Unsupervised Feature Selection by a Genetic Algorithm for Mid-Infrared Spectral Data.

Dimensional reduction of highly multidimensional datasets such as those acquired by Fourier transform infrared spectroscopy (FTIR) is a critical step in the data analysis workflow. To achieve this goal, numerous feature selection methods have been developed and applied in a supervised context, i.e., using a priori knowledge about data usually in the form of labels for classification or quantitative values for regression. For this, genetic algorithms have been largely exploited due to their flexibility and global optimization principle. However, few applications in an unsupervised context have been reported in infrared spectroscopy. The aim of this article is to propose a new unsupervised feature selection method based on a genetic algorithm using a validity index computed from KMeans partitions as a fitness function. Evaluated on a simulated dataset and validated and tested on three real-world infrared spectroscopic datasets, our developed algorithm is able to find the spectral descriptors improving clustering accuracy and simplifying the spectral interpretation of results.

Colorectal Cancer: The Contribution of CXCL12 and Its Receptors CXCR4 and CXCR7.

Colorectal cancer is one of the most common cancers, and diagnosis at late metastatic stages is the main cause of death related to this cancer. This progression to metastasis is complex and involves different molecules such as the chemokine CXCL12 and its two receptors CXCR4 and CXCR7. The high expression of receptors in CRC is often associated with a poor prognosis and aggressiveness of the tumor. The interaction of CXCL12 and its receptors activates signaling pathways that induce chemotaxis, proliferation, migration, and cell invasion. To this end, receptor inhibitors were developed, and their use in preclinical and clinical studies is ongoing. This review provides an overview of studies involving CXCR4 and CXCR7 in CRC with an update on their targeting in anti-cancer therapies.

Automatic Identification of Paraffin Pixels on FTIR Images Acquired on FFPE Human Samples.

The transfer of mid-infrared spectral histopathology to the clinic will be possible provided that its application in clinical practice is simple. Rapid analysis of formalin-fixed paraffin-embedded (FFPE) tissue section is thus a prerequisite. The chemical dewaxing of these samples before image acquisition used by the majority of studies is in contradiction with this principle. Fortunately, the in silico analysis of the images acquired on FFPE samples is possible using extended multiplicative signal correction (EMSC). However, the removal of pure paraffin pixels is essential to perform a relevant classification of tissue spectra. So far, this task was possible only if using manual and subjective histogram analysis. In this article, we thus propose a new automatic and multivariate methodology based on the analysis of optimized combinations of EMSC regression coefficients by validity indices and KMeans clustering to separate paraffin and tissue pixels. The validation of our method is performed using simulated infrared spectral images by measuring the Jaccard index between our partitions and the image model, with values always over 0.90 for diverse baseline complexity and signal-to-noise ratio. These encouraging results were also validated on real images by comparing our method with classical ones and by computing the Jaccard index between our partitions and the KMeans partitions obtained on the infrared image acquired on the same samples but after chemical dewaxing, with values always over 0.84.

Impact of pemetrexed chemotherapy on the gut microbiota and intestinal inflammation of patient-lung-derived tumor xenograft (PDX) mouse models.

Chemotherapy remains the gold standard for advanced cancer. Pemetrexed, a chemotherapeutic agent used in non-small cell lung cancer, can induce significant side effects in patients. Although microbiota's role in the efficacy and/or toxicity of chemotherapy agents has been demonstrated, the impacts of pemetrexed on the gut microbiota and on gastrointestinal inflammation remain unknown. The objective of this study was to evaluate the impact of pemetrexed and the tumor graft on the gut microbiota composition in immunodeficient mice. The faecal microbiota composition was studied with metabarcoding before, 24-h and one week after treatment. The colon epithelial barrier integrity was evaluated by histological examination, intestinal permeability measurement, and selected cytokines quantification. The tumor graft induced some variations in the microbiota composition. Pemetrexed further increased the relative abundance of Enterobacteriaceae and 3 families from the Firmicutes phylum: Enterococcaceae, Lactobacillaceae and Streptococcaceae. Pemetrexed also significantly altered the epithelial barrier integrity, which was associated with early inflammation. This pilot study shows that the association of a lung tumor graft with pemetrexed causes an alteration in the microbiota composition. Such information increases our knowledge about the impact of chemotherapy on the microbiota, which could help to minimize side effects and improve therapeutic effectiveness in the future.

Hypoxic Environment and Paired Hierarchical 3D and 2D Models of Pediatric H3.3-Mutated Gliomas Recreate the Patient Tumor Complexity.

BACKGROUND: Pediatric high-grade gliomas (pHGGs) are facing a very dismal prognosis and representative pre-clinical models are needed for new treatment strategies. Here, we examined the relevance of collecting functional, genomic, and metabolomics data to validate patient-derived models in a hypoxic microenvironment. METHODS: From our biobank of pediatric brain tumor-derived models, we selected 11 pHGGs driven by the histone H3.3K28M mutation. We compared the features of four patient tumors to their paired cell lines and mouse xenografts using NGS (next generation sequencing), aCGH (array comparative genomic hybridization), RNA sequencing, WES (whole exome sequencing), immunocytochemistry, and HRMAS (high resolution magic angle spinning) spectroscopy. We developed a multicellular in vitro model of cell migration to mimic the brain hypoxic microenvironment. The live cell technology Incucyte© was used to assess drug responsiveness in variable oxygen conditions. RESULTS: The concurrent 2D and 3D cultures generated from the same tumor sample exhibited divergent but complementary features, recreating the patient intra-tumor complexity. Genomic and metabolomic data described the metabolic changes during pHGG progression and supported hypoxia as an important key to preserve the tumor metabolism in vitro and cell dissemination present in patients. The neurosphere features preserved tumor development and sensitivity to treatment. CONCLUSION: We proposed a novel multistep work for the development and validation of patient-derived models, considering the immature and differentiated content and the tumor microenvironment of pHGGs.

Synergistic Anti-Tumor Effect of mTOR Inhibitors with Irinotecan on Colon Cancer Cells.

Advanced colorectal cancer has a poor prognosis because of metastasis formation and resistance to combined therapies. Downstream of PI3K/Akt and Ras/MAPK pathways, the mTOR kinase plays a decisive role in treatment failure. We previously established that irinotecan has antiangiogenic properties and it is known that new mammalian target of rapamycin (mTOR) catalytic AZD inhibitors, unlike rapamycin, target both mTORC1 and mTORC2. Thus, we hypothesized that the complete inhibition of the PI3K/AKT/mTOR/HIF-1α axis with mTOR catalytic inhibitors and low doses of irinotecan may have antitumor effects. We showed that the AZD8055 and AZD2014 inhibitors were much more potent than rapamycin to reduce cell viability of four colon cell lines. On the other hand, whereas AZD2014 alone inhibits migration by 40%, the drug combination led to 70% inhibition. Similarly, neither irinotecan nor AZD2014 significantly reduced cell invasion, whereas a combination of the two inhibits invasion by 70%. In vivo, irinotecan and AZD2014 combination drastically reduced ectopic patient-derived colon tumor growth and this combination was more potent than Folfox or Folfiri. Finally, the combination totally inhibited liver and lung metastases developed from orthotopic implantation of SW480 cells. Thus, the use of mTOR catalytic inhibitors, in association with other chemotherapeutic agents like irinotecan at low doses, is potentially a hope for colon cancer treatment.

RNA Aptamers Targeting Integrin α5ß1 as Probes for Cyto- and Histofluorescence in Glioblastoma.

Nucleic acid aptamers are often referred to as chemical antibodies. Because they possess several advantages, like their smaller size, temperature stability, ease of chemical modification, lack of immunogenicity and toxicity, and lower cost of production, aptamers are promising tools for clinical applications. Aptamers against cell surface protein biomarkers are of particular interest for cancer diagnosis and targeted therapy. In this study, we identified and characterized RNA aptamers targeting cells expressing integrin α5ß1. This αß heterodimeric cell surface receptor is implicated in tumor angiogenesis and solid tumor aggressiveness. In glioblastoma, integrin α5ß1 expression is associated with an aggressive phenotype and a decrease in patient survival. We used a complex and original hybrid SELEX (selective evolution of ligands by exponential enrichment) strategy combining protein-SELEX cycles on the recombinant α5ß1 protein, surrounded by cell-SELEX cycles using two different cell lines. We identified aptamer H02, able to differentiate, in cyto- and histofluorescence assays, glioblastoma cell lines, and tissues from patient-derived tumor xenografts according to their α5 expression levels. Aptamer H02 is therefore an interesting tool for glioblastoma tumor characterization.

A redox ruthenium compound directly targets PHD2 and inhibits the HIF1 pathway to reduce tumor angiogenesis independently of p53.

Targeting specific tumor metabolic needs represents an actively investigated therapeutic strategy to bypass tumor resistance mechanisms. In this study, we describe an original approach to impact the cancer metabolism by exploiting the redox properties of a ruthenium organometallic compound. This organometallic complex induced p53-independent cytotoxicity and reduced size and vascularization of patients-derived tumor explants that are resistant to platinum drugs. At the molecular level, the ruthenium complex altered redox enzyme activities and the intracellular redox state by increasing the NAD+/NADH ratio and ROS levels. Pathway analysis pointed to HIF-1 as a top deregulated metabolite pathway. Unlike cisplatin, treatment with the ruthenium complex decreased HIF1A protein levels and expression of HIF1A target genes. The rapid downregulation of HIF1A protein levels involved a direct interaction of the ruthenium compound with the redox enzyme PHD2, a HIF1A master regulator. HIF1A inhibition led to decreased angiogenesis in patient-derived xenografted using fragments of primary human colon tumors. Altogether, our results show that a ruthenium compound impacts metabolic pathways acting as anticancer agents in colon cancer via an original mechanism of action that affects redox enzymes differently than platinum-based drugs.

Fireproofing of domestic upholstered furniture: Migration of flame retardants and potential risks.

Flame retardants (FRs) are widely incorporated in polyurethane foams to decrease their fire reaction. Currently, the risks associated with the use of FRs in domestic upholstered furniture (UF) are evaluated according to FRs volatility and potency to be emitted into the atmosphere. However, exposure via contact and dermal penetration, mediated by sweat, has not been considered so far. Our study provides an identification of the latest-generation of FRs most commonly used in UF, and an evaluation of their potency to migrate into artificial sweat. First of all, an extensive literature search, along with surveys with professionals, led to the identification of twenty-two FRs and synergists commonly used in France and Europe. Then, migration into artificial sweat of various FRs embedded into synthetic or commercially available polymer matrix was studied and evidenced. These results were analysed in the light of their potential effects on human health and the environment. Based on the migration's data, it is not possible to clearly rule out potential effects of FRs on human and environment health. Therefore, the authors consider that the use of FRs in domestic upholstery does not seem to be justified due to potential risks and a lack of clear benefits.

Laminin α1 orchestrates VEGFA functions in the ecosystem of colorectal carcinoma.

BACKGROUND INFORMATION: Tumor stroma remodeling is a key feature of malignant tumors and can promote cancer progression. Laminins are major constituents of basement membranes that physically separate the epithelium from the underlying stroma. RESULTS: By employing mouse models expressing high and low levels of the laminin α1 chain (LMα1), we highlighted its implication in a tumor-stroma crosstalk, thus leading to increased colon tumor incidence, angiogenesis and tumor growth. The underlying mechanism involves attraction of carcinoma-associated fibroblasts by LMα1, VEGFA expression triggered by the complex integrin α2ß1-CXCR4 and binding of VEGFA to LM-111, which in turn promotes angiogenesis, tumor cell survival and proliferation. A gene signature comprising LAMA1, ITGB1, ITGA2, CXCR4 and VEGFA has negative predictive value in colon cancer. CONCLUSIONS: Together, we have identified VEGFA, CXCR4 and α2ß1 integrin downstream of LMα1 in colon cancer as of bad prognostic value for patient survival. SIGNIFICANCE: This information opens novel opportunities for diagnosis and treatment of colon cancer.

SNAI2 and TWIST1 in lymph node progression in early stages of NSCLC patients.

Lymph node metastasis is an important prognosis factor in non-small cell lung cancer (NSCLC) patients. The aim of this study was to investigate the role of epithelial to mesenchymal transition (EMT) in lymph node progression in the early stages of NSCLC. We studied a retrospective cohort of 160 consecutive surgically treated NSCLC patients with available frozen tumor samples for expression of EMT markers (CDH1, CTNNB1, CDH2, and VIMENTIN), inducers (TGFB1, c-MET, and CAIX), and transcription factors (EMT-TF: SNAI1, SNAI2, ZEB1, TWIST1, and TWIST2). Partial EMT was more frequent in N1-2 (N+) vs N0 patients (P < .01). TGFB1 (P = .02) as well as SNAI2 (P < .01) and TWIST1 (P = .04) were the most differentially expressed genes in N+ tumors. In this group, ZEB1 was correlated with all EMT inducers and other EMT-TFs were overexpressed depending on the inducers. CAIX was an independent prognostic factor for overall survival (IC 95% HR: 1.10-5.14, P = .03). Partial EMT is involved in lymph node progression of NSCLC patients and depends on the TGFß pathway. EMT-TFs are differentially expressed depending on EMT inducers. CAIX might be a relevant prognostic marker in early stage NSCLC.

The Balance Between Cytotoxic T-cell Lymphocytes and Immune Checkpoint Expression in the Prognosis of Colon Tumors.

Background: Immune checkpoint (ICK) expression might represent a surrogate measure of tumor-infiltrating T cell (CTL) exhaustion and therefore be a more accurate prognostic biomarker for colorectal cancer (CRC) patients than CTL enumeration as measured by the Immunoscore. Methods: The expression of ICKs, Th1, CTLs, cytotoxicity-related genes, and metagenes, including Immunoscore-like metagenes, were evaluated in three independent cohorts of CRC samples (260 microsatellite instable [MSI], 971 non-MSI). Their associations with patient survival were analyzed by Cox models, taking into account the microsatellite instability (MSI) status and affiliation with various Consensus Molecular Subgroups (CMS). PD-L1 and CD8 expression were examined on a subset of tumors with immunohistochemistry. All statistical tests were two-sided. Results: The expression of Immunoscore-like metagenes was statistically significantly associated with improved outcome in non-MSI tumors displaying low levels of both CTLs and immune checkpoints (ICKs; CMS2 and CMS3; hazard ratio [HR] = 0.63, 95% confidence interval [CI] = 0.43 to 0.92, P = .02; and HR = 0.55, 95% CI = 0.34 to 0.90, P = .02, respectively), but clearly had no prognostic relevance in CRCs displaying higher levels of CTLs and ICKs (CMS1 and CMS4; HR = 0.46, 95% CI = 0.10 to 2.10, P = .32; and HR = 1.13, 95% CI = 0.79 to 1.63, P = .50, respectively), including MSI tumors. ICK metagene expression was statistically significantly associated with worse prognosis independent of tumor staging in MSI tumors (HR = 3.46, 95% CI = 1.41 to 8.49, P = .007). ICK expression had a negative impact on the proliferation of infiltrating CD8 T cells in MSI neoplasms (median = 0.56 in ICK low vs median = 0.34 in ICK high, P = .004). Conclusions: ICK expression cancels the prognostic relevance of CTLs in highly immunogenic colon tumors and predicts a poor outcome in MSI CRC patients.

Characterization of the transcriptional and metabolic responses of pediatric high grade gliomas to mTOR-HIF-1α axis inhibition.

Pediatric high grade glioma (pHGGs), including sus-tentorial and diffuse intrinsic pontine gliomas, are known to have a very dismal prognosis. For instance, even an increased knowledge on molecular biology driving this brain tumor entity, there is no treatment able to cure those patients. Therefore, we were focusing on a translational pathway able to increase the cell resistance to treatment and to reprogram metabolically tumor cells, which are, then, adapting easily to a hypoxic microenvironment. To establish, the crucial role of the hypoxic pathways in pHGGs, we, first, assessed their protein and transcriptomic deregulations in a pediatric cohort of pHGGs and in pHGG's cell lines, cultured in both normoxic and hypoxic conditions. Secondly, based on the concept of a bi-therapy targeting in pHGGs mTORC1 (rapamycin) and HIF-1α (irinotecan), we hypothesized that the balanced expressions between RAS/ERK, PI3K/AKT and HIF-1α/HIF-2α/MYC proteins or genes may provide a modulation of the cell response to this double targeting. Finally, we could evidence three protein, genomic and metabolomic profiles of response to rapamycin combined with irinotecan. The pattern of highly sensitive cells to mTOR/HIF-1α targeting was linked to a MYC/ERK/HIF-1α over-expression and the cell resistance to a major hyper-expression of HIF-2α.

Histone hypoacetylation contributes to CXCL12 downregulation in colon cancer: impact on tumor growth and cell migration.

CXCL12 has been shown to be involved in colon cancer metastasis, but its expression level and molecular mechanisms regulating its expression remain controversial. We thus evaluated CXCL12 expression in a large cohort of colon adenomas and carcinomas, investigated for an epigenetic mechanism controlling its expression and evaluated the impact of CXCL12 levels on cell migration and tumor growth. CXCL12 expression was measured in human colon adenomas and carcinomas with transcriptome array and RT-qPCR. The promoter methylation was analyzed with whole-genome DNA methylation chips and protein expression by immunohistochemistry. We confirm a reduced expression of CXCL12 in 75% of MSS carcinomas and show that the decrease is an early event as already present in adenomas. The methylome analysis shows that the CXCL12 promoter is methylated in only 30% of microsatellite-stable tumors. In vitro, treatments with HDAC inhibitors, butyrate and valproate restored CXCL12 expression in three colon cell lines, increased acetylation of histone H3 within the CXCL12 promoter and inhibited cell migration. In vivo, valproate diminished (65%) the number of intestinal tumors in APC mutant mice, slowed down xenograft tumor growth concomitant to restored CXCL12 expression. Finally we identified loss of PCAF expression in tumor samples and showed that forced expression of PCAF in colon cancer cell lines restored CXCL12 expression. Thus, reduced PCAF expression may participate to CXCL12 promoter hypoacetylation and its subsequent loss of expression. Our study is of potential clinical interest because agents that promote or maintain histone acetylation through HDAC inhibition and/or HAT stimulation, may help to lower colon adenoma/carcinoma incidence, especially in high-risk families, or could be included in therapeutic protocols to treat advanced colon cancer.

Perioperative bevacizumab improves survival following lung metastasectomy for colorectal cancer in patients harbouring v-Ki-ras2 Kirsten rat sarcoma viral oncogene homologue exon 2 codon 12 mutations†.

OBJECTIVES: The role of perioperative chemotherapy (POC) and targeted therapies in lung metastasectomy for colorectal cancer (CRC) is still subject to debate. We aimed to evaluate whether POC and targeted therapies were associated with different outcomes according to the mutational status. METHODS: We reviewed data from 223 patients who underwent pulmonary metastasectomy for CRC from 1998 to 2015 and for whom the V-Ki-ras2 Kirsten sarcoma viral oncogene homologue (KRAS) and V-raf Murine sarcoma viral oncogene homologue B1 (BRAF) mutational statuses were known. RESULTS: A total of 167 patients (74%) underwent POC: 62 (37%) received neoadjuvant therapy, 59 (35%) were in the adjuvant setting and 46 (28%) were in both the neoadjuvant and adjuvant settings. POC did not significantly influence either the loco-regional recurrence free survival (LRRFS) (P = 0.21) or the overall survival (OS) (P = 0.29). Furthermore, in cases of adjuvant chemotherapy, outcomes were not significantly different in cases of neoadjuvant chemotherapy or both neoadjuvant and adjuvant treatment (P = 0.26 for OS, P = 0.14 for LRRFS). For patients with KRAS mutation, perioperative bevacizumab was associated with a significant improvement in both LRRFS [70 months (41.58­98.42) vs 24 months (1.15­46.86), P = 0.001] and OS [101 vs 55 months (49.77­60.23), P = 0.004]. However, this benefit was only significant in cases of KRAS exon 2 codon 12 mutations [median OS: 101 months (83.97­118.02) vs 60 months (53­66.99), P < 0.001; median LRRFS: 76 months (64.62­87.38) vs 44 months (35.27­52.73), P < 0.001]. CONCLUSION: Perioperative bevacizumab appears to be beneficial in patients with exon 2 codon 12 KRAS mutations who have undergone lung metastasectomy for CRC.

Development of a memetic clustering algorithm for optimal spectral histology: application to FTIR images of normal human colon.

The coupling between Fourier-transform infrared (FTIR) imaging and unsupervised classification is effective in revealing the different structures of human tissues based on their specific biomolecular IR signatures; thus the spectral histology of the studied samples is achieved. However, the most widely applied clustering methods in spectral histology are local search algorithms, which converge to a local optimum, depending on initialization. Multiple runs of the techniques estimate multiple different solutions. Here, we propose a memetic algorithm, based on a genetic algorithm and a k-means clustering refinement, to perform optimal clustering. In addition, this approach was applied to the acquired FTIR images of normal human colon tissues originating from five patients. The results show the efficiency of the proposed memetic algorithm to achieve the optimal spectral histology of these samples, contrary to k-means.

Fully unsupervised inter-individual IR spectral histology of paraffinized tissue sections of normal colon.

In label-free Fourier-transform infrared histology, spectral images are individually recorded from tissue sections, pre-processed and clustered. Each single resulting color-coded image is annotated by a pathologist to obtain the best possible match with tissue structures revealed after Hematoxylin-Eosin staining. However, the main limitations of this approach are the empirical choice of the number of clusters in unsupervised classification, and the marked color heterogeneity between the clustered spectral images. Here, using normal murine and human colon tissues, we developed an automatic multi-image spectral histology to simultaneously analyze a set of spectral images (8 images mice samples and 72 images human ones). This procedure consisted of a joint Extended Multiplicative Signal Correction (EMSC) to numerically deparaffinize the tissue sections, followed by an automated joint K-Means (KM) clustering using the hierarchical double application of Pakhira-Bandyopadhyay-Maulik (PBM) validity index. Using this procedure, the main murine and human colon histological structures were correctly identified at both the intra- and the inter-individual levels, especially the crypts, secreted mucus, lamina propria and submucosa. Here, we show that batched multi-image spectral histology procedure is insensitive to the reference spectrum but highly sensitive to the paraffin model of joint EMSC. In conclusion, combining joint EMSC and joint KM clustering by double PBM application allows to achieve objective and automated batched multi-image spectral histology.

Impact of EGFR mutations and KRAS amino acid substitution on the response to radiotherapy for brain metastasis of non-small-cell lung cancer.

BACKGROUND: Our study aimed to evaluate response rate (RR) to brain metastasis radiotherapy (RT), depending on the genomic status of non-small-cell lung cancer. MATERIAL & METHODS: We retrospectively reviewed 1971 non-small-cell lung cancer files of patients with EGFR and KRAS testing and focused on 157 patients who had undergone RT for brain metastasis. RESULTS: A total of 16 patients (10.2%) harbored EGFR mutations (mEGFR) and 45 patients (28.7%) KRAS (mKRAS). In univariate analysis, RR was significantly higher for mEGFR compared with wild-type EGFR/KRAS (odds ratio [OR]: 4.96; p = 0.05) or mKRAS (OR: 1.81; p = 0.03). In multivariate analysis, KRAS G12V or G12C status was associated with both poor RR (OR: 0.1; p < 0.0001) and overall survival (OR: 3.41; p < 0.0001). CONCLUSION: mEGFR are associated with higher RR to brain RT than wild-type EGFR/RAS or mKRAS.

Prognostic value of the KRAS G12V mutation in 841 surgically resected Caucasian lung adenocarcinoma cases.

BACKGROUND: Identifying patients who will experience lung cancer recurrence after surgery remains a challenge. We aimed to evaluate whether mutant forms of epidermal growth factor receptor (EGFR) and Kirsten rat sarcoma viral oncogene homolog (KRAS) (mEGFR and mKRAS) are useful biomarkers in resected non-small cell lung cancer (NSCLC). METHODS: We retrospectively reviewed data from 841 patients who underwent surgery and molecular testing for NSCLC between 2007 and 2012. RESULTS: mEGFR was observed in 103 patients (12.2%), and mKRAS in 265 (31.5%). The median overall survival (OS) and time to recurrence (TTR) were significantly lower for mKRAS (OS: 43 months; TTR: 19 months) compared with mEGFR (OS: 67 months; TTR: 24 months) and wild-type patients (OS: 55 months; disease-free survival (DFS): 24 months). Patients with KRAS G12V exhibited worse OS and TTR compared with the entire cohort (OS: KRAS G12V: 26 months vs COHORT: 60 months; DFS: KRAS G12V: 15 months vs COHORT: 24 months). These results were confirmed using multivariate analyses (non-G12V status, hazard ratio (HR): 0.43 (confidence interval: 0.28-0.65), P<0.0001 for OS; HR: 0.67 (0.48-0.92), P=0.01 for TTR). Risk of recurrence was significantly lower for non-KRAS G12V (HR: 0.01, (0.001-0.08), P<0.0001). CONCLUSIONS: mKRAS and mEGFR may predict survival and recurrence in early stages of NSCLC. Patients with KRAS G12V exhibited worse OS and higher recurrence incidences.