Subject(s)
Gastric Fundus , Hyperplasia , Stomach Neoplasms , Humans , Stomach Neoplasms/pathology , Stomach Neoplasms/metabolism , Gastric Fundus/pathology , Gastric Fundus/metabolism , Cell Transformation, Neoplastic , Mucin 5AC/metabolism , Mucin-6/metabolism , Adenocarcinoma/pathology , Adenocarcinoma/metabolism , Adenocarcinoma/genetics , Adenocarcinoma/diagnosis , Proto-Oncogene Proteins p21(ras)/genetics , Precancerous Conditions/pathology , Precancerous Conditions/metabolism , Male , Gastric Mucosa/pathology , Mutation , Middle Aged , Tumor Suppressor Protein p53/metabolism , Tumor Suppressor Protein p53/genetics , FemaleABSTRACT
The role of KRAS mutation in non-small cell lung cancer (NSCLC) initiation and progression is well-established. However, "undruggable" KRAS protein poses the research of small molecule inhibitors a significant challenge. Addressing this, proteolysis-targeting chimeras (PROTACs) have become a cutting-edge treatment method, emphasizing protein degradation. A modified ethanol injection method was employed in this study to formulate liposomes encapsulating PROTAC drug LC-2 (LC-2 LPs). Precise surface modifications using cell-penetrating peptide R8 yielded R8-LC-2 liposomes (R8-LC-2 LPs). Comprehensive cellular uptake and cytotoxicity studies unveiled that R8-LC-2 LPs depended on concentration and time, showcasing the superior performance of R8-LC-2 LPs compared to normal liposomes. In vivo pharmacokinetic profiles demonstrated the capacity of DSPE-PEG2000 to prolong the circulation time of LC-2, leading to higher plasma concentrations compared to free LC-2. In vivo antitumor efficacy research underscored the remarkable ability of R8-LC-2 LPs to effectively suppress tumor growth. This study contributed to the exploration of enhanced therapeutic strategies for NSCLC, specifically focusing on the development of liposomal PROTACs targeting the "undruggable" KRAS protein. The findings provide valuable insights into the potential of this innovative approach, offering prospects for improved drug delivery and heightened antitumor efficacy.
Subject(s)
Carcinoma, Non-Small-Cell Lung , Liposomes , Lung Neoplasms , Proteolysis , Proto-Oncogene Proteins p21(ras) , Animals , Humans , Mice , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/pharmacokinetics , Antineoplastic Agents/therapeutic use , Carcinoma, Non-Small-Cell Lung/drug therapy , Cell Line, Tumor , Cell-Penetrating Peptides , Lung Neoplasms/drug therapy , Lung Neoplasms/metabolism , Phosphatidylethanolamines/chemistry , Polyethylene Glycols/chemistry , Proteolysis/drug effects , Proteolysis Targeting Chimera/administration & dosage , Proteolysis Targeting Chimera/pharmacokinetics , Proteolysis Targeting Chimera/therapeutic use , Proto-Oncogene Proteins p21(ras)/genetics , RatsABSTRACT
Kirsten Rat Sarcoma (KRAS) is the most commonly mutated oncogene in colorectal carcinoma (CRC). We have previously reported the interactions between microsatellite instability (MSI), DNA promoter methylation, and gene expression. In this study, we looked for associations between KRAS mutation, gene expression, and methylation that may help with precision medicine. Genome-wide gene expression and DNA methylation were done in paired CRC tumor and surrounding healthy tissues. The results suggested that (a) the magnitude of dysregulation of many major gene pathways in CRC was significantly greater in patients with the KRAS mutation, (b) the up- and down-regulation of these dysregulated gene pathways could be correlated with the corresponding hypo- and hyper-methylation, and (c) the up-regulation of CDKN2A was more pronounced in tumors with the KRAS mutation. A recent cell line study showed that there were higher CDKN2A levels in 5-FU-resistant CRC cells and that these could be down-regulated by Villosol. Our findings suggest the possibility of a better response to anti-CDKN2A therapy with Villosol in KRAS-mutant CRC. Also, the more marked up-regulation of genes in the proteasome pathway in CRC tissue, especially with the KRAS mutation and MSI, may suggest a potential role of a proteasome inhibitor (bortezomib, carfilzomib, or ixazomib) in selected CRC patients if necessary.
Subject(s)
Colorectal Neoplasms , DNA Methylation , Gene Expression Regulation, Neoplastic , Mutation , Proto-Oncogene Proteins p21(ras) , Transcriptome , Humans , Colorectal Neoplasms/genetics , Colorectal Neoplasms/drug therapy , Colorectal Neoplasms/pathology , Colorectal Neoplasms/metabolism , Proto-Oncogene Proteins p21(ras)/genetics , Gene Expression Regulation, Neoplastic/drug effects , Male , Female , Middle Aged , Aged , Gene Expression Profiling , Microsatellite Instability , Epigenome , Cyclin-Dependent Kinase Inhibitor p16/genetics , Cyclin-Dependent Kinase Inhibitor p16/metabolismABSTRACT
RAS GTPases associate with the biological membrane where they function as molecular switches to regulate cell growth. Recent studies indicate that RAS proteins oligomerize on membranes, and disrupting these assemblies represents an alternative therapeutic strategy. However, conflicting reports on RAS assemblies, ranging in size from dimers to nanoclusters, have brought to the fore key questions regarding the stoichiometry and parameters that influence oligomerization. Here, we probe three isoforms of RAS [Kirsten Rat Sarcoma viral oncogene (KRAS), Harvey Rat Sarcoma viral oncogene (HRAS), and Neuroblastoma oncogene (NRAS)] directly from membranes using mass spectrometry. We show that KRAS on membranes in the inactive state (GDP-bound) is monomeric but forms dimers in the active state (GTP-bound). We demonstrate that the small molecule BI2852 can induce dimerization of KRAS, whereas the binding of effector proteins disrupts dimerization. We also show that RAS dimerization is dependent on lipid composition and reveal that oligomerization of NRAS is regulated by palmitoylation. By monitoring the intrinsic GTPase activity of RAS, we capture the emergence of a dimer containing either mixed nucleotides or GDP on membranes. We find that the interaction of RAS with the catalytic domain of Son of Sevenless (SOScat) is influenced by membrane composition. We also capture the activation and monomer to dimer conversion of KRAS by SOScat. These results not only reveal the stoichiometry of RAS assemblies on membranes but also uncover the impact of critical factors on oligomerization, encompassing regulation by nucleotides, lipids, and palmitoylation.
Subject(s)
Cell Membrane , Protein Multimerization , Proto-Oncogene Proteins p21(ras) , Proto-Oncogene Proteins p21(ras)/metabolism , Proto-Oncogene Proteins p21(ras)/genetics , Proto-Oncogene Proteins p21(ras)/chemistry , Humans , Cell Membrane/metabolism , Membrane Proteins/metabolism , Membrane Proteins/chemistry , Membrane Proteins/genetics , GTP Phosphohydrolases/metabolism , GTP Phosphohydrolases/chemistry , GTP Phosphohydrolases/genetics , Lipoylation , ras Proteins/metabolism , ras Proteins/chemistry , Guanosine Triphosphate/metabolism , Guanosine Diphosphate/metabolismABSTRACT
Recent cancer genome analyses have identified frequently mutated genes that are responsible for the development and malignant progression of cancers, including colorectal cancer (CRC). We previously constructed mouse models that carried major driver mutations of CRC, namely Apc, Kras, Tgfbr2, Trp53, and Fbxw7, in combinations. Comprehensive histological analyses of the models showed a link between mutation combinations and malignant phenotypes, such as invasion, epithelial-mesenchymal transition (EMT), and metastasis. The major cause of cancer-related death is metastasis, making it important to understand the mechanism underlying metastasis in order to develop novel therapeutic strategies. To this end, we have established intestinal tumor-derived organoids from different genotyped mice and generated liver metastasis models via transplantation of the organoids into the spleen. Through histological and imaging analyses of the transplantation models, we have determined that the combination of Apc, Kras, Tgfbr2, and Trp53 mutations promotes liver metastasis at a high incidence. We also demonstrated polyclonal metastasis of tumor cell clusters consisting of genetically and phenotypically distinct cells through our model analysis. These organoid transplantation models recapitulate human CRC metastasis, constituting a useful tool for basic and clinical cancer research as a preclinical model. We herein report the experimental protocols of the organoid culture and generation of metastasis models.
Subject(s)
Liver Neoplasms , Mutation , Organoids , Animals , Organoids/pathology , Mice , Liver Neoplasms/secondary , Liver Neoplasms/genetics , Liver Neoplasms/pathology , Genotype , Disease Models, Animal , Tumor Suppressor Protein p53/genetics , Colorectal Neoplasms/pathology , Colorectal Neoplasms/genetics , Neoplasm Metastasis , Humans , Adenomatous Polyposis Coli Protein/genetics , Receptor, Transforming Growth Factor-beta Type II/genetics , Proto-Oncogene Proteins p21(ras)/genetics , Epithelial-Mesenchymal Transition/geneticsABSTRACT
BACKGROUND: Urinary bladder cancer, is the 10th most common global cancer, diagnosed in over 600,000 people causing 200,000 deaths annually. Artemisinin and its derivatives are safe compounds that have recently been proven to possess potent anti-tumor effects in vivo, through inhibition of cancer cell growth. The aim of this study is to assess the efficiency of artemisinin as a cancer treatment alone and as a pre-treatment fore cisplatin therapy for high grade urothelial carcinoma. METHODS: Sixty male albino mice were divided into six groups, and BBN was used to induce urinary bladder cancer. Blood samples were tested for renal functions and complete blood counts, kidney and urinary bladder tissues were harvested for histopathological examination. Total RNAs from urinary bladder tissues was collected, and gene expression of FGFR3, HRAS, P53, and KDM6A was quantified using qRT-PCR. RESULTS: Compared to the induced cancer group, the results revealed that FGFR3 expression levels were down-regulated in the induced cancer group treated by artemisinin only and the induced cancer group pre-treated with artemisinin prior to cisplatin by ~ 0.86-fold and 0.4-folds, respectively, aligning with HRAS down-regulation by ~ 9.54-fold and 9.05-fold, respectively. Whereas, P53 expression levels were up-regulated by ~ 0.68-fold and 0.84-fold, respectively, in parallel with KDM6A expression, which is up-regulated by ~ 0.95-folds and 5.27-folds, respectively. Also, serum creatinine and urea levels decreased significantly in the induced cancer group treated by artemisinin alone and the induced cancer group pre-treated with artemisinin prior to cisplatin, whereas the induced cancer group treated by cisplatin their levels increased significantly. Moreover, Hb, PLT, RBC, and WBC counts improved in both cancer groups treated by artemisinin alone and pre-treated with artemisinin prior to cisplatin. Histologically, in kidney tissues, artemisinin pre-treatment significantly reduced renal injury caused by cisplatin. While Artemisinin treatment for cancer in bladder tissues reverted invasive urothelial carcinoma to moderate urothelial dysplasia. CONCLUSIONS: This study indicates that artemisinin demonstrated a significant effect in reversal of the multi-step carcinogenesis process of high grade urothelial carcinoma and could enhance the effect of cisplatin therapy using artemisinin pre-treatment.
Subject(s)
Artemisinins , Cisplatin , Gene Expression Regulation, Neoplastic , Histone Demethylases , Receptor, Fibroblast Growth Factor, Type 3 , Tumor Suppressor Protein p53 , Urinary Bladder Neoplasms , Animals , Cisplatin/pharmacology , Cisplatin/therapeutic use , Male , Artemisinins/pharmacology , Artemisinins/therapeutic use , Receptor, Fibroblast Growth Factor, Type 3/genetics , Receptor, Fibroblast Growth Factor, Type 3/metabolism , Mice , Urinary Bladder Neoplasms/drug therapy , Urinary Bladder Neoplasms/genetics , Urinary Bladder Neoplasms/pathology , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Protein p53/metabolism , Gene Expression Regulation, Neoplastic/drug effects , Histone Demethylases/metabolism , Histone Demethylases/genetics , Proto-Oncogene Proteins p21(ras)/genetics , Humans , Disease Models, Animal , Antineoplastic Agents/pharmacology , Antineoplastic Agents/therapeutic useABSTRACT
AIM: This study aims to evaluate the prognostic role of the KRAS G12C mutation in patients with advanced non-small cell lung cancer and PD-L1 expression ≥50% who are treated with immune checkpoint inhibitor monotherapy. METHODS: We conducted a systematic review of clinical studies fulfilling the following criteria: (1) enrolling patients with advanced/metastatic non-small cell lung cancer with high PD-L1 tumour expression receiving first-line therapy with anti-PD-(L)1 immune checkpoint inhibitors; (2) comparing the outcomes of patients with the KRAS G12C mutation to those without this mutation, and (3) reporting overall survival and progression-free survival (PFS). The electronic databases Medline, EMBASE, Cochrane and Google Scholar, along with reference lists, were systematically searched. RESULTS: We identified four publications that fulfilled the inclusion criteria, comprising a total of 469 patients. Of these, two studies reported hazard ratios (HR) for PFS, resulting in a final pooled patient sample of 163 for the meta-analysis. In patients with non-small cell lung cancer who received anti-PD-(L)1 monotherapy, the presence of a KRAS G12C mutation was associated with improved PFS compared to patients with KRAS wild-type tumours, with a pooled hazard ratio of 0.39 and a 95% Confidence Interval (CI) of 0.25-0.63. Among all patients with KRAS mutations, those harbouring a KRAS G12C mutation had improved PFS compared to patients with any other KRAS mutation (pooled HR 0.33, 95% CI 0.19-0.57). CONCLUSIONS: Patients with non-small cell lung cancer who have the KRAS G12C mutation and high PD-L1 expression demonstrate favourable PFS with first-line PD-(L)1 immune checkpoint inhibitor monotherapy compared to patients with KRASwt or other KRAS mutations and high PD-L1 expression.
Subject(s)
B7-H1 Antigen , Carcinoma, Non-Small-Cell Lung , Immune Checkpoint Inhibitors , Immunotherapy , Lung Neoplasms , Proto-Oncogene Proteins p21(ras) , Carcinoma, Non-Small-Cell Lung/genetics , Carcinoma, Non-Small-Cell Lung/drug therapy , Humans , Lung Neoplasms/genetics , Lung Neoplasms/drug therapy , Proto-Oncogene Proteins p21(ras)/genetics , Immunotherapy/methods , Prognosis , Immune Checkpoint Inhibitors/therapeutic use , Mutation , Progression-Free SurvivalABSTRACT
BACKGROUND: There are known disparities in incidence and outcomes of colorectal cancer (CRC) by race and ethnicity. Some of these disparities may be mediated by molecular changes in tumors that occur at different rates across populations. Genetic ancestry is a measure complementary to race and ethnicity that can overcome missing data issues and better capture genetic similarity in admixed populations. We aimed to identify somatic mutations and tumor gene expression differences associated with both genetic ancestry and imputed race and ethnicity. METHODS: Sequencing was performed with the Tempus xT NGS 648-gene panel and whole exome capture RNA-Seq for 8454 primarily late-stage CRC patients. Genetic ancestry proportions for five continental groups-Africa (AFR), American indigenous (AMR), East Asia (EAS), Europe (EUR), and South Asia (SAS)-were estimated using ancestry informative markers. To address data gaps, race and ethnicity categories were imputed, resulting in assignments for 952 Hispanic/Latino, 420 non-Hispanic (NH) Asian, 1061 NH Black, and 5763 NH White individuals. We assessed association of genetic ancestry proportions and imputed race and ethnicity categories with somatic mutations in relevant CRC genes and in 2608 expression profiles, as well as 1957 consensus molecular subtypes (CMS). RESULTS: Increased AFR ancestry was associated with higher odds of somatic mutations in APC, KRAS, and PIK3CA and lower odds of BRAF mutations. Additionally, increased EAS ancestry was associated with lower odds of mutations in KRAS, EUR with higher odds in BRAF, and the Hispanic/Latino category with lower odds in BRAF. Greater AFR ancestry and the NH Black category were associated with higher rates of CMS3, while a higher proportion of Hispanic/Latino patients exhibited indeterminate CMS classifications. CONCLUSIONS: Molecular differences in CRC tumor mutation frequencies and gene expression that may underlie observed differences by race and ethnicity were identified. The association of AFR ancestry with increased KRAS mutations aligns with higher CMS3 subtype rates in NH Black patients. The increase of indeterminate CMS in Hispanic/Latino patients suggests that subtype classification methods could benefit from enhanced patient diversity.
Subject(s)
Colorectal Neoplasms , Mutation , Humans , Colorectal Neoplasms/genetics , Male , Female , Middle Aged , Proto-Oncogene Proteins B-raf/genetics , Aged , Class I Phosphatidylinositol 3-Kinases/genetics , Proto-Oncogene Proteins p21(ras)/genetics , Biomarkers, Tumor/genetics , Adenomatous Polyposis Coli Protein/geneticsSubject(s)
Carcinoma, Non-Small-Cell Lung , Circulating Tumor DNA , Lung Neoplasms , Mutation , Proto-Oncogene Proteins p21(ras) , Humans , Lung Neoplasms/genetics , Lung Neoplasms/drug therapy , Lung Neoplasms/pathology , Lung Neoplasms/blood , Carcinoma, Non-Small-Cell Lung/genetics , Carcinoma, Non-Small-Cell Lung/drug therapy , Carcinoma, Non-Small-Cell Lung/pathology , Carcinoma, Non-Small-Cell Lung/blood , Circulating Tumor DNA/genetics , Circulating Tumor DNA/blood , Proto-Oncogene Proteins p21(ras)/geneticsSubject(s)
Carcinoma, Non-Small-Cell Lung , Circulating Tumor DNA , Lung Neoplasms , Mutation , Proto-Oncogene Proteins p21(ras) , Humans , Lung Neoplasms/genetics , Lung Neoplasms/drug therapy , Lung Neoplasms/pathology , Lung Neoplasms/blood , Carcinoma, Non-Small-Cell Lung/genetics , Carcinoma, Non-Small-Cell Lung/drug therapy , Carcinoma, Non-Small-Cell Lung/pathology , Carcinoma, Non-Small-Cell Lung/blood , Circulating Tumor DNA/genetics , Circulating Tumor DNA/blood , Proto-Oncogene Proteins p21(ras)/geneticsABSTRACT
Objective: To investigate the role of an inflammatory microenvironment induced by Porphyromonasgingivalis (P. gingivalis) in the occurrence of esophageal squamous cell carcinoma (ESCC) in mice. Methods: A total of 180 C57BL/6 mice were randomly divided into 6 groups, i.e. control group, P. gingivalis group, 4NQO group, 4NQO + P. gingivalis group, 4NQO + P. gingivalis + celecoxib group, and 4NQO + P. gingivalis + antibiotic cocktail (ABC, including metronidazole, neomycin, ampicillin, and vancomycin) group, with 30 mice in each group, using the random number table. All mice were normalized by treatment with ABC in drinking water for 2 weeks. In the following 2 weeks, the mice in the control group and the P. gingivalis group were given drinking water, while the other 4 groups were treated with 30 µg/ml 4NQO in the drinking water. In weeks 11-12, the mice in the P. gingivalis group, the 4NQO + P. gingivalis group, the 4NQO + P. gingivalis + celecoxib group, and the 4NQO + P. gingivalis + ABC group were subjected to ligation of the second molar in oral cavity followed by oral P. gingivalis infection thrice weekly for 24 weeks in weeks 11-34. In weeks 13-34, the mice in 4NQO + P. gingivalis+celecoxib group and 4NQO + P. gingivalis + ABC group were administered with celecoxib and ABC for 22 weeks, respectively. At the end of 34 weeks, gross and microscopic alterations were examined followed by RT-qPCR and immunohistochemistry to examine the expression profiles of inflammatory- and tumor-molecules in esophagi of mice. Results: At 34 weeks, 4NQO treatment alone did not affect the foci of papillary hyperproliferation, diseased area, and the thickness of the esophageal wall, but significantly enhanced the foci of hyperproliferation (median 1.00, Pï¼0.05) and mild/moderate dysplasia (median 2.00, Pï¼0.01). In addition, the expression levels of IL-6 [8.35(3.45,8.99)], IL-1ß [6.90(2.01,9.72)], TNF-α [12.04(3.31,14.08)], c-myc [2.21(1.80,3.04)], pSTAT3, Ki-67, and pH2AX were higher than those in the control group. The pathological changes of the esophageal mucosa were significantly more overt in the 4NQO + P. gingivalis group in terms of the foci of papillary hyperproliferation (median 2.00), diseased area (median 2.51 mm2), the thickness of the esophageal wall (median 172.52 µm), the foci of hyperproliferation (median 1.00, Pï¼0.05), and mild/moderate dysplasia (median 1.00, Pï¼0.01). In mice of the 4NQO + P. gingivalis group, the expression levels of IL-6 [12.27(5.35,22.08)], IL-1ß [13.89(10.04,15.96)], TNF-α [19.56(6.07,20.36)], IFN-γ [11.37(8.23,20.07)], c-myc [2.62(1.51,4.25)], cyclin D1 [4.52(2.68,7.83)], nuclear pSTAT3, COX-2, Ki-67, and pH2AX were significantly increased compared with the mice in the control group. In mice of the 4NQO + P. gingivalis group, the diseased area, invasive malignant foci as well as pSTAT3 and pH2AX expression were significantly blunted by celecoxib. Treatment with ABC markedly reduced the papillary hyperproliferative foci, invasive malignant foci, and pSTAT3 expression but not pH2AX. Conclusions: P. gingivalis promotes the occurrence of esophageal squamous cell carcinoma in mice by inducing an inflammatory microenvironment primed with 4NQO induced DNA damage. Clearance of P. gingivalis with ABC or anti-inflammatory intervention holds promise for prevention of esophageal squamous cell malignant pathogenesis via blockage of IL-6/STAT3 signaling and amelioration of inflammation.
Subject(s)
4-Nitroquinoline-1-oxide , Celecoxib , Esophageal Neoplasms , Esophageal Squamous Cell Carcinoma , Mice, Inbred C57BL , Porphyromonas gingivalis , Tumor Microenvironment , Animals , Mice , Esophageal Neoplasms/microbiology , Esophageal Neoplasms/pathology , Esophageal Squamous Cell Carcinoma/microbiology , Esophageal Squamous Cell Carcinoma/metabolism , Esophageal Squamous Cell Carcinoma/pathology , Celecoxib/pharmacology , Inflammation , Bacteroidaceae Infections/microbiology , Interleukin-6/metabolism , Anti-Bacterial Agents/pharmacology , STAT3 Transcription Factor/metabolism , Cyclooxygenase 2/metabolism , Cyclooxygenase 2/genetics , Esophagus/microbiology , Esophagus/pathology , Esophagitis/microbiology , Esophagitis/pathology , Proto-Oncogene Proteins p21(ras)/genetics , Proto-Oncogene Proteins p21(ras)/metabolism , Carcinoma, Squamous Cell/microbiology , Carcinoma, Squamous Cell/pathology , Carcinoma, Squamous Cell/metabolismSubject(s)
Colorectal Neoplasms , Mutation , Proto-Oncogene Proteins p21(ras) , Humans , Colorectal Neoplasms/genetics , Colorectal Neoplasms/drug therapy , Proto-Oncogene Proteins p21(ras)/genetics , Pyrimidines/therapeutic use , Pyridines/therapeutic use , Antineoplastic Agents/therapeutic use , PiperazinesABSTRACT
BACKGROUND: KRAS mutations frequently occur in cancers, particularly pancreatic ductal adenocarcinoma, colorectal cancer, and non-small cell lung cancer. Although KRASG12C inhibitors have recently been approved, effective precision therapies have not yet been established for all KRAS-mutant cancers. Many treatments for KRAS-mutant cancers, including epigenome-targeted drugs, are currently under investigation. Small ubiquitin-like modifier (SUMO) proteins are a family of small proteins covalently attached to and detached from other proteins in cells via the processes called SUMOylation and de-SUMOylation. We assessed whether SUMOylation inhibition was effective in KRAS-mutant cancer cells. METHODS: The efficacy of the first-in-class SUMO-activating enzyme E inhibitor TAK-981 (subasumstat) was assessed in multiple human and mouse KRAS-mutated cancer cell lines. A gene expression assay using a TaqMan array was used to identify biomarkers of TAK-981 efficacy. The biological roles of SUMOylation inhibition and subsequent regulatory mechanisms were investigated using immunoblot analysis, immunofluorescence assays, and mouse models. RESULTS: We discovered that TAK-981 downregulated the expression of the currently undruggable MYC and effectively suppressed the growth of MYC-expressing KRAS-mutant cancers across different tissue types. Moreover, TAK-981-resistant cells were sensitized to SUMOylation inhibition via MYC-overexpression. TAK-981 induced proteasomal degradation of MYC by altering the balance between SUMOylation and ubiquitination and promoting the binding of MYC and Fbxw7, a key factor in the ubiquitin-proteasome system. The efficacy of TAK-981 monotherapy in immunocompetent and immunodeficient mouse models using a mouse-derived CMT167 cell line was significant but modest. Since MAPK inhibition of the KRAS downstream pathway is crucial in KRAS-mutant cancer, we expected that co-inhibition of SUMOylation and MEK might be a good option. Surprisingly, combination treatment with TAK-981 and trametinib dramatically induced apoptosis in multiple cell lines and gene-engineered mouse-derived organoids. Moreover, combination therapy resulted in long-term tumor regression in mouse models using cell lines of different tissue types. Finally, we revealed that combination therapy complementally inhibited Rad51 and BRCA1 and accumulated DNA damage. CONCLUSIONS: We found that MYC downregulation occurred via SUMOylation inhibition in KRAS-mutant cancer cells. Our findings indicate that dual inhibition of SUMOylation and MEK may be a promising treatment for MYC-expressing KRAS-mutant cancers by enhancing DNA damage accumulation.
Subject(s)
DNA Damage , Proto-Oncogene Proteins p21(ras) , Sumoylation , Sumoylation/drug effects , Animals , Mice , Humans , Cell Line, Tumor , DNA Damage/drug effects , Proto-Oncogene Proteins p21(ras)/genetics , Proto-Oncogene Proteins p21(ras)/metabolism , Mutation , Proto-Oncogene Proteins c-myc/metabolism , Proto-Oncogene Proteins c-myc/geneticsABSTRACT
Hyperactive Ras signalling is found in most cancers. Ras proteins are only active in membrane nanoclusters, which are therefore potential drug targets. We previously showed that the nanocluster scaffold galectin-1 (Gal1) enhances H-Ras nanoclustering via direct interaction with the Ras binding domain (RBD) of Raf. Here, we establish that the B-Raf preference of Gal1 emerges from the divergence of the Raf RBDs at their proposed Gal1-binding interface. We then identify the L5UR peptide, which disrupts this interaction by binding with low micromolar affinity to the B- and C-Raf-RBDs. Its 23-mer core fragment is sufficient to interfere with H-Ras nanoclustering, modulate Ras-signalling and moderately reduce cell viability. These latter two phenotypic effects may also emerge from the ability of L5UR to broadly engage with several RBD- and RA-domain containing Ras interactors. The L5UR-peptide core fragment is a starting point for the development of more specific reagents against Ras-nanoclustering and -interactors.
Subject(s)
Peptides , Humans , Peptides/metabolism , Peptides/chemistry , Proto-Oncogene Proteins p21(ras)/metabolism , Proto-Oncogene Proteins p21(ras)/genetics , Proto-Oncogene Proteins p21(ras)/chemistry , Galectin 1/metabolism , Galectin 1/chemistry , Galectin 1/genetics , Protein Binding , Signal TransductionABSTRACT
"NeoRAS WT" refers to the loss of RAS mutations (MTs) following first-line treatment in metastatic colorectal cancer (mCRC). We evaluate the incidence and clinicopathological characteristics of NeoRAS WT mCRC using next-generation sequencing of plasma circulating tumor DNA. Patients with mCRC enrolled in the GOZILA study initially diagnosed with tissue RAS MT mCRC and received subsequent systemic therapy are eligible. NeoRAS WT is defined as the absence of detectable RAS MT in plasma and assessed in all eligible patients (Group A) and in a subgroup with at least one somatic alteration detected in plasma (Group B). Overall, 478 patients are included. NeoRAS WT prevalence is 19.0% (91/478) in Group A and 9.8% (42/429) in Group B. Absence of liver or lymph node metastasis and tissue RAS MTs other than KRAS exon 2 MTs are significantly associated with NeoRAS WT emergence. Overall, 1/6 and 2/6 patients with NeoRAS WT treated with anti-EGFR monoclonal antibodies (mAbs) show partial response and stable disease for ≥6 months, respectively. NeoRAS WT mCRC is observed at a meaningful prevalence, and anti-EGFR mAb-based therapy may be effective.
Subject(s)
Colorectal Neoplasms , Mutation , Humans , Colorectal Neoplasms/genetics , Colorectal Neoplasms/pathology , Male , Female , Middle Aged , Aged , Japan/epidemiology , ErbB Receptors/genetics , ErbB Receptors/antagonists & inhibitors , Circulating Tumor DNA/genetics , Circulating Tumor DNA/blood , Adult , High-Throughput Nucleotide Sequencing , Proto-Oncogene Proteins p21(ras)/genetics , Neoplasm Metastasis , Aged, 80 and over , Liver Neoplasms/secondary , Liver Neoplasms/genetics , Antibodies, Monoclonal/therapeutic use , Lymphatic MetastasisABSTRACT
Single-agent TAS102 (trifluridine/tipiracil) and regorafenib are FDA-approved treatments for metastatic colorectal cancer (mCRC). We previously reported that regorafenib combined with a fluoropyrimidine can delay disease progression in clinical case reports of multidrug-resistant mCRC patients. We hypothesized that the combination of TAS102 and regorafenib may be active in CRC and other gastrointestinal (GI) cancers and may in the future provide a treatment option for patients with advanced GI cancer. We investigated the therapeutic effect of TAS102 in combination with regorafenib in preclinical studies employing cell culture, colonosphere assays that enrich for cancer stem cells, and in vivo. TAS102 in combination with regorafenib has synergistic activity against multiple GI cancers in vitro including colorectal and gastric cancer, but not liver cancer cells. TAS102 inhibits colonosphere formation and this effect is potentiated by regorafenib. In vivo anti-tumor effects of TAS102 plus regorafenib appear to be due to anti-proliferative effects, necrosis and angiogenesis inhibition. Growth inhibition by TAS102 plus regorafenib occurs in xenografted tumors regardless of p53, KRAS or BRAF mutations, although more potent tumor suppression was observed with wild-type p53. Regorafenib significantly inhibits TAS102-induced angiogenesis and microvessel density in xenografted tumors, as well inhibits TAS102-induced ERK1/2 activation regardless of RAS or BRAF status in vivo. TAS102 plus regorafenib is a synergistic drug combination in preclinical models of GI cancer, with regorafenib suppressing TAS102-induced increase in microvessel density and p-ERK as contributing mechanisms. The TAS102 plus regorafenib drug combination may be further tested in gastric and other GI cancers.
Subject(s)
Drug Combinations , Drug Synergism , Gastrointestinal Neoplasms , Mutation , Neoplastic Stem Cells , Neovascularization, Pathologic , Phenylurea Compounds , Proto-Oncogene Proteins B-raf , Proto-Oncogene Proteins p21(ras) , Pyridines , Pyrrolidines , STAT3 Transcription Factor , Thymine , Trifluridine , Uracil , Xenograft Model Antitumor Assays , Humans , Trifluridine/pharmacology , Phenylurea Compounds/pharmacology , Animals , Pyridines/pharmacology , Proto-Oncogene Proteins B-raf/genetics , Proto-Oncogene Proteins B-raf/metabolism , Proto-Oncogene Proteins p21(ras)/genetics , Proto-Oncogene Proteins p21(ras)/metabolism , Neovascularization, Pathologic/drug therapy , Neovascularization, Pathologic/genetics , Neovascularization, Pathologic/metabolism , Neoplastic Stem Cells/drug effects , Neoplastic Stem Cells/metabolism , Neoplastic Stem Cells/pathology , Gastrointestinal Neoplasms/drug therapy , Gastrointestinal Neoplasms/genetics , Gastrointestinal Neoplasms/pathology , Gastrointestinal Neoplasms/metabolism , Uracil/pharmacology , Uracil/analogs & derivatives , Mice , STAT3 Transcription Factor/metabolism , STAT3 Transcription Factor/genetics , Thymine/pharmacology , Cell Line, Tumor , Pyrrolidines/pharmacology , Pyrrolidines/therapeutic use , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , MAP Kinase Signaling System/drug effects , Signal Transduction/drug effects , Cell Proliferation/drug effects , AngiogenesisABSTRACT
Importance: Among patients with metastatic colorectal cancer (mCRC), data are limited on disparate biomarker testing and its association with clinical outcomes on a national scale. Objective: To evaluate the socioeconomic and demographic inequities in microsatellite instability (MSI) and KRAS biomarker testing among patients with mCRC and to explore the association of testing with overall survival (OS). Design, Setting, and Participants: This cohort study, conducted between November 2022 and March 2024, included patients who were diagnosed with mCRC between January 1, 2010, and December 31, 2017. The study obtained data from the National Cancer Database, a hospital-based cancer registry in the US. Patients with mCRC and available information on biomarker testing were included. Patients were classified based on whether they completed or did not complete MSI or KRAS tests. Exposure: Demographic and socioeconomic factors, such as age, race, ethnicity, educational level in area of residence, median household income, insurance type, area of residence, facility type, and facility location were evaluated. Main Outcomes and Measures: The main outcomes were MSI and KRAS testing between the date of diagnosis and the date of first-course therapy. Univariable and multivariable logistic regressions were used to identify the relevant factors in MSI and KRAS testing. The OS outcomes were also evaluated. Results: Among the 41â¯061 patients included (22 362 males [54.5%]; mean [SD] age, 62.3 [10.1] years; 17.3% identified as Black individuals, 78.0% as White individuals, 4.7% as individuals of other race, with 6.5% Hispanic or 93.5% non-Hispanic ethnicity), 28.8% underwent KRAS testing and 43.7% received MSI testing. A significant proportion of patients had Medicare insurance (43.6%), received treatment at a comprehensive community cancer program (40.5%), and lived in an area with lower educational level (51.3%). Factors associated with a lower likelihood of MSI testing included age of 70 to 79 years (relative risk [RR], 0.70; 95% CI, 0.66-0.74; P < .001), treatment at a community cancer program (RR, 0.74; 95% CI, 0.70-0.79; P < .001), rural residency (RR, 0.80; 95% CI, 0.69-0.92; P < .001), lower educational level in area of residence (RR, 0.84; 95% CI, 0.79-0.89; P < .001), and treatment at East South Central facilities (RR, 0.67; 95% CI, 0.61-0.73; P < .001). Similar patterns were observed for KRAS testing. Survival analysis showed modest OS improvement in patients with MSI testing (hazard ratio, 0.93; 95% CI, 0.91-0.96; P < .001). The median (IQR) follow-up time for the survival analysis was 13.96 (3.71-29.34) months. Conclusions and Relevance: This cohort study of patients with mCRC found that older age, community-setting treatment, lower educational level in area of residence, and treatment at East South Central facilities were associated with a reduced likelihood of MSI and KRAS testing. Highlighting the sociodemographic-based disparities in biomarker testing can inform the development of strategies that promote equity in cancer care and improve outcomes for underserved populations.
Subject(s)
Biomarkers, Tumor , Colorectal Neoplasms , Healthcare Disparities , Microsatellite Instability , Proto-Oncogene Proteins p21(ras) , Humans , Colorectal Neoplasms/genetics , Colorectal Neoplasms/mortality , Colorectal Neoplasms/pathology , Male , Female , Middle Aged , Aged , Healthcare Disparities/statistics & numerical data , Proto-Oncogene Proteins p21(ras)/genetics , United States , Cohort Studies , Socioeconomic Factors , Neoplasm MetastasisABSTRACT
BACKGROUND: This study aimed to identify the associations between individual KRAS, STK11, KEAP1, or TP53 mutations, as well as the comutation status of these genes, and the tumor mutation burden (TMB) with clinical outcomes of lung adenocarcinoma patients treated with immune checkpoint inhibitors (ICIs). METHODS: We collected data from patients with lung adenocarcinoma treated with ICIs from the Center for Cancer Genomics and Advanced Therapeutics (C-CAT) database between June 2019 and August 2023. The main endpoints were the treatment response and overall survival (OS). RESULTS: Among 343 patients with lung adenocarcinoma, 61 (18%), 69 (20%), 41 (12%), and 222 (65%) patients had KRAS, STK11, KEAP1, and TP53 mutations, respectively. An overall objective response was observed in 94 of 338 patients (28%), including 2 (1%) who achieved a complete response and 92 (27%) who achieved a partial response. Patients with STK11, KEAP1, or TP53 mutations had a significantly greater TMB (P<0.001). According to the univariate analysis, the treatment response was significantly correlated with TP53 mutation in both the general (P = 0.041) and KRAS wild-type (P = 0.009) populations. KEAP1 and TP53 mutations were associated with worse OS among assessable patients (hazard ratio (HR) = 2.027, P = 0.002; HR = 1.673, P = 0.007, respectively) and among patients without KRAS mutations (HR = 1.897, P = 0.012; HR = 1.908, P = 0.004, respectively). According to the multivariate analysis, KEAP1 (HR = 1.890, P = 0.008) and TP53 (HR = 1.735, P = 0.011) mutations were found to be independent factors for OS. CONCLUSIONS: STK11, KEAP1, and TP53 mutations are significantly associated with a high TMB. TP53 mutation could affect the treatment response to some degree, and both KEAP1 and TP53 mutations resulted in inferior OS in the general patient population and in those with KRAS-wild-type lung adenocarcinoma, indicating that KEAP1 and TP53 mutations might act as prognostic factors for ICI treatment in lung adenocarcinoma patients.
Subject(s)
AMP-Activated Protein Kinase Kinases , Adenocarcinoma of Lung , Immune Checkpoint Inhibitors , Kelch-Like ECH-Associated Protein 1 , Lung Neoplasms , Mutation , Protein Serine-Threonine Kinases , Proto-Oncogene Proteins p21(ras) , Tumor Suppressor Protein p53 , Humans , Kelch-Like ECH-Associated Protein 1/genetics , Male , Female , Proto-Oncogene Proteins p21(ras)/genetics , Tumor Suppressor Protein p53/genetics , Adenocarcinoma of Lung/genetics , Adenocarcinoma of Lung/drug therapy , Adenocarcinoma of Lung/pathology , Adenocarcinoma of Lung/mortality , Aged , Middle Aged , Protein Serine-Threonine Kinases/genetics , Lung Neoplasms/genetics , Lung Neoplasms/drug therapy , Lung Neoplasms/mortality , Immune Checkpoint Inhibitors/therapeutic use , Adult , Aged, 80 and over , Treatment OutcomeABSTRACT
Current KRASG12C (OFF) inhibitors that target inactive GDP-bound KRASG12C cause responses in less than half of patients and these responses are not durable. A class of RASG12C (ON) inhibitors that targets active GTP-bound KRASG12C blocks ERK signaling more potently than the inactive-state inhibitors. Sensitivity to either class of agents is strongly correlated with inhibition of mTORC1 activity. We have previously shown that PI3K/mTOR and ERK-signaling pathways converge on key cellular processes and that inhibition of both pathways is required for inhibition of these processes and for significant antitumor activity. We find here that the combination of a KRASG12C inhibitor with a selective mTORC1 kinase inhibitor causes synergistic inhibition of Cyclin D1 expression and cap-dependent translation. Moreover, BIM upregulation by KRASG12C inhibition and inhibition of MCL-1 expression by the mTORC1 inhibitor are both required to induce significant cell death. In vivo, this combination causes deep, durable tumor regressions and is well tolerated. This study suggests that the ERK and PI3K/mTOR pathways each mitigate the effects of inhibition of the other and that combinatorial inhibition is a potential strategy for treating KRASG12C-dependent lung cancer.