OBJECTIVES: Recent research has classified lung adenocarcinoma patients with KRAS mutation into three subtypes by co-occurring genetic events in TP53 (KP subgroup), STK11/LKB1 (KL subgroup) and CDKN2A/B inactivation plus TTF-1 low expression (KC subgroup). The aim of this study was to identify valuable biomarkers by searching the candidate molecules that contribute to lung adenocarcinoma pathogenesis, especially KC subtype.
MATERIALS AND METHODS: We analyzed the publicly available database and identified the candidate REG4 using the E-GEOD-31210 dataset, and then confirmed by TCGA dataset. In addition, an independent cohort of 55 clinical samples was analyzed by quantitative real-time PCR analysis. Functional studies and RNA sequencing were performed after silencing the REG4 expression.
RESULTS: REG4, an important regulator of gastro-intestinal carcinogenesis, was highly expressed in KRAS mutant lung adenocarcinoma with low expression of TTF-1 (KC subtype). The results were validated both by gene expression analysis and immunohistochemistry study in an independent 55 clinical samples from Fudan University Shanghai Cancer Center. Further in vitro and in vivo functional assays revealed silencing REG4 expression significantly reduces cancer cell proliferation and tumorigenesis. Moreover, RNA sequencing and GSEA analysis displayed that REG4 knockdown might induce cell cycle arrest by regulating G2/M checkpoint and E2F targets.
CONCLUSION: Our results indicate that REG4 plays an important role in KRAS-driven lung cancer pathogenesis and is a novel biomarker of lung adenocarcinoma subtype. Future studies are required to clarify the underlying mechanisms of REG4 in the division and proliferation of KC tumors and its potential therapeutic value.

Mechanisms of lung squamous cell carcinoma (LSCC) development are poorly understood. Here, we report that JNK1/2 activities attenuate Lkb1-deficiency-driven LSCC initiation and progression through repressing ΔNp63 signaling. In vivo Lkb1 ablation alone is sufficient to induce LSCC development by reducing MKK7 levels and JNK1/2 activities, independent of the AMPKα and mTOR pathways. JNK1/2 activities is positively regulated by MKK7 during LSCC development. Pharmaceutically elevated JNK1/2 activities abates Lkb1 dependent LSCC formation while compound mutations of Jnk1/2 and Lkb1 further accelerate LSCC progression. JNK1/2 is inactivated in a substantial proportion of human LSCC and JNK1/2 activities positively correlates with survival rates of lung, cervical and head and neck squamous cell carcinoma patients. These findings not only determine a suppressive role of the stress response regulators JNK1/2 on LSCC development by acting downstream of the key LSCC suppresser Lkb1, but also demonstrate activating JNK1/2 activities as a therapeutic approach against LSCC.

Liver kinase B1 (

Loss of tumor suppressor liver kinase B1 (LKB1) promotes cancer cell proliferation but also leads to decreased metabolic plasticity in dealing with energy crises. Autophagy is a protective process involving self-cannibalization to maintain cellular energy homeostasis during nutrient deprivation. We developed a mouse model for

INTRODUCTION: Mutated tumor suppressor genes (TSG) such as TP53, STK11, and MGA are widely-reported. We hypothesized the presence of single mutation or co-occurring mutations in these specific genes may represent a significant therapeutic target for lung adenocarcinoma.
METHODS: We sequenced lung adenocarcinoma samples from 677 East-Asian patients, combined them with those from cBioPortal public database (including TCGA) and performed a comparative analysis between Asian and Caucasian populations.
RESULTS: East-Asian lung adenocarcinomas presented distinct driver-mutational distribution compared to that of Caucasians (79% vs 56%, p < 0.001). Similar results were observed in TSG mutations of TP53 (35% vs 46%, p = 0.150), STK11 (4% vs 17%, p = 0.006) and MGA (10% vs 4%, p = 0.166). Compared with none-mutational cases, the patients harboring TSG mutations are more likely to be male (p = 0.009), smokers (p < 0.001), and more advanced disease (p = 0.004). In addition, the TSG-mutated tumors had poorer differentiation (p < 0.001), and more likely to be solid or micropapillary-predominant adenocarcinomas (p < 0.001). Survival analysis showed that both overall survival (OS, p < 0.001) and post-recurrence survival (PRS, p < 0.001) became worse with the accumulation of TSG mutations. However, the prognostic variety was not found in Caucasian patients. Moreover, multivariate analysis proved the accumulation of TSG mutations independently predicts both unfavorable OS (HR = 0.435, 95% CI 0.245-0.774, p = 0.005) and PRS (HR = 0.491, 95% CI 0.269-0.894, p = 0.020) in East-Asian patients, adjusting all other survival-associated factors.
CONCLUSIONS: Co-occurring mutations of specific TSGs define unfavorable subgroups of lung adenocarcinoma, implying that the tumor promotion mechanisms contribute to the heterogeneity in tumor evolution. However, the Caucasian population did not show the same results, providing insights into the molecular basis underlying the striking racial disparities of this disease and evidence for different gene-panel designs for different population in the purpose of targeted therapy.

BACKGROUND: Simultaneous detection of multiple molecular biomarkers is helpful in the prediction of treatment response and prognosis for colorectal cancer (CRC) patients.
METHODS: A 22-gene panel consisting of 103 hotspot regions was utilized in the formalin-fixed paraffin-embedded (FFPE) tissue samples of 207 CRC patients, using the next-generation sequencing (NGS)-based multiplex PCR technique. Those 22 genes included AKT1, ALK, BRAF, CTNNB1, DDR2, EGFR, ERBB2, ERBB4, FBXW7, FGFR1, FGFR2, FGFR3, KRAS, MAP2K1, MET, NOTCH1, NRAS, PIK3CA, PTEN, SMAD4, STK11, and TP53.
RESULTS: Of the 207 patients, 193 had one or more variants, with 170, 20, and 3 having one, two, and three mutated genes, respectively. Of the total 414 variants identified in this study, 384, 25, and 5 were single-nucleotide variants, deletion, and insertion. The top four frequently mutated genes were TP53, KRAS, PIK3CA, and FBXW7. There was high consistency between the results of NGS-PCR technique and routine ARMS-PCR in KRAS and BRAF mutation detection. Univariate and multivariate analyses demonstrated that advanced TNM stage, elevated serum CEA, total variants number ≥ 2, AKT1 and PTEN mutation were independent predictors of shorter DFS; poor differentiation, advanced TNM stage, total variants number ≥ 2, BRAF, CTNNB1 and NRAS mutation were independent predictors of shorter OS.
CONCLUSIONS: It is feasible to detect multiple gene mutations with a 22-gene panel in FFPE CRC specimens. TNM stage and total variants number ≥ 2 were independent predictors of DFS and OS. Detection of multiple gene mutations may provide additional prognostic information to TNM stage in CRC patients.

INTRODUCTION: Heritage is the most important risk factor for breast cancer. About 15-20% of breast cancer is familial, referring to affected women who have one or more first- or second-degree relatives with the disease. The heritable component in these families is substantial, especially in families with aggregation of breast cancer with low age at onset. Identifying breast cancer susceptibility genes: Since the discovery of the highly penetrant autosomal dominant susceptibility genes BRCA1 and BRCA2 in the 1990s, several more breast cancer genes that confer a moderate to high risk of breast cancer have been identified. Furthermore, during the last decade, advances in genomic technologies have led to large scale genotyping in genome-wide association studies that have identified a considerable amount of common low penetrance loci. In total, the high risk genes, BRCA1, BRCA2, TP53, STK11, CD1 and PTEN account for approximately 20% of the familial risk. Moderate risk variants account for up to 5% of the inherited familial risk. The more than 180 identified low-risk loci explain 18% of the familial risk. Altogether more than half of the genetic background in familial breast cancer remains unclear. Other genes and low risk loci that explain a part the remaining fraction will probably be identified. Clinical aspects and future perspectives: Definitive clinical recommendations can be drawn only for carriers of germline variants in a limited number of high and moderate risk genes for which an association with breast cancer has been established. Future progress in evaluating previously identified breast cancer candidate variants and low risk loci as well as exploring new ones can play an important role in improving individual risk prediction in familial breast cancer.

As an innovative CXC chemokine, CXCL17 has a mysterious clinical significance and modulating influence on hepatocellular carcinoma (HCC). Our study examined the activity and mechanisms of CXCL17 on growth, autophagy, and metastasis of HCC. Upregulation of CXCL17 expression was observed in HCC, which is correlated with poorer histological stages and outcomes. Elevation of CXCL17 expression promoted proliferation, invasion, and migration and decreased LC-3B biosynthesis and p62 protein reduction, which are known to stimulate autophagy. However, silencing of CXCL17 inhibited the development of these cancerous phenotypes. Furthermore, AMPK was stimulated after knockdown of CXCL17. This stimulation, as well as stimulation of autophagy was caused by liver kinase B1 (LKB1), whose function is induced by knockdown CXCL17. Additionally, knockdown of CXCL17 enhanced nuclear translocation of LKB1. Altogether, these findings suggest that elevated CXCL17 expression in HCC promotes malignant reactions in malignant cells. Our research offers new evidence that chemokine CXCL17 reinforces malignant invasion and suppresses autophagy via the LKB1-AMPK pathway.

BACKGROUND: We conducted this study to identify regulatory variants in cancer-related pathway genes which can predict clinical outcomes of chemotherapy in advanced NSCLC, using a comprehensive list of regulatory SNPs prioritized by RegulomeDB.
METHODS: A total of 509 potentially functional SNPs in cancer-related pathway genes were evaluated. The SNPs were analyzed in a discovery set (n = 198), and an independent validation set (n = 181). The associations of the SNPs with chemotherapy response and overall survival (OS) were analyzed.
RESULTS: In the discovery set, 95 SNPs were significantly associated with clinical outcomes. Among the 95 SNPs, only rs10414193A > G in the intronic region of ARID3A, an eQTL for LKB1, was consistently associated with chemotherapy response and OS in the validation set. In combined analysis, the rs10414193A > G was significantly associated with worse response to chemotherapy (adjusted odds ratio = 0.63, 95% CI = 0.47-0.85, P = 0.002), and with worse OS (adjusted hazard ratio = 1.25, 95% CI = 1.08-1.45, P = 0.004). Luciferase assay showed a significantly higher LKB1 promoter activity associated with rs10414193G allele compared with rs10414193A allele (P = 0.0009).
CONCLUSIONS: Our results suggest that rs10414193A > G may be useful for the prediction of clinical outcomes of chemotherapy in advanced NSCLC.

BACKGROUND: The combination of direct sequencing and multiple ligation-dependent probe amplification (MLPA) has resulted in an 80% detection rate of serine/threonine kinase 11 (STK11) gene mutations in Peutz-Jeghers syndrome (PJS); however, this rate varies in different ethnicities.
AIMS: To test the efficacy of the combination in Chinese patients with PJS.
METHODS: PJS probands visiting our center during one year were enrolled. Sanger sequencing and MLPA were used to detect STK11 mutations. Associations between the occurrence of severe complications and risk factors were analyzed statistically.
RESULTS: We identified 47 PJS probands. Among them, 34 received an STK11 mutation test, revealing 23 point mutations and 2 exonic deletions. Nine of the mutations were splicing errors, reflecting a significantly higher proportion (p < 0.05). Laparotomy history existed for 33 of the probands, and seven families had a history of cancer. Statistical analysis revealed no associations between the occurrence of severe complications or cancers and risk factors.
CONCLUSION: The strategy achieved a high detection rate in Chinese people, validating its effectiveness. This cohort comprised a significantly higher proportion of splicing errors, reflecting the unique genetic characteristics Chinese people. No specific genotype-phenotype relationship was noted, while the wide usage of enteroscopy would benefit PJS surveillance.

BACKGROUND: Non-small-cell lung cancer (NSCLC) is a heterogeneous disease, with multiple different oncogenic mutations. Approximately 25-30% of NSCLC patients present KRAS mutations, which confer poor prognosis and high risk of tumor recurrence. About half of NSCLCs with activating KRAS lesions also have deletions or inactivating mutations in the serine/threonine kinase 11 (LKB1) gene. Loss of LKB1 on a KRAS-mutant background may represent a significant source of heterogeneity contributing to poor response to therapy.
METHODS: Here, we employed an integrated multilevel proteomics, metabolomics and functional in-vitro approach in NSCLC H1299 isogenic cells to define their metabolic state associated with the presence of different genetic background. Protein levels were obtained by label free and single reaction monitoring (SRM)-based proteomics. The metabolic state was studied coupling targeted and untargeted mass spectrometry (MS) strategy. In vitro metabolic dependencies were evaluated using 2-deoxy glucose (2-DG) treatment or glucose/glutamine nutrient limitation.
RESULTS: Here we demonstrate that co-occurring KRAS mutation/LKB1 loss in NSCLC cells allowed efficient exploitation of glycolysis and oxidative phosphorylation, when compared to cells with each single oncologic genotype. The enhanced metabolic activity rendered the viability of cells with both genetic lesions susceptible towards nutrient limitation.
CONCLUSIONS: Co-occurrence of KRAS mutation and LKB1 loss in NSCLC cells induced an enhanced metabolic activity mirrored by a growth rate vulnerability under limited nutrient conditions relative to cells with the single oncogenetic lesions. Our results hint at the possibility that energy stress induced by calorie restriction regimens may sensitize NSCLCs with these co-occurring lesions to cytotoxic chemotherapy.

The occurrence of colorectal cancer (CRC) is associated with a variety of oncogenes and tumor‑suppressor genes. As a tumor‑suppressor gene, the liver kinase B1 gene (LKB1, also known as serine/threonine kinase 11, STK11) is closely related to tumor angiogenesis, invasion and metastasis, but its molecular mechanisms remain unclear. The aim of the present study was to investigate the effects of LKB1 on the invasion and metastasis of CRC, and to explore its molecular mechanisms. By detecting the expression of LKB1 in CRC, we can provide a reference index for diagnosing the depth of invasion and lymph node metastasis. Immunohistochemistry results indicated that LKB1 expression was strongly positive in normal colon tissue and that it inhibited the production of CRC. Immunocytochemical staining showed that the expression of LKB1 was significantly decreased in adenocarcinoma and mucinous adenocarcinoma tissues, and this reduced expression induced the invasion and metastasis of CRC. In the present study, LKB1 small interfering RNA (LKB1 siRNA) was transfected into LoVo cells to observe the effect of LKB1 on the invasion and metastasis of CRC. LKB1 silencing decreased the phosphorylation of AMP‑activated protein kinase (p‑AMPK) in its downstream pathway, which increased the phosphorylation of protein kinase B (p‑AKT) and promoted tumor cell proliferation, enhancing the migration and invasion of CRC. The present study also explored the role of metformin in the LKB1 signaling pathway. Metformin inhibits the invasion and metastasis of CRC by activating p‑AMPK, thereby inhibiting the activation of p‑AKT. These results suggest that LKB1 plays an important role in the invasion and metastasis of CRC by activating AMPK, negatively regulating the AKT signaling pathway and regulating gene expression. Mutation or deletion of LKB1 is expected to be a novel therapeutic target or clinical biomarker for the prevention of the invasion and metastasis of CRC.

Background Peutz-Jeghers syndrome (PJS) is characterized by gastrointestinal polyposis, mucocutaneous pigmentation and cancer predisposition. Patients with PJS can develop large calcifying Sertoli cell tumors (LCSTs). Case presentation A patient presented at 3 years of age with delayed development, hypermobility and later also with tall stature and advanced bone age. Extensive endocrine evaluation, mutation analysis of genes associated with connective tissue disorders and a single nucleotide polymorphism (SNP) array showed no abnormalities. At 8 years of age, gynecomastia developed as well as pigmentations on the lips, both of which are associated with PJS. Mutation analysis showed a heterozygous deletion of the whole STK11 gene confirming PJS. Testicular ultrasound confirmed the presence of LCSTs. Interestingly, the previously performed SNP array did not report deletion of the STK11 gene. Conclusions We advise excluding LCSTs in children with tall stature and advanced bone age where more common causes have been eliminated. Although STK11 deletions are documented in control databases, reporting the deletion of this gene even in the absence of a phenotype is advised for patient management.

BACKGROUND/AIM: Gastric cancer (GC) with peritoneal metastasis remains difficult to treat. The anti-diabetic drug metformin exerts various antitumor effects via the 5'-adenosine monophosphate-activated protein kinase (AMPK) pathway and nuclear factor-kappa B (NF-ĸB). Therefore, we evaluated the antitumor effects of metformin for GC in vitro and on peritoneal metastasis.
MATERIALS AND METHODS: The human GC cell lines MKN1, MKN45, KATO-III and SNU-1 were used. The antiproliferative effect was evaluated in vitro with 0.5 mM or 25 mM glucose and in vivo using tumor xenograft peritoneal models of metastasis. The protein expression of AMPK, liver kinase B1 (LKB1) and NF-ĸB in tumors was examined by western blotting.
RESULTS: Metformin inhibited cell proliferation in all GC lines and sensitivity was increased under low-glucose conditions in vitro. Metformin also suppressed peritoneal metastasis. In tumors, metformin reduced the numbers of proliferating cells and NF-ĸB expression, but a similar trend was not noted for AMPK.
CONCLUSION: Metformin may be a useful drug for the treatment of GC with peritoneal metastasis.

The major types of non-small-cell lung cancer (NSCLC)-squamous cell carcinoma and adenocarcinoma-have distinct immune microenvironments. We developed a genetic model of squamous NSCLC on the basis of overexpression of the transcription factor Sox2, which specifies lung basal cell fate, and loss of the tumor suppressor Lkb1 (SL mice). SL tumors recapitulated gene-expression and immune-infiltrate features of human squamous NSCLC; such features included enrichment of tumor-associated neutrophils (TANs) and decreased expression of NKX2-1, a transcriptional regulator that specifies alveolar cell fate. In Kras-driven adenocarcinomas, mis-expression of Sox2 or loss of Nkx2-1 led to TAN recruitment. TAN recruitment involved SOX2-mediated production of the chemokine CXCL5. Deletion of Nkx2-1 in SL mice (SNL) revealed that NKX2-1 suppresses SOX2-driven squamous tumorigenesis by repressing adeno-to-squamous transdifferentiation. Depletion of TANs in SNL mice reduced squamous tumors, suggesting that TANs foster squamous cell fate. Thus, lineage-defining transcription factors determine the tumor immune microenvironment, which in turn might impact the nature of the tumor.

Many recent efforts to analyze cancer genomes involve aggregation of mutations within reference maps of molecular pathways and protein networks. Here, we find these pathway studies are impeded by molecular interactions that are functionally irrelevant to cancer or the patient's tumor type, as these interactions diminish the contrast of driver pathways relative to individual frequently mutated genes. This problem can be addressed by creating stringent tumor-specific networks of biophysical protein interactions, identified by signatures of epistatic selection during tumor evolution. Using such an evolutionarily selected pathway (ESP) map, we analyze the major cancer genome atlases to derive a hierarchical classification of tumor subtypes linked to characteristic mutated pathways. These pathways are clinically prognostic and predictive, including the TP53-AXIN-ARHGEF17 combination in liver and CYLC2-STK11-STK11IP in lung cancer, which we validate in independent cohorts. This ESP framework substantially improves the definition of cancer pathways and subtypes from tumor genome data.

RATIONALE: Adrenal hepatoid adenocarcinoma typically secretes alpha-fetoprotein (AFP). Here, we report a case of non-AFP-producing adrenal hepatoid adenocarcinoma. Next-generation sequencing (NGS) was conducted to identify gene mutations.
PATIENT CONCERNS: A 64-year-old man presented with mild back pain and unexplained weight loss for 3 months.
DIAGNOSES: Contrast-enhanced magnetic resonance imaging (MRI) showed a mass (9.9 × 9.7 × 9.1 mm) above the upper pole of the left kidney. The left renal artery and vein were compressed. The tumor was positive for CK8/18, CK19, CK7, hepatocyte marker (Hepatocyte), and Hep Par 1, but negative for AFP. Plasma AFP was 2.75 ng/mL (normal range: 0-7 ng/mL). NGS revealed mutations of the following genes: ATM, CDKN2A, EGFR, STK11, TP53, BIM, and MLH1. A diagnosis of adrenal hepatoid adenocarcinoma was established.
INTERVENTIONS: The treatment included 4 cycles of the mFOLFOX6 regimen (oxaliplatin, leucovorin, and fluorouracil), transcatheter arterial chemoembolization, and apatinib.
OUTCOMES: The patient died 9 months after the diagnosis.
LESSONS: This case highlights the importance of thorough clinical, radiological, and immunohistochemical investigation for suspected adrenal hepatoid adenocarcinoma. Metastasis from other primary tumors should be ruled out. Furthermore, AFP is not necessarily elevated in adrenal hepatoid adenocarcinoma. NGS could be helpful in establishing the diagnosis and selecting treatments.

BACKGROUND: Genetic alterations occurring in lung cancer are the basis for defining molecular subtypes and essential for targeted therapies. Exhaled breath condensate (EBC) is a form of non-invasive sample that, amongst components, contains DNA from pulmonary tissue. Next-generation sequencing (NGS) was herein used to analyze mutations in EBC from patients with lung cancer.
MATERIALS AND METHODS: EBC was collected from 26 patients with cancer and 20 healthy controls. Amplicon-based sequencing using Ion Ampliseq Colon and Lung Cancer gene panel v2 was applied.
RESULTS: The sequencing was successful in 17 patients and 20 controls. EBC from patients revealed 39 hotspot mutations occurring in: adenomatous polyposis coli (APC), v-raf murine sarcoma viral oncogene homolog B (BRAF), discoidin domain receptor tyrosine kinase 2 (DDR2), epidermal growth factor receptor (EGFR), erb-b2 receptor tyrosine kinase 4 (ERBB4), F-box and WD repeat domain containing 7 (FBXW7), fibroblast growth factor receptor 1 (FGFR1), FGFR3 (fibroblast growth factor receptor 3), Kirsten rat sarcoma viral oncogene homolog (KRAS), mitogen-activated protein kinase kinase 1 (MAP2K1), met proto-oncogene (MET), neuroblastoma RAS viral (v-ras) oncogene homolog (NRAS), phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), phosphatase and tensin homolog (PTEN), ret proto-oncogene (RET), SMAD family member 4 (SMAD4), serine/threonine kinase 11 (STK11), and tumor protein p53 (TP53) genes. EBC from controls revealed 35 hotspot mutations. The average mutant allele fraction was higher in patients than controls.
CONCLUSION: NGS can identify mutations in EBCs from patients with lung cancer. This could provide a promising non-invasive method for the assessment of gene mutations in lung cancer.

BACKGROUND: KRAS-mutant lung adenocarcinomas (LUADs) are heterogeneous and frequently occur in smokers. The heterogeneity of KRAS-mutant LUAD has been an obstacle for the drug discovery.
METHODS: We integrated multiplatform datatypes and identified two corresponding subtypes in the patients and cell lines. We further characterized the features of these two subtypes and performed drug screening to identify subtype-specific drugs. Finally, we used the defining features of the KRAS subtypes for drug sensitivity prediction.
FINDINGS: Patient-Subtype 1 (PS1) was characterized by increased smoking-related mutational signature activity, a low tumor-infiltrating lymphocyte (TIL)-associating score and STK11/KEAP1 co-mutations. Patient-Subtype 2 (PS2) was characterized by an increased smoking-related methylation signature activity, a high TIL-associating score and increased KRAS dependency. The cell line subtypes faithfully recapitulated all the patients' features. Drug screening of the two cell line subtypes yielded several potential candidates, such as cytarabine and enzastaurin for Cell-line-Subtype 1 (CS1) and a BTK inhibitor QL-XII-61 for Cell-line-Subtype 2 (CS2). The defining features, such as smoking-related methylation signature, were significantly associated with the sensitivity to several drugs.
INTERPRETATION: The heterogeneity of KRAS-mutant LUAD is associated with smoking-related genomic and epigenomic aberration along with other features such as immunogenicity, KRAS dependency and STK11/KEAP1 co-mutations. These features might be used as biomarkers for drug sensitivity prediction. FUND: This research was funded by the Young Scientists Fund of the National Natural Science Foundation of China, the Natural Science Foundation of Fujian Province, China and the Education and Research Foundation for Young Scholars of Education Department of Fujian Province, China.

NF1 is a tumor suppressor gene that negatively regulates Ras signaling. NF1 deficiency plays an important role in carcinogenesis. To investigate the frequency and clinical significance of NF1 mutation, we examined mutation status of NF1, TP53, LKB1 and RB1 in 704 surgically resected lung adenocarcinomas from East Asian patients using semiconductor-based Ion Torrent sequencing platform. Common driver events, including mutations in EGFR, KRAS, HER2, BRAF, MET, and fusions affecting ALK, RET and ROS1, were also concurrently detected. The correlation between NF1 mutations and clinicomolecular features of patients was further evaluated. Among 704 patients, 42 NF1 mutations were found in 33 patients (33/704, 4.7%), including 14 patients harboring EGFR/NF1 comutations (14/33, 42.4%). Comparing with EGFR-mutant patients, patients harboring NF1 mutations were closely associated with solid component subtype (p = 0.028). Comparing with KRAS mutations, NF1 mutations were found more common in female and never smokers (p = 0.003 and p = 0.004, respectively). Kaplan-Meier survival analysis revealed that patients harboring NF1 mutation had similar disease-free survival (DFS) and overall survival (OS) with patients with KRAS mutation. Although frequently overlapped with EGFR mutation, patients harboring NF1 mutation had significantly shorter DFS (p = 0.019) and OS (p = 0.004) than patients with EGFR mutation. During follow-up, one female patient with EGFR exon 19 deletion and NF1 Q1815X comutation showed poor response to EGFR TKIs (Gefitinib and Osimertinib) after disease relapse. In conclusion, NF1 mutations define a unique molecular and clinicopathologic subtype of lung adenocarcinoma. Examination of NF1 mutation may contribute to molecular subtyping and therapeutic intervention of lung adenocarcinoma.

The majority of studies report that liver kinase B1 (LKB1) acts as a tumor suppressor by inhibiting cell proliferation and metastasis. The present study investigated the expression pattern of LKB1 in 2 cohorts of paired hepatocellular carcinoma (HCC) and analogous non‑cancerous tissues (ANT). The results indicated that LKB1 was upregulated in HCC vs. ANT tissues, and that high expression of LKB1 was associated with a higher number of tumor foci, larger tumor size, poorer tumor differentiation, Edmondson-Steiner grade, Barcelona Clinic Liver Cancer grade and tumor-node-metastasis stage. Furthermore, high LKB1 expression was associated with poor overall survival (OS), shorter disease‑free survival and early recurrence. Univariate and multivariate analyses demonstrated that high LKB1 expression may serve as an independent prognostic marker for OS, but not for recurrence. In addition, knockdown of LKB1 expression in HCC cell lines inhibited cell proliferation and subcutaneous tumor growth by promoting cell apoptosis. Therefore, the findings of the present study suggest a proto-oncogenic role of LKB1 in HCC.

Somatic mutation analysis is a standard of practice for human cancers to identify therapeutic sensitization and resistance mutations. We performed a multigene sequencing screen to explore mutational hotspots in cancer-related genes using a semiconductor-based sequencer. DNA from oral squamous cell carcinoma samples was used as a template to amplify 207 regions from 50 cancer-related genes. Of the 80 oral squamous cell carcinoma specimens from Japanese patients, including formalin-fixed paraffin-embedded samples, 56 specimens presented at least one somatic mutation among the 50 investigated genes, and 17 of these samples showed multiple gene somatic mutations. TP53 was the most commonly mutated gene (50.0%), followed by CDKN2A (16.3%), PIK3CA (7.5%), HRAS (5.0%), MET (2.5%), and STK11 (2.5%). In total, 32 cases (40.0%) were human papillomavirus positive and they were significantly less likely to have a TP53, mutation than human papillomavirus-negative oral squamous cell carcinomas (8/32, 25.0% vs 32/48, 66.7%, p = 0.00026). We also detected copy number variations, in which segments of the genome could be duplicated or deleted from the sequencing data. We detected the tumor-specific TP53 mutation in the plasma cell-free DNA from two oral squamous cell carcinoma patients, and after surgery, the test for these mutations became negative. Our approach facilitates the simultaneous high-throughput detection of somatic mutations and copy number variations in oral squamous cell carcinoma samples.

Visual inspection of histopathology slides is one of the main methods used by pathologists to assess the stage, type and subtype of lung tumors. Adenocarcinoma (LUAD) and squamous cell carcinoma (LUSC) are the most prevalent subtypes of lung cancer, and their distinction requires visual inspection by an experienced pathologist. In this study, we trained a deep convolutional neural network (inception v3) on whole-slide images obtained from The Cancer Genome Atlas to accurately and automatically classify them into LUAD, LUSC or normal lung tissue. The performance of our method is comparable to that of pathologists, with an average area under the curve (AUC) of 0.97. Our model was validated on independent datasets of frozen tissues, formalin-fixed paraffin-embedded tissues and biopsies. Furthermore, we trained the network to predict the ten most commonly mutated genes in LUAD. We found that six of them-STK11, EGFR, FAT1, SETBP1, KRAS and TP53-can be predicted from pathology images, with AUCs from 0.733 to 0.856 as measured on a held-out population. These findings suggest that deep-learning models can assist pathologists in the detection of cancer subtype or gene mutations. Our approach can be applied to any cancer type, and the code is available at https://github.com/ncoudray/DeepPATH .

INTRODUCTION: The prostate exhibits a unique metabolism that changes during initial neoplasia to aggressive prostate cancer (PCa) and metastasis. The study of PCa metabolism thus represents a new avenue for diagnostics, particularly early diagnosis of aggressive PCa cases.
RESULTS: Here, by clustering tissue transcriptomics data from The Cancer Genome Atlas (498 PCa patients), we identified six metabolic subgroups (C1-C6) of PCa that show distinct disease-free survival (DFS) outcomes (p < 0.0001). In particular, we identified at least two subgroups (C5 & C3) that exhibit significant poor prognosis (∼70% and 30-40% relapse within the first 72 months; hazards ratios of 9.4 and 4.4, respectively, relative to the best prognosis cluster C4 that showed <20% relapse even by 120 months). We were able to reproduce the subgroups in several independent datasets including B. S. Taylor et al. (2010) data; 215 patients; DFS p = 0.00088) using a multinomial regression classifier. The subgroups displayed distinct metabolic profiles vis-à-vis normal tissues, measured as 'deregulation' observed for 20 metabolic pathways (using Pathifier; Y. Drier and E. Domany, 2013). In particular, C5 and C3 showed considerable deregulation for pathways involved in synthesis and catabolism of complex forms of lipids and carbohydrates, and these were exhibited in parallel or in the face of glycolysis, a common form of energy production in cancer cells. The subgroups were significantly over-enriched for different sets of genetic alterations [BRCA1, MSH2, FOXA1, TP53 (C5), RB1 and STK11(C3); and AR (C1); p ≤ 8.6 × 10-4], suggesting that distinct sets of alterations underpinning the PCa subgroups that 'push' the subgroups towards their unique metabolic profiles. Finally, applying the classifier to blood protein expression profiles from 42 active surveillance (AS) and 65 advanced castrate resistant PCa (ACRPC) patients (D. Olmos et al., 2012) assigned 70.77% ACPRC and interestingly reassigned 59.52% AS patients to at least one of the poor prognosis subgroups with 35.71% to the metabolically active poor-prognosis subgroup C3.
CONCLUSION: The identification of PCa subgroups displaying distinct clinical outcomes solely from metabolic expression profiles of PCa tumours reiterates the significant link between deregulated metabolism and PCa outcomes (E. Eidelman et al., 2017). On the other hand, the time to biochemical relapse (rise in PSA levels) was not indicative of early relapse seen for subgroups C3 and C5 (these show considerably late BCR compared to C4). Our study thus highlights specific processes (elevated lipid and carbohydrate metabolism pathways) that could be better indicators than PSA for early diagnosis of aggressive PCa.
AVAILABILITY: https://maxwellplus.com/research/metabolic-deregulation-in-prostate-cancer/.

Non-small cell lung carcinoma (NSCLC) accounts for significant morbidity and mortality worldwide, with most patients diagnosed at advanced stages and managed increasingly with targeted therapies and immunotherapy. In this review, we discuss diagnostic and predictive immunohistochemical markers in NSCLC, one of the most common tumors encountered in surgical pathology. We highlight 2 emerging diagnostic markers: nuclear protein in testis (NUT) for NUT carcinoma; SMARCA4 for SMARCA4-deficient thoracic tumors. Given their highly aggressive behavior, proper recognition facilitates optimal management. For patients with advanced NSCLCs, we discuss the utility and limitations of immunohistochemistry (IHC) for the "must-test" predictive biomarkers: anaplastic lymphoma kinase, ROS1, programmed cell death protein 1, and epidermal growth factor receptor. IHC using mutant-specific BRAF V600E, RET, pan-TRK, and LKB1 antibodies can be orthogonal tools for screening or confirmation of molecular events. ERBB2 and MET alterations include both activating mutations and gene amplifications, detection of which relies on molecular methods with a minimal role for IHC in NSCLC. IHC sits at the intersection of an integrated surgical pathology and molecular diagnostic practice, serves as a powerful functional surrogate for molecular testing, and is an indispensable tool of precision medicine in the care of lung cancer patients.

Peutz-Jeghers syndrome, a cancer predisposition syndrome that causes benign gastrointestinal polyps, results from a germline mutation in a single allele of STK11, the gene that encodes for LKB1. Researchers established a possible mechanism for how a STK11 mutation in T cells could lead to polyps associated with the syndrome.

BACKGROUND: Peutz-Jeghers syndrome (PJS) is caused by mutations in serine/threonine kinase 11 (STK11) gene. The increased cancer risk has been connected to P53 pathway.
METHODS: PJS probands with STK11 mutation were included in the function analysis. P53 activity elevated by STK11 mutants was investigated using dual-luciferase reporter assay in vitro after constructing expression vectors of STK11 wild type and mutants generated by site-directed substitution. The association between the P53 activity and clinicopathological factors was analysis, especially the cancer history.
RESULTS: Thirteen probands with STK11 mutations were involved, and within the mutations, c.G924A was novel. P53 activity elevation caused by 6 truncating mutations were significantly lower than that of STK11 wild type (P < 0.05). Family history of cancer was observed in 5 families. Within them, P53 activity was reduced and cancer occurred before 40 in 2 families, while it was not significantly changed and cancers happened after 45 in the other 3 families.
CONCLUSIONS: The affected P53 activity caused by STK11 mutations in PJS patients is significantly associated with protein truncation, while cancer risk in PJS can be elevated through pathways rather than P53 pathway. P53 activity test is probably a useful supporting method to predict cancer risk in PJS, which could be helpful in clinical practice.

OBJECTIVE: We previously postulated that 2-deoxyglucose (2-DG) activates multiple pro-survival pathways through IGF1R to negate its inhibitory effect on glycolysis. Here, we evaluated whether IGF1R inhibitor synergizes with 2-DG to impede the growth of non-small cell lung cancer (NSCLC).
MATERIALS AND METHODS: The activation of IGF1R signaling was assessed by the phosphorylation of IGF1R and its downstream target AKT using immunoblot. Drug dose response and combination index analyses were carried out according to the method of Chou and Talalay. Flow cytometry was used to evaluate cell cycle progression. Apoptosis was monitored by caspase-3/PARP cleavages or Annexin V staining. A subcutaneous xenograft model was used to assess this combination in vivo.
RESULTS: 2-DG induces the phosphorylation of IGF1R in its kinase domain, which can be abolished by the IGF1R inhibitor BMS-754807. Furthermore, the combination of 2-DG and BMS-754807 synergistically inhibited the survival of several non-small cell lung cancer (NSCLC) cell lines both in vitro and in vivo. The mechanistic basis of this synergy was cell line-dependent, and LKB1-inactivated EKVX cells underwent apoptosis following treatment with a subtoxic dose of 2-DG and BMS-754807. For these cells, the restoration of LKB1 kinase activity suppressed apoptosis induced by this combination but enhanced G1 arrest. In H460 cells, the addition of 2-DG did not enhance the low level of apoptosis induced by BMS-754807. However, treatment with 0.75 μM of BMS-754807 resulted in the accumulation of H460 cells with 8n-DNA content without affecting cell density increases. Hence, H460 cells may escape BMS-754807-induced G2/M cell cycle arrest through polyploidy. The inclusion of 2-DG blocked formation of the 8n-DNA cell population and restored G2/M phase cell cycle arrest.
CONCLUSION: The combination of 2-DG and IGF1R inhibitor BMS-754807 may be used to suppress the proliferation of NSCLC tumors through different mechanisms.

A hallmark of advanced tumors is a switch to aerobic glycolysis that is readily measured by [

BACKGROUND: Mutant peptides presented by MHC (major histocompatibility complex) Class II in cancer are important targets for cancer immunotherapy. Both animal studies and clinical trials in cancer patients showed that CD4 T cells specific to tumor-derived mutant peptides are essential for the efficacy of immune checkpoint blockade therapy by PD1 antibody.
RESULTS: In this study, we analyzed the next generation sequencing data of 147 lung adenocarcinoma patients from The Cancer Genome Atlas and predicted neoantigens presented by MHC Class I and Class II molecules. We found 18,175 expressed clonal somatic mutations, with an average of 124 per patient. The presentation of mutant peptides by an HLA(human leukocyte antigen) Class II molecule, HLA DRB1, were predicted by NetMHCIIpan3.1. 8804 neo-peptides, including 375 strong binders and 8429 weak binders were found. For HLA DRB1*01:01, 54 strong binders and 896 weak binders were found. The most commonly mutated genes with predicted neo-antigens are KRAS, TTN, RYR2, MUC16, TP53, USH2A, ZFHX4, KEAP1, STK11, FAT3, NAV3 and EGFR.
CONCLUSIONS: Our results support the feasibility of discovering individualized HLA Class II presented mutant peptides as candidates for immunodiagnosis and immunotherapy of lung adenocarcinoma.