Protein expression of the amino acid transporter SLC7A5 in tumor tissue is prognostic in early-stage colorectal cancer.

Proteins overexpressed in early-stage cancers may serve as early diagnosis and prognosis markers as well as targets for cancer therapies. In this study, we examined the expression of an essential amino acid carrier SLC7A5 (LAT1, CD98, or 4F2 light chain) in cancer tissue from two well-annotated cohorts of 575 cases of early-stage and 106 cases of late-stage colorectal cancer patients. Immunohistochemistry showed SLC7A5 overexpression in 72.0% of early-stage and 56.6% of late-stage cases. SLC7A5 expression was not influenced by patient gender, age, location, or mismatch repair status, although it appeared to be slightly less prevalent in tumors of mucinous differentiation or with lymphovascular invasion. Statistical analyses revealed a positive correlation between SLC7A5 overexpression and both overall survival and disease-free survival in early-stage but not late-stage cancers. Co-expression analyses of the TCGA and CPTAC colorectal cancer cohorts identified a network of gene transcripts positively related to SLC7A5, with its heterodimer partner SLC3A2 having the highest co-expression score. Network analysis uncovered the SLC7A network to be significantly associated with ncRNA such as tRNA processing and the mitotic cell cycle. Since SLC7A5 is also a marker of activated lymphocytes such as NK, T, and B lymphocytes, SLC7A5 overexpression in early colorectal cancers might trigger a strong anti-tumor immune response which could results in better clinical outcome. Overall, our study provides clear evidence of differential SLC7A5 expression and its prognostic value for early-stage colorectal cancer, although the understanding of its functions in colorectal tumorigenesis and cancer immunity is currently rather limited and awaits further characterization.

Proteogenomic characterization of primary colorectal cancer and metastatic progression identifies proteome-based subtypes and signatures.

Metastatic progression of colorectal adenocarcinoma (CRC) remains poorly understood and poses significant challenges for treatment. To overcome these challenges, we performed multiomics analyses of primary CRC and liver metastases. Genomic alterations, such as structural variants or copy number alterations, were enriched in oncogenes and tumor suppressor genes and increased in metastases. Unsupervised mass spectrometry-based proteomics of 135 primary and 123 metastatic CRCs uncovered distinct proteomic subtypes, three each for primary and metastatic CRCs, respectively. Integrated analyses revealed that hypoxia, stemness, and immune signatures characterize these 6 subtypes. Hypoxic CRC harbors high epithelial-to-mesenchymal transition features and metabolic adaptation. CRC with a stemness signature shows high oncogenic pathway activation and alternative telomere lengthening (ALT) phenotype, especially in metastatic lesions. Tumor microenvironment analysis shows immune evasion via modulation of major histocompatibility complex (MHC) class I/II and antigen processing pathways. This study characterizes both primary and metastatic CRCs and provides a large proteogenomics dataset of metastatic progression.

Impact of central nervous system metastasis after complete resection of lung adenocarcinomas harboring common EGFR mutation - A real-world database study in Japan: The CReGYT-01 EGFR study.

OBJECTIVES: To clarify the impact of central nervous system (CNS) metastasis on performance status (PS) at relapse, on subsequent treatment(s), and on survival of patients with lung adenocarcinoma harboring common epidermal growth factor receptor (EGFR) mutation. METHODS: We conducted the multicenter real-world database study for patients with radical resections for lung adenocarcinomas between 2015 and 2018 at 21 centers in Japan. EGFR mutational status was examined at each center. RESULTS: Of 4181 patients enrolled, 1431 underwent complete anatomical resection for lung adenocarcinoma harboring common EGFR mutations. Three-hundred-and-twenty patients experienced disease relapse, and 78 (24%) had CNS metastasis. CNS metastasis was significantly more frequent in patients with conventional adjuvant chemotherapy than those without (30% vs. 20%, P = 0.036). Adjuvant chemotherapy did not significantly improve relapse-free survival at any pathological stage (adjusted hazard ratio for stage IA2-3, IB, and II-III was 1.363, 1.287, and 1.004, respectively). CNS metastasis did not affect PS at relapse. Subsequent treatment, mainly consisting of EGFR-tyrosine kinase inhibitors (TKIs), could be equally given in patients with or without CNS metastasis (96% vs. 94%). Overall survival after relapse was equivalent between patients with and without CNS metastasis. CONCLUSION: The efficacy of conventional adjuvant chemotherapy may be limited in patients with lung adenocarcinoma harboring EGFR mutations. CNS metastasis is likely to be found in practice before deterioration in PS, and may have little negative impact on compliance with subsequent EGFR-TKIs and survival after relapse. In this era of adjuvant TKI therapy, further prospective observational studies are desirable to elucidate the optimal management of CNS metastasis.

Early-Stage Loss of GALNT6 Predicts Poor Clinical Outcome in Colorectal Cancer.

Colorectal adenocarcinomas arise from luminal lining epithelium of the colorectal tract which is covered with highly glycosylated mucins. Mucin O-glycosylation is initiated by a family of polypeptide N-acteylgalactosaminyltransferases (GALNTs). This study examined GALNT6 protein expression in 679 colorectal tumors, including 574 early-stage and 105 late-stage cancers. GALNT6 expression in cancer tissue varied widely between patients ranging from high levels to complete loss. Loss of GALNT6 occurred in 9.9% of early-stage and 15.2% of late-stage cancers and was more prevalent in grade 3 or MSI subtype tumors. Survival analyses revealed that loss of GALNT6 expression is prognostic of reduced overall survival, and univariate and multivariate analyses demonstrated that loss of GALNT6 is an independent risk variable. We also analyzed 508-case TCGA and 63-case CPTAC colorectal cancer cohorts for all members of the GALNT enzyme family, the mucin family, as well as KRAS and BRAF mutations. GLANT6 mRNA expression showed no strong correlation with other GALNTs or mucins but was significantly higher in KRAS mutated or BRAF wild-type early-stage cancers. Using large cohorts of patients and different approaches, this study shows that loss of GALNT6 enzyme in early-stage colorectal cancer predicts poor clinical outcomes.

One-step nucleic acid amplification for intraoperative diagnosis of lymph node metastasis in lung cancer patients: a single-center prospective study.

One-step nucleic acid amplification (OSNA) is a rapid intraoperative molecular detection technique for sentinel node assessment via the quantitative measurement of target cytokeratin 19 (CK19) mRNA to determine the presence of metastasis. It has been validated in breast cancer but its application in lung cancer has not been adequately investigated. 214 LNs from 105 patients with 100 primary lung cancers, 2 occult primary lung tumors, and 3 metastatic lung tumors, who underwent surgical lung resection with LN dissection between February 2018 and January 2020, were assessed. Resected LNs were divided into two parts: one was snap-frozen for OSNA and the other underwent rapidly frozen histological examination. Intraoperatively collected LNs were evaluated by OSNA using loop-mediated isothermal amplification and compared with intraoperative pathological diagnosis as a control. Among 214 LNs, 14 were detected as positive by OSNA, and 11 were positive by both OSNA and intraoperative pathological diagnosis. The sensitivity and specificity of OSNA was 84.6% and 98.5%, respectively. The results of 5 of 214 LNs were discordant, and the remainder all matched (11 positive and 198 negative) with a concordance rate of 97.7%. Although the analysis of public mRNA expression data from cBioPortal showed that CK19 expression varies greatly depending on the cancer type and histological subtype, the results of the five cases, except for primary lung cancer, were consistent. OSNA provides sufficient diagnostic accuracy and speed and can be applied to the intraoperative diagnosis of LN metastasis for non-small cell lung cancer.

Tumor stromal nicotinamide N-methyltransferase overexpression as a prognostic biomarker for poor clinical outcome in early-stage colorectal cancer.

In a quest for prognostic biomarkers in early-stage colorectal cancer, we investigated NNMT (nicotinamide N-methyltransferase) in large cohorts of patients. Immunohistochemical examination of 679 patients illustrates that NNMT protein is predominantly expressed in the cancer stroma at varying levels, and about 20% of cancer tissues overexpress NNMT when compared to levels observed in normal colorectal mucosa. Clinical correlation analyses of 572 patients with early-stage cancers reveal that NNMT protein overexpression is significantly associated with shorter overall and disease-free survival, but no such correlation is found in late-stage colorectal cancer. Analyses of TCGA and CPTAC colorectal cancer cohorts show that NNMT mRNA expression is positively correlated with protein levels, is significantly higher in CIMP-high or MSI subtypes than in CIMP-low or MSS subtypes, and is positively correlated with its paralog INMT but not with its interaction partners such as PNMT, ADK, APP, ATF6, BMF, BRD4, CDC37, or CRYZ. In early-stage cancers, NNMT expression is higher in BRAF-mutated than in BRAF wild type tumors but is not affected by KRAS or PIK3CA mutation status. As a cancer stromal protein with important roles in metabolism and cancer epigenetics, NNMT is emerging as a promising biomarker for risk stratification of early-stage cancers.

Pulmonary Enteric Adenocarcinoma Harboring a BRAF G469V Mutation.

Pulmonary enteric adenocarcinoma (PEAC) is a rare subtype of lung cancer that should be differentiated from colorectal cancer metastasis. Little is known about its genetic background. An 84-year-old male with adenocarcinoma of the lung underwent left upper lobectomy. The histology of the surgical specimen was suggestive of PEAC. Gastrointestinal and colorectal fiberscopy revealed no evidence of colorectal cancer. Next-generation sequencing of the tumor identified a G469V substitution in serine/threonine-protein kinase B-raf (BRAF). Based on the higher prevalence of the G469 substitution in BRAF-mutant lung adenocarcinoma than in BRAFmutant colorectal cancer, the tumor likely originated from the lung. Identification of mutational genotype may be of some help in distinguishing PEAC from the lung metastasis of colorectal cancer.

Chronic Lung Injury After Trimodality Therapy for Locally Advanced Non-Small Cell Lung Cancer.

BACKGROUND: Trimodality therapy is a treatment option for patients with locally advanced non-small cell lung cancer (LA-NSCLC). Thoracic radiation has both early (radiation pneumonitis) and late (chronic lung injury [CLI]) adverse effects on the lung. While CLI is expected to result in various problems in long-term survivors, these manifestations have not been precisely investigated. METHODS: We enrolled 112 LA-NSCLC patients who had received induction chemoradiotherapy followed by surgery, and then undergone follow-up computed tomography (CT) every 6 months for greater than 1 year. All chest CT images were reviewed to evaluate any injury of the pulmonary parenchyma. RESULTS: CLI at 1 year after surgery and its progression were observed in 94 (84%) and 38 (34%) patients, respectively. Progressive lung fibrosis as the first manifestation of CLI progression was most frequent after right middle and lower lobectomy. Cavity formation was the subsequent manifestation after progressive lung fibrosis , and chronic infection was the final stage of CLI. The cumulative rate of chronic infection was 76.4% at 10 years in patients with cavity formation. Ten patients with chronic infection included 7 cases of pulmonary aspergillosis and 2 cases of cavity infections with methicillin-resistant Staphylococcus aureus or Stenotrophomonas maltophilia. Among them, 4 patients required surgical interventions including completion pneumonectomy or fenestration. CONCLUSIONS: CLI is a common incidence after trimodality therapy for LA-NSCLC. CLI frequently results in cavity formation, which is a precursor of highly refractory chronic infections requiring surgical intervention. Appropriate management needs to be established for CLI developing after trimodality therapy.

The neutrophil-to-lymphocyte ratio as a novel independent prognostic factor for multiple metastatic lung tumors from various sarcomas.

PURPOSE: Sarcomas are among the most refractory malignant tumors and often recur as pulmonary metastasis. Although the presence of a high neutrophil-to-lymphocyte ratio (NLR) has been associated with the prognosis of several malignancies, the relationship between the NLR and sarcoma with pulmonary metastasis is unclear. We investigated the impact of the NLR in patients who underwent surgical resection for metastatic lung tumors from various sarcomas. METHODS: The subjects of this retrospective study were 158 patients with metastatic lung tumors from various sarcomas, who underwent initial pulmonary metastasectomy between 2006 and 2015. We examined the clinicopathological variables, including the NLR and the characteristics of surgical procedures. Survival was estimated by the Kaplan-Meier method and prognostic factors were evaluated by multivariate analysis. RESULTS: Multivariate analysis revealed significantly better survival of the group with an NLR < 2.26 immediately before the most recent pulmonary metastasectomy, in addition to such factors as the largest resected lesion being < 22 mm, a disease-free interval of > 2 years, and 3 or more pulmonary metastasectomies. CONCLUSION: The NLR immediately before the most recent pulmonary metastasectomy is a novel independent prognostic factor, which may be helpful when considering repeated pulmonary metastasectomy.

A Simple Prognostic Benefit Scoring System for Sarcoma Patients with Pulmonary Metastases: Sarcoma Lung Metastasis Score.

BACKGROUND: Pulmonary metastasectomy could be considered one of the treatment options for disease control in sarcoma patients with pulmonary metastases; however, there is little consensus regarding the suitable criteria for predicting the likely outcomes in these patients. The aim of this study was to establish a prognostic benefit scoring system based on preoperatively examined prognostic factors for sarcoma patients with pulmonary metastases. METHODS: This was a single-center, retrospective cohort study conducted in a cohort of 135 sarcoma patients who underwent a first pulmonary metastasectomy at Okayama University Hospital between January 2006 and December 2015. Based on the results of a multivariable logistic regression analysis performed to determine the factors influencing 3-year mortality, a Sarcoma Lung Metastasis Score was created and its correlation with 3-year survival was analyzed. RESULTS: The results of the multivariate analysis revealed significant differences in the disease-free interval (< 2 years vs. ≥ 2 years; odds ratio (OR) 4.22, 95% confidence interval (CI) 1.67-10.70), maximum tumor diameter (≥ 15 mm vs. < 15 mm; OR 3.86, 95% CI 1.75-8.52), and number of pulmonary metastases (≥ 6 vs. < 6; OR 2.65, 95% CI 1.06-6.620). The Sarcoma Lung Metastasis Score, which was defined as the total score of these three factors, reliably predicted 3-year survival (score: 0, 89.5%; 1, 63.2%; 2, 39.0%; 3, 10.5%). CONCLUSIONS: Our newly proposed simple Sarcoma Lung Metastasis Score appears to be a useful prognostic predictor for sarcoma patients with pulmonary metastases, in that it could be helpful for the selection of appropriate treatments for these patients.

Antitumor Effects of Pan-RAF Inhibitor LY3009120 Against Lung Cancer Cells Harboring Oncogenic BRAF Mutation.

BACKGROUND/AIM: The therapeutic strategy for patients with non-small-cell lung cancer (NSCLC) harboring the BRAF non-V600E mutation has not been established. LY3009120, a newly discovered pan-RAF inhibitor, has shown strong antitumor effects in cancers with various BRAF genotypes. This study investigated the antitumor effects of LY3009120 in NSCLC cells harboring the BRAF non-V600E mutation. MATERIALS AND METHODS: We examined the antitumor effects of LY3009120 by MTS assay and flow cytometry. We analyzed the expression status of proteins by western blot. The mouse xenograft models were used for the in vivo experiments. RESULTS: LY3009120 suppressed BRAF-related downstream pathway molecules and induced cleavage of poly ADP-ribose polymerase in all examined NSCLC cell lines. LY3009120 also inhibited in vivo tumor growth in NSCLC cells harboring the BRAF non-V600E mutation. CONCLUSION: LY3009120 is a potent therapeutic agent for patients with BRAF non-V600E mutant NSCLC.

YES1 activation induces acquired resistance to neratinib in HER2-amplified breast and lung cancers.

Molecular-targeted therapies directed against human epidermal growth factor receptor 2 (HER2) are evolving for various cancers. Neratinib is an irreversible pan-HER tyrosine kinase inhibitor and has been approved by the FDA as an effective drug for HER2-positive breast cancer. However, acquired resistance of various cancers to molecular-targeted drugs is an issue of clinical concern, and emergence of resistance to neratinib is also considered inevitable. In this study, we established various types of neratinib-resistant cell lines from HER2-amplified breast and lung cancer cell lines using several drug exposure conditions. We analyzed the mechanisms of emergence of the resistance in these cell lines and explored effective strategies to overcome the resistance. Our results revealed that amplification of YES1, which is a member of the SRC family, was amplified in two neratinib-resistant breast cancer cell lines and one lung cancer cell line. Knockdown of YES1 by siRNA and pharmacological inhibition of YES1 by dasatinib restored the sensitivity of the YES1-amplified cell lines to neratinib in vitro. Combined treatment with dasatinib and neratinib inhibited tumor growth in vivo. This combination also induced downregulation of signaling molecules such as HER2, AKT and MAPK. Our current results indicate that YES1 plays an important role in the emergence of resistance to HER2-targeted drugs, and that dasatinib enables such acquired resistance to neratinib to be overcome.

Acquired resistance mechanisms to afatinib in HER2-amplified gastric cancer cells.

Cancer treatment, especially that for breast and lung cancer, has entered a new era and continues to evolve, with the development of genome analysis technology and the advent of molecular targeted drugs including tyrosine kinase inhibitors. Nevertheless, acquired drug resistance to molecular targeted drugs is unavoidable, creating a clinically challenging problem. We recently reported the antitumor effect of a pan-HER inhibitor, afatinib, against human epidermal growth factor receptor 2 (HER2)-amplified gastric cancer cells. The purpose of the present study was to identify the mechanisms of acquired afatinib resistance and to investigate the treatment strategies for HER2-amplified gastric cancer cells. Two afatinib-resistant gastric cancer cell lines were established from 2 HER2-amplified cell lines, N87 and SNU216. Subsequently, we investigated the molecular profiles of resistant cells. The activation of the HER2 pathway was downregulated in N87-derived resistant cells, whereas it was upregulated in SNU216-derived resistant cells. In the N87-derived cell line, both MET and AXL were activated, and combination treatment with afatinib and cabozantinib, a multikinase inhibitor that inhibits MET and AXL, suppressed the cell growth of cells with acquired resistance both in vitro and in vivo. In the SNU216-derived cell line, YES1, which is a member of the Src family, was remarkably activated, and dasatinib, a Src inhibitor, exerted a strong antitumor effect in these cells. In conclusion, we identified MET and AXL activation in addition to YES1 activation as novel mechanisms of afatinib resistance in HER2-driven gastric cancer. Our results also indicated that treatment strategies targeting individual mechanisms of resistance are key to overcoming such resistance.

Droplet digital PCR as a novel system for the detection of microRNA­34b/c methylation in circulating DNA in malignant pleural mesothelioma.

Malignant pleural mesothelioma (MPM) is a rare malignancy arising from the pleura that is difficult to diagnose, contributing to its dismal prognosis. Previously, we reported that the degree of microRNA (miR)­34b/c methylation in circulating DNA is associated with the development of MPM. Herein, we present a newly developed droplet digital PCR (ddPCR)­based assay for the detection of miR­34b/c methylation in circulating DNA in patients with MPM. We originally prepared two probes within a short amplicon of 60 bp, designing one from the positive strand and the other from the complementary strand. The two probes functioned cooperatively, and our established assay detected DNA methylation accurately in the preliminary validation. We subsequently verified this assay using clinical samples. Serum samples from 35 cases of MPM, 29 cases of pleural plaque and 10 healthy volunteers were collected from 3 different institutions and used in this study. We divided the samples into 2 groups (group A, n=33; group B, n=41). A receiver­operating characteristic curve analysis using the samples in group A determined the optimal cut­off value for the diagnosis of MPM, with a sensitivity of 76.9% and a specificity of 90%. On the other hand, the use of the same criterion yielded a sensitivity of 59.1% and a specificity of 100% in group B, and corresponding values of 65.7 and 94.9% for the entire cohort, indicating a moderate sensitivity and a high specificity. In addition, when the analysis was focused on stage II or more advanced MPM, the sensitivity improved to 81.8%, suggesting the possibility that the methylated allele frequency in MPM may be associated with the stage of disease progression. On the whole, the findings of this study indicate that miR­34b/c methylation in circulating DNA is a promising biomarker for the prediction of disease progression in patients with MPM.

Ganetespib in Epidermal Growth Factor Receptor-Tyrosine Kinase Inhibitor-resistant Non-small Cell Lung Cancer.

BACKGROUND: The 90-kDa heat-shock protein (HSP90) is a chaperone protein expressed at high levels in cancer cells and is involved in the folding or stabilization of several client proteins, including epidermal growth factor receptor (EGFR). Ganetespib is a second-generation HSP90 inhibitor with a potent antitumor effect against various cancer types. MATERIALS AND METHODS: This study examined the antitumor effect of ganetespib in EGFR-mutant non-small cell lung cancer (NSCLC) cells and experimentally established EGFR-tyrosine kinase inhibitor (TKI)-resistant cells harboring various resistance mechanisms, including EGFR T790M mutation, met proto-oncogene amplification, and epithelial-mesenchymal transition. RESULTS: Ganetespib showed a potent antitumor effect at low concentrations, suppressing EGFR-related downstream pathway molecules and inducing cleavage of poly ADP-ribose polymerase in all examined EGFR-TKI-resistant cell lines in vitro. Ganetespib also inhibited in vivo tumor growth in resistant cells harboring EGFR T790M. CONCLUSION: Ganetespib might be a promising therapeutic option for the treatment of patients with EGFR-TKI-resistant NSCLC.

Anti-tumor effect of neratinib against lung cancer cells harboring HER2 oncogene alterations.

Human epidermal growth factor receptor 2 (HER2) is a member of the ErbB family of receptor tyrosine kinases. Numerous studies have reported the amplification and overexpression of HER2 in several types of cancer, including non-small cell lung cancer (NSCLC). However, the benefits of HER2-targeted therapy have not been fully established. In the present study, the anti-tumor effect of neratinib, an irreversible pan-HER tyrosine kinase inhibitor (TKI), against NSCLC cells harboring HER2 alterations was investigated. The sensitivity of normal bronchial epithelial cells (BEAS-2B) ectopically overexpressing wild-type or mutant HER2 to neratinib was assessed. Furthermore, the anti-tumor activity of neratinib in several NSCLC cell lines harboring HER2 alterations was determined in vitro and in vivo, and the association between their genetic alterations and sensitivity to neratinib treatment was investigated. BEAS-2B cells ectopically overexpressing wild-type HER2 or mutants (A775insYVMA, G776VC, G776LC, P780insGSP, V659E, G660D and S310F) exhibited constitutive autophosphorylation of HER2, as determined by western blotting. While these BEAS-2B cells were sensitive to neratinib, they were insensitive to erlotinib, a first-generation epidermal growth factor receptor-TKI. Neratinib also exerted anti-proliferative effects on HER2-altered (H2170, Calu-3 and H1781) NSCLC cell lines. Neratinib was also demonstrated to exert strong tumor growth inhibitory activity in mouse xenograft models using HER2-altered lung cancer cells. The results of the present study strongly suggest that neratinib has potential as a promising therapeutic option for the treatment of HER2-altered NSCLC.

Application of amplicon-based targeted sequencing with the molecular barcoding system to detect uncommon minor EGFR mutations in patients with treatment-naïve lung adenocarcinoma.

BACKGROUND: In lung cancer, epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor sensitizing mutations co-existing with rare minor EGFR mutations are known as compound mutations. These minor EGFR mutations can lead to acquired resistance after EGFR tyrosine kinase inhibitor treatment, so determining the mutation status of patients is important. However, using amplicon-based targeted deep sequencing based on next-generation sequencing to characterize mutations is prone to sequencing error. We therefore assessed the benefit of incorporating molecular barcoding with high-throughput sequencing to investigate genomic heterogeneity in treatment-naïve patients who have undergone resection of their non-small cell lung cancer (NSCLC) EGFR mutations. METHODS: We performed amplicon-based targeted sequencing with the molecular barcoding system (MBS) to detect major common EGFR mutations and uncommon minor mutations at a 0.5% allele frequency in fresh-frozen lung cancer samples. RESULTS: Profiles of the common mutations of EGFR identified by MBS corresponded with the results of clinical testing in 63 (98.4%) out of 64 cases. Uncommon mutations of EGFR were detected in seven cases (10.9%). Among the three types of major EGFR mutations, patients with the G719X mutation had a significantly higher incidence of compound mutations than those with the L858R mutation or exon 19 deletion (p = 0.0052). This was validated in an independent cohort from the Cancer Genome Atlas dataset (p = 0.018). CONCLUSIONS: Our findings demonstrate the feasibility of using the MBS to establish an accurate NSCLC patient genotype. This work will help understand the molecular basis of EGFR compound mutations in NSCLC, and could aid the development of new treatment modalities.

Activation of AXL as a Preclinical Acquired Resistance Mechanism Against Osimertinib Treatment in EGFR-Mutant Non-Small Cell Lung Cancer Cells.

Osimertinib (AZD9291) has an efficacy superior to that of standard EGFR-tyrosine kinase inhibitors for the first-line treatment of patients with EGFR-mutant advanced non-small cell lung cancer (NSCLC). However, patients treated with osimertinib eventually acquire drug resistance, and novel therapeutic strategies to overcome acquired resistance are needed. In clinical or preclinical models, several mechanisms of acquired resistance to osimertinib have been elucidated. However, the acquired resistance mechanisms when osimertinib is initially used for EGFR-mutant NSCLC remain unclear. In this study, we experimentally established acquired osimertinib-resistant cell lines from EGFR-mutant NSCLC cell lines and investigated the molecular profiles of resistant cells to uncover the mechanisms of acquired resistance. Various resistance mechanisms were identified, including the acquisition of MET amplification, EMT induction, and the upregulation of AXL. Using targeted next-generation sequencing with a multigene panel, no secondary mutations were detected in our resistant cell lines. Among three MET-amplified cell lines, one cell line was sensitive to a combination of osimertinib and crizotinib. Acquired resistance cell lines derived from H1975 harboring the T790M mutation showed AXL upregulation, and the cell growth of these cell lines was suppressed by a combination of osimertinib and cabozantinib, an inhibitor of multiple tyrosine kinases including AXL, both in vitro and in vivo. Our results suggest that AXL might be a therapeutic target for overcoming acquired resistance to osimertinib. IMPLICATIONS: Upregulation of AXL is one of the mechanisms of acquired resistance to osimertinib, and combination of osimertinib and cabozantinib might be a key treatment for overcoming osimertinib resistance.

exSSSRs (extracellular S100 soil sensor receptors)-Fc fusion proteins work as prominent decoys to S100A8/A9-induced lung tropic cancer metastasis.

Within the "seed and soil" theory of organ tropic cancer metastasis is a growing compilation of evidence that S100A8/A9 functions as a soil signal that attracts cancer cells to certain organs, which prove beneficial to their growth. S100A8/A9-sensing receptors including Toll-like receptor 4 (TLR4), advanced glycation end products (RAGE), and also important receptors we recently succeeded in identifying (EMMPRIN, NPTNß, MCAM, and ALCAM) have the potential to become promising therapeutic targets. In our study, we prepared extracellular regions of these novel molecules and fused them to human IgG2-Fc to extend half-life expectancy, and we evaluated the anti-metastatic effects of the purified decoy proteins on metastatic cancer cells. The purified proteins markedly suppressed S100A8/A9-mediated lung tropic cancer metastasis. We hence expect that our novel biologics may become a prominent medicine to prevent cancer metastasis in clinical settings through cutting the linkage between "seed and soil".

Newly developed anti-S100A8/A9 monoclonal antibody efficiently prevents lung tropic cancer metastasis.

The metastatic dissemination of cancer cells to remote areas of the body is the most problematic aspect in cancer patients. Among cancers, melanomas are notoriously difficult to treat due to their significantly high metastatic potential even during early stages. Hence, the establishment of advanced therapeutic approaches to regulate metastasis is required to overcome the melanoma disease. An accumulating mass of evidence has indicated a critical role of extracellular S100A8/A9 in melanoma distant metastasis. Lung S100A8/A9 is induced by melanoma cells from distant organs and it attracts these cells to its enriched lung environment since melanoma cells possess several receptors that sense the S100A8/A9 ligand. We hence aimed to develop a neutralizing antibody against S100A8/A9 that would efficiently block melanoma lung metastasis. Our protocol provided us with one prominent antibody, Ab45 that efficiently suppressed not only S100A8/A9-mediated melanoma mobility but also lung tropic melanoma metastasis in a mouse model. This prompted us to make chimeric Ab45, a chimera antibody consisting of mouse Ab45-Fab and human IgG2-Fc. Chimeric Ab45 also showed significant inhibition of the lung metastasis of melanoma. From these results, we have high hopes that the newly produced antibody will become a potential biological tool to block melanoma metastasis in future clinical settings.