Faculty Bibliography

Adaptive resistance to MEK inhibitors (MEK-Is) typically occurs via induction of genes for different receptor tyrosine kinases (RTKs) and/or their ligands, even in tumors of the same histotype, making combination strategies challenging. SHP2 (PTPN11) is required for RAS/ERK pathway activation by most RTKs, and might provide a common resistance node. We found that combining the SHP2 inhibitor SHP099 with a MEK-I inhibited the proliferation of multiple cancer cell lines in vitro. PTPN11 knockdown/MEK-I treatment had similar effects, while expressing SHP099 binding-defective PTPN11 mutants conferred resistance, demonstrating that SHP099 is on-target. SHP099/trametinib was highly efficacious in xenograft and/or genetically engineered models of KRAS-mutant pancreas, lung, and ovarian cancer and in wild type RAS-expressing triple negative breast cancer. SHP099 inhibited activation of KRAS mutants with residual GTPase activity, impeded SOS/RAS/MEK/ERK1/2 reactivation in response to MEK-Is and blocked ERK1/2-dependent transcriptional programs. We conclude that SHP099/MEK-I combinations could have therapeutic utility in multiple malignancies.

Cancer heterogeneity is a notorious hallmark of this disease, and it is desirable to tailor effective treatments for each individual patient. Drug combinations have been widely accepted in cancer treatment for better therapeutic efficacy as compared to a single compound. However, experimental complexity and cost grow exponentially with more target compounds under investigation. The primary challenge remains to efficiently perform a large-scale drug combination screening using a small number of patient primary samples for testing. Here, a scalable, easy-to-use, high-throughput drug combination screening scheme is reported, which has the potential of screening all possible pairwise drug combinations for arbitrary number of drugs with multiple logarithmic mixing ratios. A "Christmas tree mixer" structure is introduced to generate a logarithmic concentration mixing ratio between drug pairs, providing a large drug concentration range for screening. A three-layer structure design and special inlets arrangement facilitate simple drug loading process. As a proof of concept, an 8-drug combination chip is implemented, which is capable of screening 172 different treatment conditions over 1032 3D cancer spheroids on a single chip. Using both cancer cell lines and patient-derived cancer cells, effective drug combination screening is demonstrated for precision medicine.

Bladder cancer is a significant health problem. It is estimated that more than 16,000 people will die this year in the United States from bladder cancer. While 75% of bladder cancers are non-invasive and unlikely to metastasize, about 25% progress to an invasive growth pattern. Up to half of the patients with invasive cancers will develop lethal metastatic relapse. Thus, understanding the mechanism of invasive progression in bladder cancer is crucial to predict patient outcomes and prevent lethal metastases. In this article, we present a three-dimensional cancer invasion model which allows incorporation of tumor cells and stromal components to mimic in vivo conditions occurring in the bladder tumor microenvironment. This model provides the opportunity to observe the invasive process in real time using time-lapse imaging, interrogate the molecular pathways involved using confocal immunofluorescent imaging and screen compounds with the potential to block invasion. While this protocol focuses on bladder cancer, it is likely that similar methods could be used to examine invasion and motility in other tumor types as well.

The biological properties of pancreatic cancer stem cells (PCSCs) remain incompletely defined and the central regulators are unknown. By bioinformatic analysis of a human PCSC-enriched gene signature, we identified the transcription factor HNF1A as a putative central regulator of PCSC function. Levels of HNF1A and its target genes were found to be elevated in PCSCs and tumorspheres, and depletion of HNF1A resulted in growth inhibition, apoptosis, impaired tumorsphere formation, decreased PCSC marker expression, and downregulation of POU5F1/OCT4 expression. Conversely, HNF1A overexpression increased PCSC marker expression and tumorsphere formation in pancreatic cancer cells and drove pancreatic ductal adenocarcinoma (PDA) cell growth. Importantly, depletion of HNF1A in xenografts impaired tumor growth and depleted PCSC marker-positive cells in vivo. Finally, we established an HNF1A-dependent gene signature in PDA cells that significantly correlated with reduced survivability in patients. These findings identify HNF1A as a central transcriptional regulator of PCSC properties and novel oncogene in PDA.

We investigated whether intrapancreatic coagulation, with deposition of the fibrinogen-γ dimer (Fib-γD) and hypoxia, affect the severity of acute pancreatitis (AP) in mice. Pancreata of mice with AP induced by administration of cerulein or by L-arginine, or from patients with pancreatitis, had increased deposition of Fib-γD compared with control pancreata. Heparin administration protected mice from cerulein-induced AP and prevented Fib-γD formation. Cerulein administration resulted in activation and stabilization of hypoxia-inducible factor-1α (HIF1α) in pancreata of oxygen-dependent degradation domain-luciferase HIF1α reporter mice. Cerulein also led to induction of genes regulated by HIF1α, including Vegfa and Ero1a, before evidence of Fib-γD deposition or histologic features of AP. Expression of tissue factor, which is regulated by vascular endothelial growth factor, also increased following cerulein administration. Mice with acinar cell-specific disruption of Hif1a (Hif1a^Ac-/-) developed spontaneous endoplasmic reticulum stress and less severe AP, but did not accumulate Fib-γD following administration of cerulein. Feeding mice increased pancreatic expression of HIF1α, indicating a physiologic role in the exocrine pancreas. Therefore, HIF1α has bifunctional roles, in exocrine pancreas homeostasis and progression of AP that is promoted by intrapancreatic coagulation.

Hepatocyte nuclear factor 1A (HNF1A) is a transcription factor essential to normal pancreas and liver development and homeostasis. HNF1A has been shown to be upregulated in pancreatic cancer stem cells (CSC), a proposed key driver of pancreatic ductal adenocarcinoma (PDAC), relative to other pancreatic cancer cells. A CSC gene signature consisting of 50 upregulated genes, including HNF1A, was previously identified by a microarray. This signature predicts worse survival compared to HNF1A expression alone. This research aims to establish an HNF1A-dependent gene signature that could potentially be used as a biomarker for pancreatic cancer. Using quantitative reverse transcriptase PCR, multiple CSC signature genes were shown to respond to HNF1A knockdown and overexpression. Responsive genes varied across cell lines, suggesting a complex pathway significantly affected by HNF1A transcriptional control

In recent years, as the high burden of surgical disease and poor access to surgical care in low- and middle-income countries have gained recognition as major public health problems, interest in global health has surged among surgical trainees and faculty. Traditionally, clinical volunteerism was at the forefront of the high-income country response to the significant burden of surgical disease in low- and middle-income countries. However, sustainable strategies for providing surgical care in low- and middle-income countries increasingly depend on bilateral clinical, research, and education collaborations to ensure effective resource allocation and contextual relevance. Academic global surgery creates avenues for interested surgeons to combine scholarship and education with their clinical global surgery passions through incorporation of basic/translational, education, clinical outcomes, or health services research with global surgery. Training in global health, either within residency or through advanced degrees, can provide the necessary skills to develop and sustain such initiatives. We further propose that creating cross-continental, bidirectional collaborations can maximize funding opportunities. Academic institutions are uniquely positioned to lead longitudinal and, importantly, sustainable global surgery efforts. However, for the individual global surgeon, the career path forward may be unclear. This paper reviews the development of academic global surgery, delineates the framework and factors critical to training global surgeons, and proposes models for establishing an academic career in this field. Overall, with determination, the academic global surgeon will not only carve out a niche of expertise but will define this critical field for future generations.

Usp9x has emerged as a potential therapeutic target in some hematologic malignancies and a broad range of solid tumors including brain, breast, and prostate. To examine Usp9x tumorigenicity and consequence of Usp9x inhibition in human pancreatic tumor models, we carried out gain- and loss-of-function studies using established human pancreatic tumor cell lines (PANC1 and MIAPACA2) and four spontaneously immortalized human pancreatic patient-derived tumor (PDX) cell lines. The effect of Usp9x activity inhibition by small molecule deubiquitinase inhibitor G9 was assessed in 2D and 3D culture, and its efficacy was tested in human tumor xenografts. Overexpression of Usp9x increased 3D growth and invasion in PANC1 cells and up-regulated the expression of known Usp9x substrates Mcl-1 and ITCH. Usp9x inhibition by shRNA-knockdown or by G9 treatment reduced 3D colony formation in PANC1 and PDX cell lines, induced rapid apoptosis in MIAPACA2 cells, and associated with reduced Mcl-1 and ITCH protein levels. Although G9 treatment reduced human MIAPACA2 tumor burden in vivo, in mouse pancreatic cancer cell lines established from constitutive (8041) and doxycycline-inducible (4668) KrasG12D/Tp53R172H mouse pancreatic tumors, Usp9x inhibition increased and sustained the 3D colony growth and showed no significant effect on tumor growth in 8041-xenografts. Thus, Usp9x inhibition may be therapeutically active in human PDAC, but this activity was not predicted from studies of genetically engineered mouse pancreatic tumor models.

OBJECTIVES: Definitive chemoradiotherapy for unresectable pancreatic cancer has traditionally involved 5-fluorouracil-based chemotherapy. Our institution has a long history of combining gemcitabine and radiotherapy (RT), and performed a retrospective review of all patients treated in this manner. MATERIALS AND METHODS: We reviewed the records of 180 patients treated from 1999 to 2012. Mean RT dose was 40.9 Gy in 2.2-Gy fractions, and targeted only radiographically apparent disease. Ninety-six percent of patients received full-dose gemcitabine-based chemotherapy with RT. Kaplan-Meier was used to analyze time-to-event endpoints, and Cox regression models were used to assess significant prognostic variables. RESULTS: Eighty-nine percent of patients completed RT without a toxicity-related treatment break. Median follow-up was 10.2 months. Twenty-nine percent of patients had a radiographic decrease in primary tumor size following treatment. Median overall survival was 11.8 months, time to distant metastasis (TDM) was 6.7 months, and time to local recurrence (TLR) was 8.3 months. On multivariate analysis, male sex, lower performance status, and higher posttreatment CA 19-9 level predicted for worse overall survival. Posttreatment, CA 19-9 was also associated with TDM and TLR, and radiographic tumor response was associated with better TLR. CONCLUSION: Definitive chemoradiation using full-dose gemcitabine is well tolerated and achieves survival outcomes comparable to reported trials in the literature.

We present "Labyrinth," a label-free microfluidic device to isolate circulating tumor cells (CTCs) using the combination of long loops and sharp corners to focus both CTCs and white blood cells (WBCs) at a high throughput of 2.5 mL/min. The high yield (>90%) and purity (600 WBCs/mL) of Labyrinth enabled us to profile gene expression in CTCs. As proof of principle, we used previously established cancer stem cell gene signatures to profile single cells isolated from the blood of breast cancer patients. We observed heterogeneous subpopulations of CTCs expressing genes for stem cells, epithelial cells, mesenchymal cells, and cells transitioning between epithelial and mesenchymal. Labyrinth offers a cell-surface marker-independent single-cell isolation platform to study heterogeneous CTC subpopulations.