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Covalent KRAS(G12C) inhibitors have recently shown promising efficacy in the clinic. However, the rapid development of drug resistance compromises their long-term benefits, indicating the need for additional approaches to treat KRAS-mutated cancers. Targeted protein degradation represents one such potential approach. We have therefore explored applying Chaperone-mediated Protein Degradation (CHAMP) technology to degrade KRAS(G12C) protein. CHAMPs are heterobifunctional small molecules that mediate protein degradation by inducing proximity between a target protein and the HSP90 chaperone complex, resulting in target ubiquitination and degradation by the proteasome. Due HSP90 being highly activated in cancer cells, CHAMPs also preferentially accumulate in tumors relative to normal tissues, resulting in an improved therapeutic index relative to typical inhibitors. In order to chemically induce KRAS(G12C) degradation, CHAMP compounds were synthesized by covalently coupling KRAS(G12C)- and HSP90-binding moieties through a short linker. CHAMP treatment of KRAS(G12C)-mutated cell lines resulted in formation of a KRAS(G12C)-CHAMP-HSP90 ternary complex and subsequent proteasome-dependent KRAS(G12C) degradation and inhibition of cell proliferation. In contrast, combination treatment with KRAS(G12C) and HSP90 inhibitors did not result in substantial KRAS(G12C) degradation. Importantly, CHAMPs retained potency in cell line models of resistance to covalent KRAS(G12C) inhibitors. Further, in mouse xenograft models, CHAMPs displayed prolonged pharmacokinetics in tumors relative to plasma and normal tissues and strongly inhibited tumor growth at tolerated doses. Based on these findings, KRAS(G12C)-degrading CHAMPs represents a promising approach to the treatment of KRAS(G12C)-mutated cancers. Citation Format: Kevin P. Foley, Chenghao Ying, Yaya Wang, Yan Dai, Zhiyong Wang, Jinhua Li, Zimo Yang, Yuetong Sun, Hao Xin Zhou, Thomas L. Prince, Guoqiang Wang, Weiwen Ying. Chemically induced chaperone-mediated protein degradation (CHAMP) of KRAS(G12C) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1649.

Molecular hydrogen (H2 ) has been proven to be an effective agent that can cure multiple organ diseases by reducing oxidative stress. Although the protective effect of hydrogen on acute pancreatitis (AP) has been confirmed, its molecular mechanism is still unclear. In this article, we aimed to investigate the changes in pancreatic cell protein expression associated with the protective effect of H2 against AP and attempted to uncover the molecular mechanism underlying this process. A proteomic analysis identified 73 differentially expressed proteins and generated the protein-protein interaction networks of these proteins. The results triggered our interest in mitogen-activated protein kinase (MAPK) and heat shock cognate 71 kDa protein (Hsc70). The subsequent in vitro experiments showed that H2 treatment inhibited the phosphorylation of extracellular signal-regulated kinase (ERK), c-jun N-terminal kinase (JNK), and p38 MAPK, and activated NF-κB and the expression of tumor necrosis factor α and interleukin-1β, while simultaneously preventing the translocation of phospho-ERK, phospho-JNK, and phospho-p38 from the cytoplasm to the nucleus. Furthermore, Hsc70 expression was upregulated by H2 administration. The animal experimental results were consistent with those of the in vitro experiments. In conclusion, H2 treatment can ameliorate the inflammatory response and reduce the expression of inflammatory mediators during the early phase of AP by inhibiting the MAPK pathways and increasing Hsc70 expression.

We aimed to investigate the relationship between the synthesis of hydrogen sulfide (H(2)S) and the pancreatic acinar cell apoptosis in severe acute pancreatitis (SAP) rats, as well as analyse the potential apoptotic pathway involved in this process. Sixty rats had been equally divided into four groups: sham, SAP, SAP + sodium hydrosulfide (NaHS) and SAP + DL-propargylglycine (PAG). 24 h after SAP induction, all surviving animals of each group were sacrificed to collect blood and tissue samples for the following measurements: the level of serum H(2)S as well as the levels of tumor necrosis factor-alpha (TNF-α), interleukin-10 (IL-10), H(2)S synthesizing activity, CSE mRNA and protein expression, maleic dialdehyde (MDA) and myeloperoxidase (MPO) activity, the expression of Bax, Bcl-2, caspase-3, -8 and -9, the release of cytochrome c and the activation of nuclear factor-kappa B (NF-κB), ERK1/2, JNK1/2 and p38 in pancreas. Furthermore, in situ detection of cell apoptosis was examined and the severity of pancreatic damage was analyzed by pathological grading and scoring. Results Significant differences in every index except IL-10 had been found between the SAP, NaHS and PAG groups (P 0.05). Treatment with PAG obviously induced the pancreatic acinar cell apoptosis as well as improved all the pathological changes and inflammatory parameters. In contrast, administration of NaHS significantly attenuated apoptosis in the pancreas and aggravated the severity of pancreatic damage. Moreover, the expressions of caspase-3, -8, -9 and the release of cytochrome c were all increased in the apoptotic cells, and the activity of NF-κB as well as the phosphorylation of ERK1/2, JNK1/2 and p38 decreased accompanying with the reduction of the serum H(2)S level. H(2)S plays a pivotal role in the regulation of pancreatic acinar cell apoptosis in SAP rats. The present results showed that inhibition of H(2)S synthesis provided protection for SAP rats via inducing acinar cell apoptosis. This process acted through both extrinsic and intrinsic apoptotic pathways, and may be regulated by reducing the activity of NF-κB.

To investigate the function of nuclear factor (NF)-κB in the epithelial to mesenchymal transition induced by hypoxia in pancreatic cancer cells.For cultured pancreatic cancer cells (BxPC-3 and Panc-1) under hypoxic and normoxic conditions, the differences in the morphology were observed by optical microscope. The expression of markers of epithelial and mesenchymal phenotypes, E-cadherin, vimentin and N-cadherin, were determined by Western blot. NF-κB P65 activity was measured by electrophoretic mobility shift assay. Invasion and gemcitabine resistance of pancreatic cancer cells were evaluated in matrigel invasion assay and cell counting kit-8 assay. Both molecular and pharmacologic means of inhibiting NF-κB P65 were used in these hypoxic cells and then the above resulting phenotypes were compared with those of the control-treated cells.After cultured pancreatic cancer cells under hypoxic conditions for 48 h, normoxic cells exhibited a polygonal shape and formed tight clusters of cells, whereas hypoxic cells took on an elongated, fibroblastoid morphology associated with a more highly invasive character and resistance to gemcitabine; hypoxic cells exhibited an suppression of E-cadherin and increase in vimentin and N-cadherin expression. NF-κB P65 activity was elevated in hypoxic cells. On the contrary, on molecular or pharmacologic inhibition of NF-κB P65, hypoxic cells regained expression of E-cadherin, lost expression of N-cadherin, and reversed their highly invasive and drug resistant phenotype.Pancreatic cancer cells underwent epithelial to mesenchymal transition exposed to hypoxia, exhibited highly invasive and drug resistant phenotype. Inhibition of NF-κB P65 under hypoxic conditions, pancreatic cancer cells regained expression of E-cadherin, lost expression of N-cadherin, and reversed their highly invasive and drug resistant phenotype.

Dihydroartemisinin (DHA) has recently shown antitumor activity in human pancreatic cancer cells. However, its effect on antiangiogenic activity in pancreatic cancer is unknown, and the mechanism is unclear. This study was aimed to investigate whether DHA would inhibit angiogenesis in human pancreatic cancer.Cell viability and proliferation, tube formation of human umbilical vein endothelial cells (HUVECs), nuclear factor (NF)-κB DNA-binding activity, expressions of vascular endothelial growth factor (VEGF), interleukin (IL)-8, cyclooxygenase (COX)-2, and matrix metalloproteinase (MMP)-9 were examined in vitro. The effect of DHA on antiangiogenic activity in pancreatic cancer was also assessed using BxPC-3 xenografts subcutaneously established in BALB/c nude mice.DHA inhibited cell proliferation and tube formation of HUVECs in a time- and dose-dependent manner and also reduced cell viability in pancreatic cancer cells. DHA significantly inhibited NF-κB DNA-binding activity, so as to tremendously decrease the expression of NF-κB-targeted proangiogenic gene products: VEGF, IL-8, COX-2, and MMP-9 in vitro. In vivo studies, DHA remarkably reduced tumor volume, decreased microvessel density, and down-regulated the expression of NF-κB-related proangiogenic gene products.Inhibition of NF-κB activation is one of the mechanisms that DHA inhibits angiogenesis in human pancreatic cancer. We also suggest that DHA could be developed as a novel agent against pancreatic cancer.

Background: Epithelial to mesenchymal transition (EMT) induced by hypoxia is one of the critical causes of treatment failure in different types of human cancers. NF-kB is closely involved in the progression of EMT. Compared with HIF-1a, the correlation between NF-kB and EMT during hypoxia has been less studied, and although the phenomenon was observed in the past, the molecular mechanisms involved remained unclear. Methodology/Principal Findings: Here, we report that hypoxia or overexpression of hypoxia-inducible factor-1a (HIF-1a) promotes EMT in pancreatic cancer cells. On molecular or pharmacologic inhibition of NF-kB, hypoxic cells regained expression of E-cadherin, lost expression of N-cadherin, and attenuated their highly invasive and drug-resistant phenotype. Introducing a pcDNA3.0/HIF-1a into pancreatic cancer cells under normoxic conditions heightened NF-kB activity, phenocopying EMT effects produced by hypoxia. Conversely, inhibiting the heightened NF-kB activity in this setting attenuated the EMT phenotype. Conclusions/Significance: These results suggest that hypoxia or overexpression of HIF-1a induces the EMT that is largely dependent on NF-kB in pancreatic cancer cells.

To summary the experience of the surgical comprehensive treatment of severe acute pancreatitis (SAP).From July 1999 to December 2009, a total of 506 patients suffered SAP were admitted with a mean APACHE II score 12.8 ± 4.6. There were 270 male and 236 female, aged from 16 to 89 years, mean age 43 years. SAP patients were treated by the SAP treatment team which consisted of pancreatic specialized and multidisciplinary doctors. Two hundreds and thirty-four cases (46.2%) received non-operative treatment and 272 cases (53.8%) received surgical intervention.In 506 cases, 445 patients were cured and 52 patients died (31 died in early stage, 21 died in later stage), 9 cases discharged automatically. The overall incidence of complication, overall mortality and overall curative rate were 29.4% (149/506), 10.3% (52/506) and 87.9% (445/506), respectively. The incidences of complication in non-operative group and in surgical intervention group were 27.8% (65/234) and 30.9% (84/272), respectively (P0.05). The mortality in non-operative group and in surgical intervention group were 9.4% (22/234) and 11.0% (30/272), respectively (P0.05). The curative rates in non-operative group and in surgical intervention group were 90.6% (212/234) and 85.7% (233/272), respectively (P0.05).Patients should be treated in ICU in the early phase of the disease when APACHE II score10. Pancreatic specialized and multidisciplinary team treatment, appropriate choice of timing, indication and procedure of surgical intervention and details of drainage are vital to the prognosis of SAP.

Despite rapid advances in chemotherapy and surgical resection strategies, pancreatic cancer remains the fourth leading cause of cancer related deaths in the United States with a 5-year survival rate of less than 5%. Therefore, novel therapeutic agents for the prevention and treatment of pancreatic cancer are urgently needed. The aim of this study was to investigate the effect of pristimerin, a quinonemethide triterpenoid compound isolated from Celastraceae and Hippocrateaceae, on inhibition of cell proliferation and induction of apoptosis in three pancreatic cancer cells, BxPC-3, PANC-1 and AsPC-1, in both monotherapy and in combination with gemcitabine. Treatment with pristimerin decreased the cell proliferation of all three pancreatic cancer cells in a dose- and time-dependent manner. Treatment of pancreatic cancer cells with pristimerin also resulted in G1-phase arrest which was strongly associated with a marked decrease in the level of cyclins (D1 and E) and cyclin-dependent kinases (cdk2, cdk4 and cdk6 ) with concomitant induction of WAF1/p21 and KIP1/p27. Pristimerin treatment also resulted in apoptotic cell death, cleavage of caspase-3, modulation in the expressions of Bcl-2 family proteins, inhibition of the translocation and DNA-binding activity of NF-κB. In addition, pristimerin potentiated the growth inhibition and apoptosis inducing effects of gemcitabine in all three pancreatic cancer cells, at least in part, by inhibiting constitutive as well as gemcitabine-induced activation of NF-κB in both its DNA-binding activity and transcriptional activity. Taken together, these data provide the first evidence that pristimerin has strong potential for development as a novel agent against pancreatic cancer.