this group’s death rate was 11%. Of these seven deaths, all were related to multiorgan failure. Five patients in this group eventually required surgery for organized necrosis, with no deaths. Of the 34 patients with infected necrosis, 31 underwent surgery and 3 underwent percutaneous drainage. Only four (12%) of these patients died, all of multiorgan failure. Of the total 11 patients who died, few if any would have been candidates for earlier surgical intervention. ConclusionsThese results suggest that conservative strategies can be applied successfully to manage most patients with necrotizing pancreatitis, although some will eventually require surgery for symptomatic organized necrosis. Few if any patients seem likely to benefit from a more aggressive strategy.

RNA interference targeting the M2 subunit of ribonucleotide reductase enhances pancreatic adenocarcinoma chemosensitivity to gemcitabine.

Ribonucleotide reductase is emerging as an important determinant of gemcitabine chemoresistance in human cancers. Activity of this enzyme, which catalyses conversion of ribonucleotide 5′-diphosphates to their 2′-deoxynucleotides, is modulated by levels of its M2 subunit (RRM2). Here we show that RRM2 overexpression is associated with gemcitabine chemoresistance in pancreatic adenocarcinoma cells, and that suppression of RRM2 expression using RNA interference mediated by small interfering RNA (siRNA) enhances gemcitabine-induced cytotoxicity in vitro. We demonstrate the ability of systemically administered RRM2 siRNA to suppress tumoral RRM2 expression in an orthotopic xenograft model of pancreatic adenocarcinoma. Synergism between RRM2 siRNA and gemcitabine results in markedly suppressed tumor growth, increased tumor apoptosis and inhibition of metastasis. Our findings confirm the importance of RRM2 in pancreatic adenocarcinoma gemcitabine chemoresistance. This is the first demonstration that systemic delivery of siRNA-based therapy can enhance the efficacy of an anticancer nucleoside analog.

EphA2: a determinant of malignant cellular behavior and a potential therapeutic target in pancreatic adenocarcinoma

The EphA2 receptor tyrosine kinase is overexpressed in a variety of human cancers. We sought to characterize the role of EphA2 in pancreatic adenocarcinoma and, using RNA interference (RNAi) mediated by small interfering RNA (siRNA), we determined the effects of suppressing EphA2 expression in vitro and in vivo. EphA2 expression in PANC1, MIAPaCa2, BxPC3 and Capan2 cells was assessed by Northern and Western blot. We artificially overexpressed EphA2 by transient transfection and suppressed EphA2 expression using RNAi. Cellular invasiveness was quantified by modified Boyden chamber assay. Anoikis was induced by anchorage-independent polyHEMA culture and caspase 3 activity was quantified fluorometrically. Focal adhesion kinase (FAK) phosphorylation was assessed by immunoprecipitation. EphA2 siRNA treatment was assessed in a nude mouse xenograft model. Pancreatic adenocarcinoma cells differentially express EphA2. Inherent and induced EphA2 overexpression is associated with increased cellular invasiveness and anoikis resistance. EphA2 siRNA suppresses EphA2 expression, cellular invasiveness, anoikis resistance and FAK phosphorylation in vitro and retards tumor growth and inhibits metastasis in vivo. EphA2 is both a determinant of malignant cellular behavior and a potential therapeutic target in pancreatic adenocarcinoma.

Is severity of necrotizing pancreatitis increased in extended necrosis and infected necrosis

Introduction We previously reported that organ failure occurs in 50% of patients with necrotizing pancreatitis, that extended pancreatic necrosis (greater than 50% necrosis) is not associated with an increased prevalence of organ failure or infected necrosis, and that the prevalence of organ failure is similar in sterile necrosis and infected necrosis. Aims To analyze these relations in a larger group of patients and to evaluate other factors that might have prognostic significance. Methodology We reviewed 1,110 consecutive cases of acute pancreatitis between January 1, 1995, and January 1, 2000. Necrosis was documented by contrast-enhanced CT. A p value less than 0.05 was considered significant. Results Ninety-nine patients (9%) had necrotizing pancreatitis; 52% had organ failure. Patients with extended pancreatic necrosis did not have increased prevalence of organ failure or infected necrosis but did have an increased need for intubation and an increased mortality rate associated with multiple organ failure. Patients with infected necrosis did not have an increased prevalence of organ failure but did have a marginally increased prevalence of multiple organ failure and increased need for intubation. Overall mortality was 14% and was markedly increased among patients with organ failure at admission (47%) and among patients who had multiple organ failure during the hospitalization (49%). Conclusion Although severity of necrotizing pancreatitis was somewhat increased in extended pancreatic necrosis and infected necrosis, mortality was more strongly linked to organ failure at admission and multiple organ failure during hospitalization.

Inhibition of Src Tyrosine Kinase Impairs Inherent and Acquired Gemcitabine Resistance in Human Pancreatic Adenocarcinoma Cells

Purpose: We tested the hypotheses that Src tyrosine kinase overactivity represents a chemoresistance mechanism and that Src inhibition may enhance gemcitabine cytotoxicity in pancreatic adenocarcinoma cells. Experimental Design: Pancreatic adenocarcinoma cells PANC1, MiaPaCa2, Capan2, BxPC3, and PANC1GemRes, a stably gemcitabine-resistant subline of PANC1, were exposed to combinations of gemcitabine and Src tyrosine kinase inhibitor 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2). Src expression, phosphorylation (Tyr-416), and activity were analyzed by immunoblotting and in vitro kinase assay. Expression of the M2 subunit of ribonucleotide reductase (RRM2), a putative chemoresistance enzyme, was quantified by Northern and Western blot. Cellular proliferation was quantified by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Apoptosis was characterized by YO-PRO-1/propidium iodide staining, fluorometric caspase profiling, and caspase inhibition (Z-Val-Ala-Asp-fluoromethyl ketone). The effects of constitutively active and dominant negative Src were determined. The therapeutic efficacy of PP2 in combination with gemcitabine was tested in nude mice orthotopically xenografted with PANC1GemRes. Results: Greater gemcitabine resistance was associated with higher Src phosphorylation and activity, both of which were higher in PANC1GemRes, relative to PANC1; total Src levels were alike. PANC1GemRes overexpressed RRM2. PP2 enhanced inherent gemcitabine chemosensitivity and attenuated gemcitabine resistance in PANC1GemRes. Constitutively active Src increased gemcitabine chemoresistance; dominant negative Src impaired gemcitabine chemoresistance. PP2 augmented gemcitabine-induced caspase-mediated apoptosis, suppressed RRM2 expression, and decreased activity of the RRM2-regulating transcription factor E2F1 in PANC1GemRes. PP2 and gemcitabine in combination substantially decreased tumor growth and inhibited metastasis in vivo. Conclusions: Increased Src tyrosine kinase activity represents a potential chemoresistance mechanism and a promising therapeutic target warranting further investigation in gemcitabine-resistant pancreatic adenocarcinoma.

CEACAM6 gene silencing impairs anoikis resistance and in vivo metastatic ability of pancreatic adenocarcinoma cells

Anoikis is the apoptotic response induced in normal cells by inadequate or inappropriate adhesion to substrate. It is postulated that resistance to anoikis facilitates tumorigenesis and metastasis. Carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6) is an immunoglobulin superfamily member overexpressed in a number of human cancers and implicated in anoikis resistance. We tested the effect of CEACAM6 gene silencing on anoikis in pancreatic adenocarcinoma cell lines. Anoikis was induced in PANC1, Capan2, MiaPaCa2 and MiaAR (a MiaPaCa2-derived anoikis-resistant subline) by culture in poly-2-hydroxyethylmethacrylate-coated wells. Anoikis was quantified by YO-PRO-1/propidium iodide staining and flow cytometry. The role of caspase activation was determined using fluorometric profiling and the caspase inhibitor Z-Val-Ala-Asp-fluoromethyl ketone (Z-VAD-fmk). CEACAM6 expression was suppressed by RNA interference. Using a nude mouse orthotopic xenograft model, we assessed the effect of this treatment on in vivo metastatic ability. Anoikis resistance was associated with increased CEACAM6 expression. CEACAM6-specific short interfering ribonucleic acid (siRNA), but not control siRNA, increased susceptibility to caspase-mediated anoikis, an effect abrogated by Z-VAD-fmk, and decreased Akt phosphorylation (Ser-473) under anchorage-independent conditions. CEACAM6 gene silencing reversed the acquired anoikis resistance of MiaAR and inhibited its in vivo metastatic ability. CEACAM6 warrants further investigation as a novel therapeutic target for the treatment of pancreatic adenocarcinoma.

Implementing Resident Work Hour Limitations: Lessons from the New York State Experience

ObjectiveTo determine the impact of work hour limitations imposed by the 405 (Bell) Regulations as perceived by general surgery residents in New York State. Summary Background DataNew Accreditation Council for Graduate Medical Education (ACGME) requirements on resident duty hours are scheduled to undergo nationwide implementation in July 2003. State regulations stipulating similar resident work hour limitations have already been enacted in New York. MethodsA statewide survey of residents enrolled in general surgery residencies in New York was administered. ResultsMost respondents reported general compliance with 405 Regulations in their residency programs, a finding corroborated by reported work hours and call schedules. Whereas a majority of residents reported improved quality of life as a result of the work hour limitations, a substantial portion reported negative impacts on surgical training and quality and continuity of patient care. Negative perceptions of the impact of duty hour restrictions were more prevalent among senior residents and residents at academic medical centers than among junior residents and residents at community hospitals. ConclusionsImplementation of resident work hour limitations in general surgery residencies may have negative consequences for patient care and resident education. As surgical residency programs develop strategies for complying with ACGME requirements, these negative consequences must be addressed.

Prostaglandin E2 enhances pancreatic cancer invasiveness through an Ets-1-dependent induction of matrix metalloproteinase-2

Accumulating evidence suggests an important role for cyclooxygenase-2 (COX-2) in the pathogenesis of a wide range of malignancies. Here we tested the hypothesis that the COX-2 product prostaglandin E2 (PGE2) increases cellular invasive potential by inducing matrix metalloproteinase-2 (MMP-2) expression and activity through an extracellular signal-regulated kinase (ERK)/Ets-1-dependent mechanism in pancreatic cancer. PANC-1 and MIAPaCa-2 pancreatic cancer cells were treated with PGE2 or rofecoxib, a selective COX-2 inhibitor. MMP-2 expression and activity were assayed using Western blot analysis and zymography, respectively. MMP-2 promoter activity was analyzed with a luciferase-based assay. Ets-1 activity was analyzed using gel shift assay. Ets-1 expression was specifically silenced using RNA interference. Cellular invasive and migratory potentials were determined using a Boyden chamber assay with or without Matrigel, respectively. Exogenous PGE2 induced MMP-2 expression and activity and increased ERK1/2 phosphorylation, Ets-1 binding activity, and MMP-2 promoter activity. PGE2 also increased cellular migratory and invasive potentials. The mitogen-activated protein kinase kinase inhibitor PD98059 and Ets-1 silencing each abolished PGE2-induced increases in MMP-2 expression. PD98059 and Ets-1 silencing each abrogated the effect of PGE2 on cellular invasive potential but not on cellular migratory potential. Rofecoxib suppressed MMP-2 expression and activity, Ets-1 binding activity, MMP-2 promoter activity, and cellular migratory and invasive potentials. These results suggest that PGE2 mediates pancreatic cancer cellular invasiveness through an ERK/Ets-1-dependent induction of MMP-2 expression and activity. They also suggest that COX-2 inhibition may represent a strategy to inhibit invasive potential in pancreatic cancer.

Systemic siRNA-Mediated Gene Silencing: A New Approach to Targeted Therapy of Cancer

Objective:RNA interference (RNAi), mediated by small interfering RNA (siRNA), silences genes with a high degree of specificity and potentially represents a general approach for molecularly targeted anticancer therapy. The aim of this study was to evaluate the ability of systemically administered siRNA to silence gene expression in vivo and to assess the effect of this approach on tumor growth using a murine pancreatic adenocarcinoma xenograft model. Summary Background Data:Carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6) is widely overexpressed in human gastrointestinal cancer. Overexpression of CEACAM6 promotes cell survival under anchorage independent conditions, a characteristic associated with tumorigenesis and metastasis. Methods:CEACAM6 expression was quantified by real-time polymerase chain reaction (PCR) and Western blot. Mice (n = 10/group) were subcutaneously xenografted with 2 × 106 BxPC3 cells (which inherently overexpress CEACAM6). Tumor growth, CEACAM6 expression, cellular proliferation (Ki-67 immunohistochemistry), apoptosis, angiogenesis (CD34 immunohistochemistry), and survival were compared for mice administered either systemic CEACAM6-specific or control single-base mismatch siRNA over 6 weeks, following orthotopic tumor implantation. Results:Treatment with CEACAM6-specific siRNA suppressed primary tumor growth by 68% versus control siRNA (P < 0.05) and was associated with a decreased proliferating cell index, impaired angiogenesis and increased apoptosis in the xenografted tumors. CEACAM6-specific siRNA completely inhibited metastasis (0% of mice versus 60%, P < 0.05) and significantly improved survival, without apparent toxicity. Conclusions:Our data demonstrate the efficacy of systemically administered siRNA as a therapeutic modality in experimental pancreatic cancer. This novel therapeutic strategy may be applicable to a broad range of cancers and warrants investigation in patients with refractory disease.

Work hours reform: perceptions and desires of contemporary surgical residents

BACKGROUND New Accreditation Council for Graduate Medical Education (ACGME) requirements on resident duty hours are scheduled to undergo nationwide implementation in July 2003. General surgery residents, because of their long duty hours, are likely to be among those most affected by changes imposed to comply with the ACGME requirements. There are few contemporary data on their attitudes toward work hours reform. STUDY DESIGN The study entailed a region-wide survey of residents enrolled in general surgery residencies in New England to characterize the perceptions and desires of surgical residents on the issue of work hours reform. RESULTS Respondents reported working a mean of 105 +/- 0.7 hours per week, considerably more than the 80-hour limit stipulated by the ACGME. Of the respondents, 81% reported that sleep deprivation had negatively affected their work. A strong majority of respondents believe that work hours reform would improve their quality of life but less than one half expect it to have a positive impact on patient care. A greater percentage of senior residents than junior residents (p < 0.05) have negative perceptions of work hour limitations, particularly with respect to consequences for patient care. Other findings suggest that residents who have actually experienced work hour restrictions are less positive about such restrictions than these residents who had not yet experienced them. CONCLUSIONS Changes imposed by residency programs to comply with work hour requirements might have detrimental effects on senior residents and patient care. The impact of such changes should be carefully monitored as the ACGME requirements are implemented.