Regulation Of KRAS Research Articles

Anti-proteolytic regulation of KRAS by USP9X/NDRG3 in KRAS-driven cancer development

Cancers with activating mutations of KRAS show a high prevalence but remain intractable, requiring innovative strategies to overcome the poor targetability of KRAS. Here, we report that KRAS expression is post-translationally up-regulated through deubiquitination when the scaffolding function of NDRG3 (N-Myc downstream-regulated gene 3) promotes specific interaction between KRAS and a deubiquitinating enzyme, USP9X. In KRAS-mutant cancer cells KRAS protein expression, downstream signaling, and cell growth are highly dependent on NDRG3. In conditional KrasG12D knock-in mouse models of pancreatic ductal adenocarcinoma, Ndrg3 depletion abolishes Kras protein expression and suppresses intraepithelial neoplasia formation in pancreas. Mechanistically, KRAS protein binds to the C-terminal serine/threonine-rich region of NDRG3, subsequently going through deubiquitination by USP9X recruited to the complex. This interaction can be disrupted in a dominant-negative manner by a C-terminal NDRG3 fragment that binds KRAS but is defective in USP9X binding, highly suppressing KRAS protein expression and KRAS-driven cell growth. In summary, KRAS-driven cancer development critically depends on the deubiquitination of KRAS protein mediated by USP9X/NDRG3, and KRAS-addicted cancers could be effectively targeted by inhibiting the KRAS-NDRG3 interaction.

The molecular mechanism underlying KRAS regulation on STK31 expression in pancreatic ductal adenocarcinoma.

KRAS gene mutations are common in pancreatic ductal adenocarcinoma (PDAC), but targeting mutant KRAS is still challenging. Here, an endoribonuclease-prepared small interfering RNA (esiRNA) library was used to screen new kinases that play critical roles in PDAC driven by KRAS gene mutations, and serine/threonine kinase 31 (STK31) was identified and characterized as a potential therapeutic target for KRAS-mutant PDAC. Our results showed that STK31 was upregulated in KRAS-mutant PDAC patients with poor survival and highly expressed in PDAC cell lines with KRASG12D mutation. Inhibition of STK31 in KRAS-mutant cell lines significantly reduced PDAC cell growth invitro and hindered tumor growth invivo. Gain and loss of function experiments revealed that STK31 is a downstream target of KRAS in PDAC. A pharmacological inhibition assay showed MAPK/ERK signaling involved in STK31 regulation. The further mechanistic study validated that c-Jun, regulated by KRAS/MAPK signaling, directly modulates the transcription level of STK31 by binding to its promoter region. Through RNA sequencing, we found that the cell cycle regulators CCNB1 and CDC25C are downstream targets of STK31. Taken together, our results indicate that STK31, which is the downstream target of the KRAS/MAPK/ERK/c-Jun signaling pathway in KRAS-mutant PDAC, promotes PDAC cell growth by modulating the expression of the cell cycle regulators CCNB1 and CDC25C.

Abstract 1671: Exploring the mechanism of PP2A regulated cell death via macropinocytosis in pancreatic cancer

Abstract Pancreatic Ductal Adenocarcinoma (PDAC) is the 4th leading cause of cancer-related death in the United States. KRAS is an oncogene mutated in 90% of PDAC patients. Oncogenic KRAS leads to aberrant cell proliferation and survival. KRAS is considered hard to target because resistance mechanisms to KRAS inhibitors are common highlighting the need to understand alternative strategies. PDAC tumor microenvironment evolves with cancer cell progression leading to vascular remodeling and blood vessel collapse. This makes the tumors nutrient deplete. To circumvent this, cells employ KRAS-dependent macropinocytosis- a nutrient scavenging pathway. Macropinocytosis is the process by which cells take up extracellular material by membrane ruffling to form vesicles which then fuse with lysosomes to release nutrients. Targeting KRAS-driven macropinocytosis can be crucial to limiting nutrients to the cell to prevent tumor growth. Protein Phosphatase 2A (PP2A), a heterotrimeric Serine/Threonine phosphatase, is a downstream regulator of KRAS. Its activity is repressed by mutated KRAS. We have seen that the activation of PP2A with small molecular activator, DT061, prevents macropinosomes from fusing with lysosomes which leads to cell death. But how DT061 regulated PP2A activity affects the macropinocytosis pathway is still unknown. The purpose of this study is to understand the mechanism of cell death brough upon by the activation of PP2A with DT061. Membrane ruffling is critical for the initiation of macropinocytosis. Rac is a GTPase usually involved with membrane ruffling. To show that Rac is needed for DT061 mediated macropinocytosis, we used EHT 1864, a Rac family inhibitor. Cells were treated with EHT 1864 before being treated with DT061 and stained with crystal violet. The inhibition of Rac prevented DT061 driven cell death, indicating that Rac is crucial to DT061 driven cell death. High molecular weight TMR-Dextran can only be taken up by the cells through macropinocytosis. Cells were treated with TMR-Dextran and EHT 1864. Cells with Rac inhibition showed lower uptake of TMR-Dextran than cells that were treated with DT061. Together these findings indicate to us that Rac plays a crucial role in the initiation of macropinocytosis and the regulation of cell death caused by the activation of PP2A. Understanding the pathway of DT061 mediated cell death can be vital to developing this concept as a therapeutic method of treatment for PDAC patients who currently have the lowest 5-year survival rate (12%) of all cancers. Citation Format: Indiraa Doraivel, Garima Baral, Claire M. Pfeffer, Brittany L. Allen-Petersen. Exploring the mechanism of PP2A regulated cell death via macropinocytosis in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1671.

The effects of miR-217 inhibitor and mimic in the progression of Kirsten rat sarcoma viral oncogene homologue driven cancers

BackgroundKirsten rat sarcoma viral oncogene homologue (KRAS) is one of the most frequently mutated proto-oncogenes in approximately 90% of pancreatic ductal carcinoma (PDAC) and 45% of colorectal cancer (CC) cases. Studies in the past have identified microRNA-217 (miR-217) as a potential tumour-suppressing miRNA that is downregulated in various cancers. Using in silico prediction algorithms, several studies have identified miR-217 as a potential regulator of KRAS, and we investigated its role in PDAC and CC progression. MethodThe study was carried out in KRAS-driven cancer (KDC) cell lines PANC-1 (pancreatic cancer) and SW-480 (CC), which have mutant KRAS gene expression. The KDC cells are transfected with specific oligonucleotides for miR-217, anti-miR-217, and a negative control in serum-free media using lipofectamine. After fixing the IC50, using specific primers, gene expression studies were carried out by qPCR for KRAS downstream targets and genes associated with apoptosis and cell cycle. Anti-migration and anti-apoptotic effects were studied using the transwell migration assay and annexin-V/PI staining methods, and mitochondrial morphology was observed using a transmission electron microscope. ResultsThe present study demonstrates that overexpression of miR-217 in KDC cells mitigates proliferation and migration and promotes cell cycle arrest and apoptosis of KDC cells via the MAPK/ERK signalling pathway. Besides, decreased miR-217 expression rescues KDC cells from the effects mediated by KRAS downstream signalling. ConclusionThe outcome of the study indicates miR-217 suppresses tumour growth and promotes apoptosis in KDC and that these effects are associated with down regulation of MAPK/ERK signalling.

CRNDE mediated hnRNPA2B1 stability facilitates nuclear export and translation of KRAS in colorectal cancer

Development of colorectal cancer (CRC) involves activation of Kirsten rat sarcoma viral oncogene homolog (KRAS) signaling. However, the post-transcriptional regulation of KRAS has yet to be fully characterized. Here, we found that the colorectal neoplasia differentially expressed (CRNDE)/heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNPA2B1) axis was notably elevated in CRC and was strongly associated with poor prognosis of patients, while also significantly promoting CRC cell proliferation and metastasis both in vitro and in vivo. Furthermore, CRNDE maintained the stability of hnRNPA2B1 protein by inhibiting E3 ubiquitin ligase TRIM21 mediated K63 ubiquitination-dependent protein degradation. CRNDE/hnRNPA2B1 axis facilitated the nuclear export and translation of KRAS mRNA, which specifically activated the MAPK signaling pathway, eventually accelerating the malignant progression of CRC. Our findings provided insight into the regulatory network for stable hnRNPA2B1 protein expression, and the molecular mechanisms by which the CRNDE/hnRNPA2B1 axis mediated KRAS nucleocytoplasmic transport and translation, deeply underscoring the bright future of hnRNPA2B1 as a promising biomarker and therapeutic target for CRC.By hindering hnRNPA2B1 from binding to the E3 ubiquitin ligase TRIM21, whose mediated ubiquitin-dependent degradation was thereby inhibited, CRNDE protected the stability of hnRNPA2B1’s high protein expression in CRC. Supported by the high level of the oncogenic molecule CRNDE, hnRNPA2B1 bound to KRAS mRNA and promoted KRAS mRNA nucleus export to enter the ribosomal translation program, subsequently activating the MAPK signaling pathway and ultimately accelerating the malignant progression of CRC.

Association of the rs8720 and rs12587 KRAS Gene Variants with Colorectal Cancer in a Mexican Population and Their Analysis In Silico.

Colorectal cancer (CRC) is a major global health challenge and one of the top 10 cancers in Mexico. Lifestyle and genetic factors influence CRC development, prognosis, and therapeutic response; identifying risk factors, such as the genes involved, is critical to understanding its behavior, mechanisms, and prognosis. The association between KRAS gene variants (rs8720 and rs12587) and CRC in the Mexican population was analyzed. We performed in silico analysis and analyzed 310 healthy individuals and 385 CRC patients using TaqMan assays and real-time PCR. The CC and GG genotypes of rs8720 and rs12587 were identified as CRC risk factors (p < 0.05). The CC and TC genotypes of the rs8720 were associated with rectal cancer, age over 50 years, moderately differentiated histology, and advanced cancer stage. TG and GG genotypes of the rs12587 variant were a risk factor in the CRC group, in patients with stage I-II, males, and stage III-IV non-chemotherapy response. The TG haplotype is protected against CRC. The combined CCGG genotype was linked to CRC risk. In silico analysis revealed that the rs12587 and rs8720 variants could influence KRAS gene regulation via miRNAs. In conclusion, rs8720 and rs12587 variants of the KRAS gene were associated with CRC risk and could influence KRAS regulation via miRNAs.

KRAS regulation of miRNA: Stepping on the brake to go faster

In this issue of Molecular Cell, Shui etal.1 use a systems biology approach to unravel a paradoxical role of microRNA in oncogenic KrasG12D regulation of gene and protein expression.

Design of Orally-bioavailable Tetra-cyclic phthalazine SOS1 inhibitors with high selectivity against EGFR

KRAS mutations (G12C, G12D, etc.) are implicated in the oncogenesis and progression of many deadliest cancers. Son of sevenless homolog 1 (SOS1) is a crucial regulator of KRAS to modulate KRAS from inactive to active states. We previously discovered tetra-cyclic quinazolines as an improved scaffold for inhibiting SOS1-KRAS interaction. In this work, we report the design of tetra-cyclic phthalazine derivatives for selectively inhibiting SOS1 against EGFR. The lead compound 6c displayed remarkable activity to inhibit the proliferation of KRAS(G12C)-mutant pancreas cells. 6c showed a favorable pharmacokinetic profile in vivo, with a bioavailability of 65.8% and exhibited potent tumor suppression in pancreas tumor xenograft models. These intriguing results suggested that 6c has the potential to be developed as a drug candidate for KRAS-driven tumors.

Abstract 2218: The molecular mechanism underlying KRAS regulation on STK31 expression in pancreatic ductal adenocarcinoma

Abstract KRAS is frequently mutated in pancreatic ductal adenocarcinoma (PDAC). However, targeting mutant KRAS is still challenging. Kinase inhibitors are ideal targeted therapeutics for mutant KRAS-driven cancer. In our previous study, an esiRNA screening was prepared to identify some vital kinases that took part in KRAS mutant-driven pancreatic cancer. The STK31 was identified as a potential therapeutic target in KRAS mutant PDAC. This study aimed to investigate the underlying mechanism of STK31 in KRAS mutant PDAC as well as its therapeutic potential. The results demonstrated that STK31 was upregulated in KRAS mutant PDAC patients with poor survival. STK31 was also highly expressed in PDAC cell lines with KRAS mutant background. Inhibition of STK31 in KRAS mutant cell lines significantly reduced PDAC cell proliferation, colony formation, and migration. Gain and loss of function experiments revealed that STK31 was a downstream target of KRAS in PDAC. Pharmacological inhibition of several signaling pathways involved in KRAS showed MAPK/ERK signaling involved in STK31 regulation. Then, further mechanistic study validated that c-Jun, regulated by KRAS/MAPK/ERK signaling, directly regulated the transcription level of STK31 by binding to its promoter region. In summary, our results show that STK31 promotes PDAC cell proliferation under the control of the KRAS/MAPK/ERK/c-Jun transcriptional axis in PDAC. STK31 may act as a potential target in PDAC treatment. Citation Format: Yuting Liu, Shing Chun Tang, Yangchao Chen. The molecular mechanism underlying KRAS regulation on STK31 expression in pancreatic ductal adenocarcinoma [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 2218.

Design and Structural Optimization of Orally Bioavailable SOS1 Inhibitors for the Treatment of KRAS-Driven Carcinoma.

KRAS mutations (G12C, G12D, etc.) are implicated in the oncogenesis and progression of many refractory cancers. Son of sevenless homolog 1 (SOS1) is a key regulator of KRAS to modulate KRAS from inactive to active states. Herein, we disclosed efficacy-improving tetra-cyclic quinazoline derivatives as an enhanced scaffold for inhibiting the SOS1-KRAS interaction. Compound 37, which conjugated 1-carbonitrile-cyclopropane to tetra-cyclic quinazoline, showed a twofold higher oral drug exposure and 2.5-fold longer half-life than BI-3406 in CD-1 mouse plasma. In a Mia-paca-2 xenograft model, 37 administrated alone inhibited tumor growth by 71%. Preclinical investigations demonstrated that 37 had a limited inhibition of CYP and hERG. Overall, our studies showed that 37 was a promising drug candidate for treatment of KRAS-driven cancer.

KRAS Affects the Lipid Composition by Regulating Mitochondrial Functions and MAPK Activation in Bovine Mammary Epithelial Cells.

Kirsten rat sarcoma viral oncogene homolog (KRAS), or guanosine triphosphatase KRAS, is a proto-oncogene that encodes the small guanosine triphosphatase transductor protein. Previous studies have found that KRAS can promote cytokine secretion, cell chemotaxis, and survival. However, its effects on milk fat synthesis in bovine mammary epithelial cells are unclear. In this study, the effects of KRAS inhibition on cell metabolism, autophagy, oxidative stress, endoplasmic reticulum stress, mitochondrial function, and lipid composition as well as the potential mechanisms were detected in an immortalized dairy cow mammary epithelial cell line (MAC-T). The results showed that inhibition of KRAS changed the lipid composition (especially the triglyceride level), mitochondrial functions, autophagy, and endoplasmic reticulum stress in cells. Moreover, KRAS inhibition regulated the levels of the mammalian target of rapamycin and mitogen-activated protein kinase (extracellular regulated protein kinases, c-Jun N-terminal kinases, p38) activation. These results indicated that regulation of KRAS would affect the synthesis and composition of milk fat. These results are also helpful for exploring the synthesis and secretion of milk fat at the molecular level and provide a theoretical basis for improving the percentage of fat in milk and the yield of milk from cows.

Indoloquinoline-Mediated Targeted Downregulation of KRAS through Selective Stabilization of the Mid-Promoter G-Quadruplex Structure.

KRAS is a well-validated anti-cancer therapeutic target, whose transcriptional downregulation has been demonstrated to be lethal to tumor cells with aberrant KRAS signaling. G-quadruplexes (G4s) are non-canonical nucleic acid structures that mediate central dogmatic events, such as DNA repair, telomere elongation, transcription and splicing events. G4s are attractive drug targets, as they are more globular than B-DNA, enabling more selective gene interactions. Moreover, their genomic prevalence is increased in oncogenic promoters, their formation is increased in human cancers, and they can be modulated with small molecules or targeted nucleic acids. The putative formation of multiple G4s has been described in the literature, but compounds with selectivity among these structures have not yet been able to distinguish between the biological contribution of the predominant structures. Using cell free screening techniques, synthesis of novel indoloquinoline compounds and cellular models of KRAS-dependent cancer cells, we describe compounds that choose between KRAS promoter G4near and G4mid, correlate compound cytotoxic activity with KRAS regulation, and highlight G4mid as the lead molecular non-canonical structure for further targeting efforts.

Mutant KRAS modulates colorectal cancer cells invasive response to fibroblast-secreted factors through the HGF/C-MET axis.

Genetic alterations influencethe malignant potential of cancer cells, and so does the tumor microenvironment. Herein, we combined the study of KRAS oncogenic effects in colorectal cancer cells with the influence of fibroblast-derived factors. Results revealed that mutant KRAS regulates cell fate through both autonomous and nonautonomous signaling mechanisms. Specifically, processes such as proliferation and cell-cell aggregation were autonomously controlled by mutant KRAS independently of the stimulation with fibroblasts conditioned media. However, cancer cell invasion revealed to be a KRAS-dependent nonautonomous effect, resulting from the cooperation between fibroblast-derived HGF and mutant KRAS regulation of C-MET expression. C-MET downregulation upon KRAS silencing rendered cells less responsive to HGF and thus less invasive. Yet, in one cell line, KRAS inhibition triggered invasion upon stimulation with fibroblasts conditioned media. Inhibition of PIK3CA oncogene did not promote invasion, thus showing a KRAS-specific effect. Moreover, the invasive capacity also depended on the HGF-C-MET axis. Overall, our study awards oncogenic KRAS an important role in modulating the response to fibroblast-secreted factors either by promoting or impairing invasion, and depicts the HGF-C-MET axis as a putative therapeutic target to impair the invasive properties of mutant KRAS cancer cells.

Mechanistic Insights into the Long-range Allosteric Regulation of KRAS Via Neurofibromatosis Type 1 (NF1) Scaffold Upon SPRED1 Loading

Allosteric regulation is the most direct and efficient way of regulating protein function, wherein proteins transmit the perturbations at one site to another distinct functional site. Deciphering the mechanism of allosteric regulation is of vital importance for the comprehension of both physiological and pathological events in vivo as well as the rational allosteric drug design. However, it remains challenging to elucidate dominant allosteric signal transduction pathways, especially for large and multi-component protein machineries where long-range allosteric regulation exits. One of the quintessential examples having long-range allosteric regulation is the ternary complex, SPRED1–RAS–neurofibromin type 1 (NF1, a RAS GTPase–activating protein), in which SPRED1 facilitates RAS–GTP hydrolysis by interacting with NF1 at a distal, allosteric site from the RAS binding site. To address the underlying mechanism, we performed extensive Gaussian accelerated molecular dynamics simulations and Markov state model analysis of KRAS–NF1 complex in the presence and absence of SPRED1. Our findings suggested that SPRED1 loading allosterically enhanced KRAS–NF1 binding, but hindered conformational transformation of the NF1 catalytic center for RAS hydrolysis. Moreover, we unveiled the possible allosteric pathways upon SPRED1 binding through difference contact network analysis. This study not only provided an in-depth mechanistic insight into the allosteric regulation of KRAS by SPRED1, but also shed light on the investigation of long-range allosteric regulation among complex macromolecular systems.

Targeting KRAS Regulation with PolyPurine Reverse Hoogsteen Oligonucleotides.

KRAS is a GTPase involved in the proliferation signaling of several growth factors. The KRAS gene is GC-rich, containing regions with known and putative G-quadruplex (G4) forming regions. Within the middle of the G-rich proximal promoter, stabilization of the physiologically active G4mid structure downregulates transcription of KRAS; the function and formation of other G4s within the gene are unknown. Herein we identify three putative G4-forming sequences (G4FS) within the KRAS gene, explore their G4 formation, and develop oligonucleotides targeting these three regions and the G4mid forming sequence. We tested Polypurine Reverse Hoogsteen hairpins (PPRHs) for their effects on KRAS regulation via enhancing G4 formation or displacing G-rich DNA strands, downregulating KRAS transcription and mediating an anti-proliferative effect. Five PPRH were designed, two against the KRAS promoter G4mid and three others against putative G4FS in the distal promoter, intron 1 and exon 5. PPRH binding was confirmed by gel electrophoresis. The effect on KRAS transcription was examined by luciferase, FRET Melt2, qRT-PCR. Cytotoxicity was evaluated in pancreatic and ovarian cancer cells. PPRHs decreased activity of a luciferase construct driven by the KRAS promoter. PPRH selectively suppressed proliferation in KRAS dependent cancer cells. PPRH demonstrated synergistic activity with a KRAS promoter selective G4-stabilizing compound, NSC 317605, in KRAS-dependent pancreatic cells. PPRHs selectively stabilize G4 formation within the KRAS mid promoter region and represent an innovative approach to both G4-stabilization and to KRAS modulation with potential for development into novel therapeutics.

ZC3HAV1 promotes the proliferation and metastasis via regulating KRAS in pancreatic cancer.

Proliferation and metastasis are important malignant features of pancreatic cancer (PC), but the underlying molecular mechanism is unclear. ZC3HAV1, a PARP family member of proteins-enzymes, has been considered to play a significant part in a variety of biological processes. Nonetheless, the functions of ZC3HAV1 in developing PC are still unknown. This research aims to explore the biological function and the expression of ZC3HAV1 shown in PC. In our study, PCR analysis suggested that ZC3HAV1 was expressed at a high level in PC tissues and cell lines, and high ZC3HAV1 expression was remarkably related to poor prognosis. The functional assays indicated that upregulated ZC3HAV1 accelerated PC cell proliferation along with colony formation capacities in vitro. Subsequently, ZC3HAV1 could upregulate cyclin D1 and CDK2 and also promote G1/S transition in cells of PC. What’s more, we also discovered that ZC3HAV1 promotes the migration and the invasion of PC cells. It upregulates the expression of EMT (epithelial-mesenchymal transition) relevant markers. Conversely, the functional assays showed that ZC3HAV1 knockdown significantly reduced tumorigenesis. Using bioinformatics analysis and immunoprecipitation assays we found that ZC3HAV1 could directly bind to KRAS and positively regulate its expression. Furthermore, ZC3HAV1 overexpression activated MAPK signaling by increasing p-ERK levels. Conversely, knockdown of KRAS attenuated ZC3HAV1-mediated promotion of proliferation and invasion in cells of PC. The result indicated that ZC3HAV1 was in relation to poor prognosis and accelerated the proliferation and metastasis of PC cells by regulation of KRAS. Our research may offer brand-new evidence to diagnose and treat PC in clinic.

MO18-6 The tumor suppression potential of RNA pseudouridine synthase family via microRNA regulation

MicroRNAs are non-coding RNAs consisting of about 22 nucleotides that regulate a large number of mRNAs. Among them, let-7 is one of the first microRNAs to be identified and has tumor suppressive potential through regulation of KRAS and MYC mRNAs. We have shown that TruB1, one of the tRNA pseudouridine synthetases, regulates let-7 relatively specifically by cell-based screening (Kurimoto et al. EMBO J. 2020). In the present study, to evaluate the specificity of the let-7-mediated tumor suppressor ability of TruB1, we examined the effects of whole RNA pseudouridine synthetase on the regulation of its let-7 in vitro experiments and in the reported literatures. The results showed that DKC1 and PUS10 also regulate microRNAs of different sub-subset from let-7 in the reported literatures, whereas TruB2 may regulate microRNA subset including let-7. We will discuss about clinical impact of TruB2 on several cancers by TCGA database. These new functions of RNA-pseudouridine enzymes might have the impact on the tumor-suppressive potential. These results are significant from a molecular biological point of view, leading to a clarification of the regulatory mechanism.

Dynamic Regulation of Expression of KRAS and Its Effectors Determines the Ability to Initiate Tumorigenesis in Pancreatic Acinar Cells.

Pancreatic acinar cells are a cell type of origin for pancreatic cancer that become progressively less sensitive to tumorigenesis induced by oncogenic Kras mutations after birth. This sensitivity is increased when Kras mutations are combined with pancreatitis. Molecular mechanisms underlying these observations are still largely unknown. To identify these mechanisms, we generated the first CRISPR-edited mouse models that enable detection of wild-type and mutant KRAS proteins in vivo. Analysis of these mouse models revealed that more than 75% of adult acinar cells are devoid of detectable KRAS protein. In the 25% of acinar cells expressing KRAS protein, transcriptomic analysis highlighted a slight upregulation of the RAS and MAPK pathways. However, at the protein level, only marginal pancreatic expression of essential KRAS effectors, including C-RAF, was observed. The expression of KRAS and its effectors gradually decreased after birth. The low sensitivity of adult acinar cells to Kras mutations resulted from low expression of KRAS and its effectors and the subsequent lack of activation of RAS/MAPK pathways. Pancreatitis triggered expression of KRAS and its effectors as well as subsequent activation of downstream signaling; this induction required the activity of EGFR. Finally, expression of C-RAF in adult pancreas was required for pancreatic tumorigenesis. In conclusion, our study reveals that control of the expression of KRAS and its effectors regulates the sensitivity of acinar cells to transformation by oncogenic Kras mutations. SIGNIFICANCE: This study generates new mouse models to study regulation of KRAS during pancreatic tumorigenesis and highlights a novel mechanism through which pancreatitis sensitizes acinar cells to Kras mutations.

Discoveries in the redox regulation of KRAS.

Oncogenic KRAS is one of the most common drivers of human cancer. Despite intense research, no effective therapy to directly inhibit oncogenic KRAS has yet been approved and KRAS mutant tumors remain associated with a poor prognosis. This short review discusses the current knowledge of the redox regulation of RAS and examines the newest findings on cysteine 118 (C118) as a potential novel target for KRAS inhibition.

PTPN2 regulates the activation of KRAS and plays a critical role in proliferation and survival of KRAS-driven cancer cells

RAS genes are the most commonly mutated in human cancers and play critical roles in tumor initiation, progression, and drug resistance. Identification of targets that block RAS signaling is pivotal to develop therapies for RAS-related cancer. As RAS translocation to the plasma membrane (PM) is essential for its effective signal transduction, we devised a high-content screening assay to search for genes regulating KRAS membrane association. We found that the tyrosine phosphatase PTPN2 regulates the plasma membrane localization of KRAS. Knockdown of PTPN2 reduced the proliferation and promoted apoptosis in KRAS-dependent cancer cells, but not in KRAS-independent cells. Mechanistically, PTPN2 negatively regulates tyrosine phosphorylation of KRAS, which, in turn, affects the activation KRAS and its downstream signaling. Consistently, analysis of the TCGA database demonstrates that high expression of PTPN2 is significantly associated with poor prognosis of patients with KRAS-mutant pancreatic adenocarcinoma. These results indicate that PTPN2 is a key regulator of KRAS and may serve as a new target for therapy of KRAS-driven cancer.