Experimental animal models of pancreatic cancer (review)

Pancreatic adenocarcinoma is currently the fifth leading cause of cancer-related death in the United States and is largely refractory to conventional therapies. The average survival from diagnosis to death is 4-6 months. The major cause of death is rapid development of metastasis involving the lymph nodes, liver, lungs, or peritoneum. To understand its etiology and eventually to make prevention possible and effective, appropriate carcinogenesis models will certainly help shed more light on the process of pancreatic carcinogenesis and help us understand the effects of environmental and genetic elements on pancreatic cancer development. The development of new treatment strategies to control cancer metastasis is of immediate urgency. Fulfillment of this difficult task relies on our knowledge of the cellular and molecular biology of pancreatic cancer metastasis, and the use of relevant animal models will certainly help define each aspect of this complicated process.     

Challenges and advances in mouse modeling for human pancreatic tumorigenesis and metastasis

Pancreatic cancer is critical for developed countries, where its rate of diagnosis has been increasing steadily annually. In the past decade, the advances of pancreatic cancer research have not contributed to the decline in mortality rates from pancreatic cancer—the overall 5-year survival rate remains about 5% low. This number only underscores an obvious urgency for us to better understand the biological features of pancreatic carcinogenesis, to develop early detection methods, and to improve novel therapeutic treatments. To achieve these goals, animal modeling that faithfully recapitulates the whole process of human pancreatic cancer is central to making the advancements. In this review, we summarize the currently available animal models for pancreatic cancer and the advances in pancreatic cancer animal modeling. We compare and contrast the advantages and disadvantages of three major categories of these models: (1) carcinogen-induced; (2) xenograft and allograft; and (3) genetically engineered mouse models. We focus more on the genetically engineered mouse models, a category which has been rapidly expanded recently for their capacities to mimic human pancreatic cancer and metastasis, and highlight the combinations of these models with various newly developed strategies and cell-lineage labeling systems.     

Histo-molecular oncogenesis of pancreatic cancer: From precancerous lesions to invasive ductal adenocarcinoma

Pancreatic cancer is a lethal malignancy, whose precursor lesions are pancreatic intraepithelial neoplasm, intraductal papillary mucinous neoplasm, intraductal tubulopapillary neoplasm, and mucinous cystic neoplasm. To better understand the biology of pancreatic cancer, it is fundamental to know its precursors and to study the mechanisms of carcinogenesis. Each of these precursors displays peculiar histological features, as well as specific molecular alterations. Starting from such pre-invasive lesions, this review aims at summarizing the most important aspects of carcinogenesis of pancreatic cancer, with a specific focus on the recent advances and the future perspectives of the research on this lethal tumor type.     

The multiple facets of drug resistance: one history,different approaches

Some cancers like melanoma and pancreatic and ovarian cancers, for example, commonly display resistance to chemotherapy, and this is the major obstacle to a better prognosis of patients. Frequently, literature presents studies in monolayer cell cultures, 3D cell cultures or in vivo studies, but rarely the same work compares results of drug resistance in different models. Several of these works are presented in this review and show that usually cells in 3D culture are more resistant to drugs than monolayer cultured cells due to different mechanisms. Searching for new strategies to sensitize different tumors to chemotherapy, many methods have been studied to understand the mechanisms whereby cancer cells acquire drug resistance. These methods have been strongly advanced along the years and therapies using different drugs have been increasingly proposed to induce cell death in resistant cells of different cancers. Recently, cancer stem cells (CSCs) have been extensively studied because they would be the only cells capable of sustaining tumorigenesis. It is believed that the resistance of CSCs to currently used chemotherapeutics is a major contributing factor in cancer recurrence and later metastasis development. This review aims to appraise the experimental progress in the study of acquired drug resistance of cancer cells in different models as well as to understand the role of CSCs as the major contributing factor in cancer recurrence and metastasis development, describing how CSCs can be identified and isolated.     

Vitamin E in the management of pancreatic cancer: A scoping review

Pancreatic cancer is the leading cause of cancer mortality worldwide. Research investigating effective management strategies for pancreatic cancer is ongoing. Vitamin E, consisting of both tocopherol and tocotrienol, has demonstrated debatable effects on pancreatic cancer cells. Therefore, this scoping review aims to summarize the effects of vitamin E on pancreatic cancer. In October 2022, a literature search was conducted using PubMed and Scopus since their inception. Original studies on the effects of vitamin E on pancreatic cancer, including cell cultures, animal models and human clinical trials, were considered for this review. The literature search found 75 articles on this topic, but only 24 articles met the inclusion criteria. The available evidence showed that vitamin E modulated proliferation, cell death, angiogenesis, metastasis and inflammation in pancreatic cancer cells. However, the safety and bioavailability concerns remain to be answered with more extensive preclinical and clinical studies. More in-depth analysis is necessary to investigate further the role of vitamin E in the management of pancreatic cancers.     

NF-κB in pancreatic cancer

Although the genetic profile of pancreatic cancer is emerging as a result of much research, the role of specific genetic alterations that initiate tumorigenesis and produce its cardinal clinical features of locally aggressive growth, metastasis, and chemotherapy resistance remains unresolved. Recently, a number of studies have shown that the inhibition of constitutive NF-κB activation, one of the frequent molecular alterations in pancreatic cancer, inhibits tumorigenesis and metastasis. It also sensitizes pancreatic cancer cell lines to anticancer agent-induced apoptosis. Therefore because of the crucial role of NF-κB in pancreatic cancer, it is a potential target for developing novel therapeutic strategies for the disease. In vivo and in vitro models that mimic the tumorigenic phenotypes in the appropriate histological and molecular concert would be very useful for confirming the suspected role of the pancreatic cancer signature genetic lesions and better understanding the molecular basis of this disease.     

Nek2 siRNA therapy using a portal venous port–catheter system for liver metastasis in pancreatic cancer

Nek2 (NIMA‐related kinase 2) is a serine‐threonine kinase and human homolog of the mitotic regulator NIMA of Aspergillus nidulan. We reported the efficiency of Nek2 siRNA in several cancer xenograft models using cholangiocarcinoma, breast cancer and colorectal cancer. Pancreatic cancer is difficult to treat due to its rapid progression and resistance to chemotherapy. Novel treatments are urgently required to improve survival in pancreatic cancer, and siRNA are a promising therapeutic option. However, finding an in vivo drug delivery system of siRNA remains a major problem for clinical application. In this study, the overexpression of Nek2 was identified in pancreatic cancer cell lines. Nek2 siRNA inhibited tumor growth in a subcutaneous xenograft mouse model of pancreatic cancer, prolonged the survival time in an intraperitoneal xenograft mouse model and efficiently prevented the progression of liver metastasis using a portal venous port–catheter system. Taken together, Nek2 is an effective therapeutic target in pancreatic cancer. An adequate delivery system is considered important in treating advanced pancreatic cancer, such as peritoneal dissemination and liver metastasis. Further investigations are required on the safety and side effects of the portal venous port–catheter system. We hope that Nek2 siRNA will be a novel therapeutic strategy for pancreatic cancer with liver metastasis and peritoneal dissemination.     

Early detection markers in Pancreas Cancer

The role of early detection in cancer has shown improved survival for certain cancers, including colon cancer, cervical cancer and breast cancer. The possibility for early detection of pancreatic cancer may be realized by an improved understanding of the histology and molecular genetics of precursor lesions and cancerous lesions in pancreatic cancer and the development of sensitive and specific screening tests (both invasive and non-invasive) to detect early pancreatic cancer. The NIH-NCI Early Detection Research Network (EDRN) in Pancreatic Cancer has been focussed on the development and validation of new biomarkers for the detection of early pancreatic cancer. This review will focus on our understanding of the histologic and molecular model of pancreatic carcinogenesis, current strategies and limitations of screening for pancreatic cancer and the development and validation of new biomarkers for the early detection of pancreatic cancer.     

乳腺癌移植转移模型的建立与应用

乳腺癌是女性最常见的恶性肿瘤之一,而转移是乳腺癌患者死亡的主要原因。乳腺癌转移模型是筛选乳腺癌转移相关基因、探讨肿瘤转移分子机制和评价抗转移实验性治疗疗效的重要工具。乳腺癌转移模型主要包括自发性转移模型、诱发性转移模型、转基因肿瘤转移模型和移植性转移模型。移植性肿瘤转移模型以其操作简单、重复性好和生物学性状稳定等优点应用最为广泛。乳腺癌移植性转移模型按建立方法可分为实验性转移模型和自发性转移模型。乳腺癌实验性转移模型是直接将瘤细胞注入血液循环系统后在远处驻留增殖形成转移灶,实验周期短、转移发生率高,费用低,但缺少原发灶的形成和肿瘤细胞从原发灶逃逸的过程。乳腺癌自发性转移模型是将瘤细胞或瘤块接种于皮下、肌肉和乳腺脂肪垫组织,在局部形成原发癌后自发转移,涉及了从原发灶产生到转移灶形成的乳腺癌转移的完整过程,是较好的研究乳腺癌转移机制的工具。近年来对移植转移模型转移灶的检测有了较大进展,采用活体动物成像技术不但能在肿瘤发生早期探测到各组织器官内的微小转移灶,还可以动态监测肿瘤细胞在动物体内的转移状态。本文对乳腺癌移植性转移模型的分类、模型建立方法、转移灶的检测,以及该类模型在乳腺癌转移分子机制和抗转移治疗研究中的应用进行综述。     

Genetic reduction of insulin-like growth factor-1 mimics the anticancer effects of calorie restriction on cyclooxygenase-2-driven pancreatic neoplasia

Risk of pancreatic cancer, the fourth deadliest cancer in the United States, is increased by obesity. Calorie restriction (CR) prevents obesity, suppresses carcinogenesis in many models, and reduces serum levels of IGF-1. In the present study, we examined the impact of CR on a model of inflammation-associated pancreatitis and pancreatic dysplasia, with a focus on the mechanistic contribution of systemic IGF-1. Administration of a 30% CR diet for 14 weeks decreased serum IGF-1 levels and hindered pancreatic ductal lesion formation and dysplastic severity, relative to a higher calorie control diet, in transgenic mice overexpressing COX-2 [bovine keratin-5 promoter (BK5.COX-2)]. These findings in CR mice correlated with reductions in Ki-67-positive cells, vascular luminal size, VEGF expression, and phosphorylation and total expression of downstream mediators of the IGF-1 pathway. Cell lines derived from BK5.COX-2 ductal lesions (JC101 cells) formed pancreatic tumors in wild-type FVB mice that were significantly reduced in size by a 14-week CR regimen, relative to the control diet. To further understand the impact of circulating levels of IGF-1 on tumor growth in this model, we orthotopically injected JC101 cells into liver-specific IGF-1-deficient (LID) mice. The approximate 65% reduction of serum IGF-1 levels in LID mice resulted in significantly decreased burden of JC101 tumors, despite modestly elevated levels of circulating insulin and leptin. These data show that CR prevents development of dysplasia and growth of pancreatic cancer through alterations in IGF-1, suggesting that modulation of this pathway with dietary and/or pharmacologic interventions is a promising pancreatic cancer prevention strategy.     

小鼠胰腺癌肝转移模型及相关分子表达的研究

  目的  经脾注射法建立小鼠胰腺癌肝转移模型，初步研究部分转移相关分子的基因表达水平变化。   方法  用脾脏注射法在12只C57/BL6小鼠中建立胰腺癌肝转移模型，6只小鼠注射PBS缓冲液作为对照。术后4周处死并解剖小鼠，观察腹腔内转移情况和肝脏成瘤的情况。采用实时荧光定量PCR检测小鼠脾脏胰腺癌组织、胰腺癌肝转移组织及肝转移灶旁肝组织中相关转移分子的基因表达情况，比较其差异。   结果  实验组12只小鼠的肝脏转移率为83.33%，解剖后观察脾脏表面均见单一瘤结节，其中10只小鼠肝脏表面及切面见多发散在的转移瘤结节，肝脏成瘤率83.33%，肝脏瘤组织细胞形态学符合胰腺癌的特征。在构建成功的10只胰腺癌肝转移小鼠模型中，CCL-17、CCL-22、CD44等分子在脾脏癌灶与肝转移灶的基因表达量高于癌旁肝组织，相对表达率 < 1（P < 0.05）；Ang-2、BAK、BAX等分子在脾脏癌灶与肝转移灶的基因表达量低于癌旁肝组织相对表达率>1 < 1（P < 0.05）。   结论  经脾注射法成功建立小鼠胰腺癌肝转移模型，能较好模拟胰腺癌体内肝转移过程。小鼠胰腺癌肝转移模型中相关转移分子基因表达水平的变化趋势与人体内胰腺癌肝转移基本一致，可为筛选抗肿瘤药物和研究胰腺癌转移行为提供帮助。      

Effect of angiostatin on liver metastasis of pancreatic cancer in hamsters.

The liver is the most common site of metastasis in pancreatic cancer, and there are no promising strategies to treat it. Angiostatin, a kringle-containing fragment of plasminogen, is a potent inhibitor of angiogenesis. The effect of angiostatin on liver metastasis in pancreatic cancer was investigated by using our established hamster model of liver metastasis. Pancreatic cancer cells (PGHAM-1, 1 x 10(6)) derived from N-nitrosobis(2-oxopropyl)amine (BOP)-induced pancreatic tumor in Syrian golden hamsters were transplanted into the spleen of female hamsters, and the animals were subcutaneously injected with angiostatin and saline. Subsequently, the macroscopic appearance of liver surface metastases was evaluated. In addition, histological sections of the liver metastases were analyzed for neovascularization, proliferation, and apoptosis on the basis of von Willebrand factor, argyrophilic nucleolar organizer region (Ag-NOR), and TdT-mediated dUTP-biotin nick end labeling (TUNEL) staining, respectively. The results showed significant tumor growth retardation and inhibition of angiogenesis in metastatic liver tumors in response to treatment with angiostatin. Moreover, the metastases remained in a nearly dormant state due to a balance between apoptosis and proliferation of the tumor, with no detectable side effects. This is the first experimental trial of angiostatin on pancreatic cancer and liver metastasis. The results suggest that angiostatin therapy could be effective against liver metastases of pancreatic cancer.     

Innovative Therapeutic Strategies Targeting Colorectal Cancer Stem Cells

Colorectal cancer is the fourth most common cause of cancer-related death. Although many advances in the treatment of this disease have been made, a large number of patients develop metastasis and resistance to current therapies. The current evidence indicates that cancer stem cells (CSCs) and epithelial-to-mesenchymal transition (EMT) have crucial roles in colorectal carcinogenesis and metastasis. It is also very important to understand the mechanisms that allow the survival of CSCs, such as metabolic reprogramming, which permits them to obtain specific properties or the activation of alternative signaling pathways in response to first-line therapies. In this review, we discuss the failure of conventional therapies for colorectal cancer and provide a brief analysis of new therapeutic strategies for targeting non-responsive CSC. We highlight the use of combination therapies, such as horizontal or vertical targeting, as the most efficient strategy for eradicating these subpopulations of cancer cells.     

Effect of Angiostatin on Liver Metastasis of Pancreatic Cancer in Hamsters

The liver is the most common site of metastasis in pancreatic cancer, and there are no promising strategies to treat it. Angiostatin, a kringle-containing fragment of plasminogen, is a potent inhibitor of angiogenesis. The effect of angiostatin on liver metastasis in pancreatic cancer was investigated by using our established hamster model of liver metastasis. Pancreatic cancer cells (PGHAM-1, 1x10⁶) derived from N-nitrosobis(2-oxopropyl)amine (BOP)-induced pancreatic tumor in Syrian golden hamsters were transplanted into the spleen of female hamsters, and the animals were subcutaneously injected with angiostatin and saline. Subsequently, the macroscopic appearance of liver surface metastases was evaluated. In addition, histological sections of the liver metastases were analyzed for neovascularization, proliferation, and apoptosis on the basis of von Willebrand factor, argyrophilic nucleolar organizer region (Ag-NOR), and TdT-mediated dUTP-biotin nick end labeling (TUNEL) staining, respectively. The results showed significant tumor growth retardation and inhibition of angiogenesis in metastatic liver tumors in response to treatment with angiostatin. Moreover, the metastases remained in a nearly dormant state due to a balance between apoptosis and proliferation of the tumor, with no detectable side effects. This is the first experimental trial of angiostatin on pancreatic cancer and liver metastasis. The results suggest that angiostatin therapy could be effective against liver metastases of pancreatic cancer.     

Hepatoma-derived growth factor involved in the carcinogenesis of gastric epithelial cells through promotion of cell proliferation by Erk1/2 activation

Hepatoma-derived growth factor (HDGF) is related to tumorigenesis and the development of cancer; it is an independent factor associated with the prognosis of liver cancer, non-small cell lung cancer and pancreatic cancer. However, the molecular mechanism by which HDGF participates in gastric carcinogenesis and development as well as its functional regulation during the development of gastric precancerous lesions needs to be further analyzed. In the present study, we analyzed the effect of HDGF transfection on the proliferation and on the changes of mitogen-activated protein kinase (MAPK), Akt, and nuclear factor-kB (NF-kB) pathways in gastric cancer AGS cells. HDGF transfection significantly activated Erk1/2 in AGS cells and promoted anchorage-independent growth. Further studies showed that HDGF expression gradually increased in the gastric carcinogenesis process and HDGF showed a high expression in poorly differentiated adenocarcinoma prone to lymphoid metastasis; these findings suggest that HDGF is involved in the gastric carcinogenesis process and promotes proliferation and metastasis via Erk1/2 activation.     

The pathobiological impact of cigarette smoke on pancreatic cancer development (review)

Despite extensive efforts, pancreatic cancer remains incurable. Most risk factors, such as genetic disposition, metabolic diseases or chronic pancreatitis cannot be influenced. By contrast, cigarette smoking, an important risk factor for pancreatic cancer, can be controlled. Despite the epidemiological evidence of the detrimental effects of cigarette smoking with regard to pancreatic cancer development and its unique property of being influenceable, our understanding of cigarette smoke-induced pancreatic carcinogenesis is limited. Current data on cigarette smoke-induced pancreatic carcinogenesis indicate multifactorial events that are triggered by nicotine, which is the major pharmacologically active constituent of tobacco smoke. In addition to nicotine, a vast number of carcinogens have the potential to reach the pancreatic gland, where they are metabolized, in some instances to even more toxic compounds. These metabolic events are not restricted to pancreatic ductal cells. Several studies show that acinar cells are also greatly affected. Furthermore, pancreatic cancer progenitor cells do not only derive from the ductal epithelial lineage, but also from acinar cells. This sheds new light on cigarette smoke-induced acinar cell damage. On this background, our objective is to outline a multifactorial model of tobacco smoke-induced pancreatic carcinogenesis.     

Vascular endothelial growth factor-D induces lymphangiogenesis and lymphatic metastasis in models of ductal pancreatic cancer

The presence of lymphatic metastases is a strong indicator for poor prognosis in patients with ductal pancreatic cancer. In order to better understand the mechanisms controlling lymphatic growth and lymph node metastasis in human ductal pancreatic cancer, we analyzed the expression pattern of the vascular endothelial growth factor-D (VEGF-D), its receptor VEGF-receptor-3 (VEGFR-3) and the lymphatic endothelium-specific hyaluronan receptor LYVE-1 in a panel of 19 primary human ductal pancreatic tumors and 10 normal pancreas specimens. We further addressed the biological function of VEGF-D for induction of lymphatic metastasis in a nude mouse xenograft model using two human ductal pancreatic cancer cell lines with overexpression of VEGF-D. Compared to normal human pancreas, pancreatic cancer tissue showed overexpression of VEGF-D and VEGFR-3 in conjunction with a high lymphatic vascularization as determined by immunohistochemistry and in situ hybridization. Tumors derived from VEGF-D-overexpressing cells had a higher microvessel density compared to their mock-controls, as determined based on CD31 immunohistochemistry. Importantly, these tumors also revealed a significant induction of intra- and peritumoral lymphatics, as judged from immunohistochemical detection of LYVE-1 expression. This was associated with a significant increase in lymphatic vessel invasion by tumor cells and an increased rate of lymphatic metastases, as indicated by pan-cytokeratin reactive cells in lymph nodes. Our results suggest that VEGF-D plays a pivotal role in stimulating lymphangiogenesis and lymphatic metastasis in human ductal pancreatic cancer, and therefore represents a novel therapeutic target for this devastating disease.     

Werner syndrome as a hereditary risk factor for exocrine pancreatic cancer: Potential role of WRN in pancreatic tumorigenesis and patient-tailored therapy

Advanced age is considered a risk factor for pancreatic cancer, but this relationship at the molecular and genetic level remains unclear. We present a clinical case series focusing on an association between pancreatic adenocarcinoma and Werner syndrome (WS) that is an autosomal recessive genetic disorder characterized by accelerated aging and cancer predisposition, and is caused by loss-of-function mutations in the WS RecQ helicase gene (WRN). Although pancreatic adenocarcinoma mostly occurs in a sporadic fashion, a minority of cases occurs in the context of susceptible individuals with hereditary syndromes. While WS has not been previously recognized as a risk factor for developing malignant tumors of the exocrine pancreas, the clinicopathologic features of three reported patients suggest a contributory role of WRN deficiency in pancreatic carcinogenesis. Molecular genetic analyses support the role of WRN as a tumor suppressor gene, although recent evidence reveals that WRN can alternatively promote oncogenicity depending on the molecular context. Based upon the clinico-pathologic features of these patients and the role of WRN in experimental models, we propose that its loss-of-function predisposes the development of pancreatic adenocarcinoma through epigenetic silencing or loss-of-heterozygosity of WRN. To test this hypothesis, we are investigating the mechanistic role of WRN in pancreatic cancer models including a pancreatic adenocarcinoma cell line generated from a human with WS. These studies are expected to provide new insight into the relationship between aging and pancreatic tumorigenesis, and facilitate development of novel strategies for patient-tailored interventions in this deadly malignancy.Key words: Werner syndrome, WRN, RecQ helicase, exonuclease, pancreatic adenocarcinoma, tumor suppressor gene, adult-onset progeria, DNA repair syndrome     

Pancreatic adenocarcinoma pathology: changing “landscape”

Pancreatic cancer is a devastating disease. At time of diagnosis the disease is usually advanced and only a minority of patients are eligible for surgical resection. The overall 5-year survival is 6%. However, survival of patients with early stage pancreatic cancer is significantly better. To improve the prognosis of patients with pancreatic cancer, it is essential to diagnose and treat pancreatic cancer in the earliest stage. Prevention of pancreatic cancer by treating noninvasive precursor lesions just before they invade tissues can potentially lead to even better outcomes. Pancreatic carcinogenesis results from a stepwise progression in which accumulating genetic alterations drive neoplastic progression in well-defined precursor lesions, ultimately giving rise to an invasive adenocarcinoma. A thorough understanding of the genetic changes that drive pancreatic carcinogenesis can lead to identification of biomarkers for early detection and targets for therapy. Recent next-generation sequencing (NGS) studies have shed new light on our understanding of the natural history of pancreatic cancer and the precursor lesions that give rise to these cancers. Importantly, there is a significant window of opportunity for early detection and treatment between the first genetic alteration in a cell in the pancreas and development of full-blown pancreatic cancer. The current views on the pathology and genetics of pancreatic carcinogenesis that evolved from studies of pancreatic cancer and its precursor lesions are discussed in this review.