Lee Gorden

Matrix metalloproteinases in colorectal cancer: Is it worth talking about?

Matrix metalloproteinases (MMPs), a family of extracellular matrix degrading enzymes, are expressed in various stages of colorectal cancer (CRC) and correlate with survival and prognosis. There is considerable evidence in preclinical models that MMP inhibitors (MMPIs) are effective at multiple stages of CRC tumor progression, including reducing the number of intestinal adenomas, inhibiting the growth and establishment of primary CRC tumors, and reducing metastasis to the lung and liver. However, clinical trials with MMPIs in other tumor types have been largely unsuccessful, raising the question as to whether MMPs represent therapeutic targets in CRC. This review focuses on the expression, role, and contribution of MMP family members to various stages of CRC tumor progression. The conclusion is that there is considerable evidence to suggest that MMP inhibition may be an effective strategy if applied at either end of the tumor progression spectrum; the prevention of adenomas, or the treatment of micrometastatic disease.

Marked Plasmacytosis and Immunoglobulin Abnormalities Following Infusion of Streptokinase

Marked plasmacytosis is an uncommon clinical finding associated with plasma cell dyscrasias and certain reactive states, particularly serum sickness. Moreover, serum sickness-like reactions are a well-recognized complication of therapy with streptokinase. In this report, the authors describe a patient who developed a transient, but striking, plasmacytosis and an unexplained fever following streptokinase treatment for a pulmonary embolus. An evaluation for multiple myeloma was completely negative except for the occurrence of serum monoclonal-like proteins which largely disappeared over an eight month period.

Adiponectin receptor agonists inhibit leptin induced pSTAT3 and in vivo pancreatic tumor growth

Obesity is a significant risk factor for pancreatic cancer, harboring a chronic inflammatory condition characterized by dysregulation of the adipokines, leptin and adiponectin, that in turn alter oncogenic signaling pathways. We and others have shown that leptin promotes the proliferation and an invasive potential of pancreatic cancer cells through STAT3 mediated signaling. However, the role of adiponectin on the tumorigenicity of pancreatic cancer has not been elucidated. Adiponectin represents an important negative regulator of cytokines, which acts through two receptors, ADIPOR1 and ADIPOR2, to elicit pro-apoptotic, anti-inflammatory, and anti-angiogenic responses. We show that the level and expression of both adiponectin receptors are decreased in pancreatic tumors relative to normal pancreatic tissue. In vitro stimulation with adiponectin or a small molecule adiponectin receptor agonist, AdipoRon, increases apoptosis while inhibiting pancreatic cancer cell proliferation, colony formation, and anchorage independent growth. In addition, adiponectin receptor agonism inhibits leptin mediated STAT3 activation. In vivo, treatment of mice with AdipoRon inhibits orthotopic pancreatic tumor growth. These results demonstrate that adiponectin receptor activation is a key regulator of pancreatic cancer growth and AdipoRon provides a rational agent for the development of novel therapeutic strategies for pancreatic cancer.

Presurgical Planning for Hepatobiliary Malignancies: Clinical and Imaging Considerations

There are many considerations in the evaluation of liver malignancy before planned surgical treatment. This article focuses on interpretation of MR imaging of the liver for surgical treatment planning of hepatocellular carcinoma, colorectal cancer metastases, and hilar cholangiocarcinoma. Clinical status, anatomic variants, future liver remnant, and underlying liver disease are all important factors in the decision to proceed with liver resection. The primary objective of preoperative imaging is to correctly identify patients who are candidates for curative intervention and to accurately stage their disease. Treatment planning for these complex patients is best done with a multidisciplinary team approach.

Abstract 2802: High fat diet increases development of hepatocellular carcinoma in glycine N-methyltransferase deficient mice

Introduction: Hepatocellular carcinoma (HCC) is the third leading cause of cancer deaths worldwide. HCC typically arises in patients with chronic liver disease or cirrhosis, yet it is increasingly associated with non-alcoholic fatty liver disease (NAFLD), specifically nonalcoholic steatohepatitis (NASH) in the absence of cirrhosis. NAFLD is associated with obesity, metabolic syndrome, and/or patients with type II diabetes. Our previous studies have shown that high fat diet induced hepatic steatosis increases proliferation of hepatocytes and the growth of malignant tumors in a murine model. Glycine N-MethylTransferase (GNMT) expression is lost in over 95% of HCC, and mice deficient in GNMT develop spontaneous HCC by 6 months of age. We hypothesized that GNMT deficient mice would have an increased susceptibility for the development and growth of HCC when a fed high fat diet. Methods: Wildtype and GNMT deficient mice were placed on lean diet (LD, 13% calories from fat) or high fat diet (HFD, 42% calories from fat) at eight weeks of age. An initial cohort of mice were sacrificed after 3 months on diet (6 months of age) to assess for early tumor burden. A second cohort of mice was analyzed by magnetic resonance imaging (MRI) after 6 months on diet (9 months of age) and then sacrificed to assess for late stage disease. All mice were assessed for body weight, liver weight, pancreatic weight, and proliferative index (Ki67). Results: GNMT deficient mice failed to gain weight when placed on HFD, which remained at levels equivalent to wildtype LD mice. At three months of age, wildtype mice on HFD had significantly enlarged livers due to hepatic steatosis. HFD fed GNMT deficient mouse livers were nearly 50% the size of wildtype livers and contained only minimal fatty deposits. Further, livers from HFD and LD fed GNMT mice were equivalent after 3 months, yet they were larger than wildtype mice fed LD. After six months on diet, MRI analysis showed significantly larger livers in HFD fed GNMT mice compared to LD fed GNMT mice due to extensive tumor burden. All wildtype mice lacked any tumors after six months regardless of diet. Histological analysis revealed a heightened cellular proliferation via Ki67 staining in GNMT deficient livers compared to wildtype livers. In comparison, GNMT silencing also occurs in pancreatic cancer, yet none of the GNMT deficient mice developed pancreatic tumors. However, small focal areas of pancreatitis were detected regardless of diet. Additionally, pancreatic weight was significantly decreased in HFD fed GNMT deficient mice compared the LD GNMT deficient mice. Conclusions: While high fat diet did not induce obesity in GNMT deficient mice, it significantly increased cellular proliferation and primary tumor growth in the liver. Understanding dietary factors that impact the microenvironment of the liver and contribute to HCC development and progression is vital to finding new therapeutics for this malignancy. Citation Format: Michael N. VanSaun, Alisha Mendonsa, Fanuel Messaggio, Nagaraj Nagathihalli, Lee Gorden. High fat diet increases development of hepatocellular carcinoma in glycine N-methyltransferase deficient mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2802. doi:10.1158/1538-7445.AM2017-2802

Abstract A46: AdipoRon suppresses ERK and STAT3 to inhibit pancreatic cancer growth

Introduction: The increasing incidence of pancreatic cancer is associated with a rising prevalence of obesity, a documented risk factor for the disease. Obesity harbors a systemic chronic inflammatory disorder characterized by increased production and secretion of pro-inflammatory adipokines leptin, TNF-α, and IL-6; while exhibiting a decrease in the anti-inflammatory adipokine; adiponectin. Dysregulation of these factors is thought to be a key mechanism of obesity associated cancers, contributing to increased activation of mitogenic pathways including PI3K and MAPK. Adiponectin represents an important negative regulator of leptin, TNF-α and IL-6. We previously demonstrated that adiponectin inhibits pancreatic cancer proliferation and tumor growth, however, the molecular mechanisms by which adiponectin regulates these processes are unknown. We hypothesize that Adiponectin Receptor (AdipoR) agonists elicit anti-tumor effects through suppression of RAS-MAPK mediated pathways and its downstream signaling components in pancreatic cancer progression. Experimental Procedures: The anti-tumor effects of AdipoRon, a novel small molecule agonist of the AdipoR, were assessed in vitro on human (MiaPaca-2 and Panc-1) and murine (P-4313 and K8484) pancreatic cancer cell lines. Cells were treated with AdipoRon in a dose-dependent manner and then assayed for cellular proliferation, apoptosis, colony formation and anchorage-independent growth. The effect of AdipoRon on activation of key RAS-MAPK signaling regulators was investigated by immunoblot analysis. To determine whether AdipoRon could inhibit the effects of obesity associated pro-tumorigenic cytokines, human and mouse pancreatic cancer cells were exposed to plasma collected from obese mice or specifically with recombinant cytokines. To determine whether AdipoRon could inhibit tumor growth in vivo, mice were orthotopically injected in the pancreas with the murine KrasG12D mutant P-4313 cell line. Tumors were allowed to establish for two weeks and treated with either vehicle or AdipoRon. Tumor size and number of Ki67 positive cells were assessed. Results: Compared to vehicle treatment, in vitro assessment confirmed that AdipoRon was highly effective at inhibiting cell proliferation, increasing apoptosis and preventing colony formation for all pancreatic cell lines tested. Anchorage independent growth was drastically reduced for both Panc1 (3.8 fold) and MiaPaca-2 (5.1 fold) cell lines in the presence of AdipoRon. Treatment of both murine and human pancreatic cancer cell lines with AdipoRon caused a significant dose dependent decrease in pSTAT3, pERK1, and pERK2 with a simultaneous increase in pAMPK. Importantly, AdipoRon completely antagonized the stimulatory effects of obese plasma or recombinant IL-6 on the activation of pSTAT3. Administration of AdipoRon to P-4313 orthotopic pancreatic tumor bearing mice resulted in four fold decrease in tumor size and a 50% reduction in tumor cell proliferation. Conclusions: AdipoRon, an adiponectin receptor agonist, suppresses KRAS signaling mediators ERK and STAT3 while simultaneously increasing AMPK resulting in inhibition of pancreatic cancer proliferation and tumor growth. Targeting of adiponectin receptors can provide a viable therapeutic strategy for the treatment of pancreatic cancer. Citation Format: Fanuel Messaggio, Alisha M. Mendonsa, Jason A. Castellanos, Casey Roberts, Nagaraj S. Nagathihalli, Nipun B. Merchant, Lee D. Gorden, Michael N. VanSaun.{Authors}. AdipoRon suppresses ERK and STAT3 to inhibit pancreatic cancer growth. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2016 May 12-15; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2016;76(24 Suppl):Abstract nr A46.

Abstract A38: Adiponectin receptor agonists cause inhibition of pancreatic cancer proliferation

Adiponectin is an anti-atherogenic hormone which is typically decreased in the setting of obesity, diabetes, and pancreatitis. Adiponectin and/or its receptor levels have been inversely correlated with risk for pancreatic cancer. The purpose of this study was to determine the effect of adiponectin on pancreatic cancer cells and to investigate the primary molecular pathways that it regulates. We have previously shown that pancreatic cancer cells primarily produce adiponectin receptor 1. In our current studies, recombinant adiponectin and a novel small molecular adiponectin agonist, AdipoRon, were used to stimulate human and murine pancreatic cancer cell lines in vitro. Proliferation was assessed through an EdU incorporation assay, metabolism was measured through MTT, and apoptosis was assessed through Annexin V staining. Results demonstrate that adiponectin stimulation leads to a decrease in proliferation of pancreatic cancer cells. Further, AdipoRon elicits a more potent suppression of proliferation and an additional increase in apoptosis. To better understand the molecular mechanisms activated by adiponectin stimulation, we performed western blot analysis of treated cells. We were able to confirm that AMPK was a direct target of both adiponectin and AdipoRon. Further, we were able to show that FBS stimulated phosphorylation of Akt was diminished in the presence of AdipoRon. Our data demonstrate that adiponectin and adiponectin receptor agonists act in a suppressive manner against pancreatic cancer cell proliferation. Current studies will determine whether adiponectin agonists can additionally decrease proliferation in vivo. Citation Format: Alisha Mendonsa, Lee Gorden, Michael Nathan VanSaun. Adiponectin receptor agonists cause inhibition of pancreatic cancer proliferation. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Innovations in Research and Treatment; May 18-21, 2014; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2015;75(13 Suppl):Abstract nr A38.

Abstract 4984: Contribution of MMP13 to tumor development in the steatotic liver microenvironment

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA With the rising prevalence of obesity there has been a marked increase in the incidence of non alcoholic fatty liver disease (NAFLD). Epidemiologically, NAFLD has been linked to an increased risk for development of primary liver cancer (HCC), however the mechanisms involved are not known. The liver is also a frequent site of metastasis for several types of cancers. To determine the effect of steatosis (fatty liver) on tumor metastasis to the liver, we used a mouse model of diet induced steatosis coupled with the splenic injection model of metastasis to the liver and found a significant increase in the number of tumors in the steatotic livers. To evaluate what molecular changes distinguish the steatotic liver from normal liver, microarray analysis was performed and demonstrated that MMP13, a member of the Matrix Metalloproteinase family recognized to be involved in liver disease and cancer progression, is significantly upregulated in the steatotic liver compared to normal livers of mice. We evaluated MMP13 expression in human patients with NAFLD and found that MMP13 is elevated with the progression of NAFLD. We hypothesize that increased MMP13 levels in the steatotic liver contribute to a more permissive microenvironment for the establishment of liver tumors. To test this hypothesis, mice genetically deficient in MMP13 or control wildtype mice, with and without steatosis, were injected with syngeneic MC38 colon cancer cells. Examination of the two groups demonstrated a significant decrease in the number of tumors in the MMP13 null mice (P <0.01). Additionally, since the tumor cells themselves express MMP13, wildtype mice were injected with either shRNA control or MMP13 knockdown MC38 colon cancer cells to determine the role of tumor cell derived MMP13. Using transwell migration assay and the platypus invasion system with parental and MMP13 shRNA knockdown MC38 cells we found that loss of MMP13 decreases the invasive and migratory properties of the metastatic cancer cells in vitro. In conclusion, we found that MMP13 is elevated in the setting of steatosis and that loss of both stromal and tumor derived MMP13 lead to decreased number of metastatic foci in the liver. Tumor derived MMP13 effects tumor cell migration and invasion invitro. MMP13 may thus be a potential target to control initiation and growth of metastatic cancer of the liver. Citation Format: Alisha Maria Mendonsa, Michael N. VanSaun, Lee Gorden. Contribution of MMP13 to tumor development in the steatotic liver microenvironment. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4984. doi:10.1158/1538-7445.AM2014-4984

Abstract LB-378: Establishment of hepatic metastases in a steatotic microenvironment is enhanced by stromal derived MMP-12

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC Non-alcoholic fatty liver disease (NAFLD), encompassing steatosis and progression to non-alcoholic steatohepatitis (NASH) are liver disorders of increasing clinical significance. We have shown that hepatic steatosis establishes a permissive microenvironment for metastatic seeding and tumor progression in the liver. We have identified MMP-12 (macrophage metalloelastase) as an important molecular component associated with hepatic steatosis and steatohepatitis. Wildtype mice in the C57Bl/6 background were fed a 42% fat diet for three months to induce hepatic steatosis. Affymetrix microarray analysis was performed on steatotic vs. normal murine livers to determine candidate genes altered between these liver microenvironments. Our results noted 715 significant changes in gene expression, of which matrix metalloproteinase 12 was among the most significantly up regulated genes in the steatotic microenvironment. Importantly, we have detected an upregulation of MMP-12 in human steatotic and steatohepatitis samples. MMP-12 deficient mice gained weight and developed diet-induced hepatic steatosis similar to wildtype mice. To determine whether MMP-12 affected metastasis, normal and steatotic MMP-12 deficient mice were tested with an experimental metastasis model via splenic injection of MC38 tumor cells. The number of resultant tumors per unit of tissue area in wildtype steatotic livers showed a 25 fold increase compared to tumors in MMP-12 deficient mice. Comparison of tumors between regular diet livers in wildtype and MMP12−/−mice was not significant. Immunohistochemical staining for F4/80 showed a reduction in the number of positive cells in MMP-12 deficient steatotic livers. Conclusions: Modulation of host factors is known to be important in tissue/site specific susceptibility to cancer metastases. MMP-12 can influence immune-mediated injury response by processing latent TNF alpha and regulating macrophage recruitment. Upregulation of matrix metalloproteinase 12 suggest a role for this protease in inflammatory mediated events in the distinct microenvironments of steatosis and steatohepatitis. The molecular mechanisms underlying the decreased establishment of metastatic tumors in MMP-12 deficient mice requires further investigation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr LB-378.