Anaplastic Lymphoma Receptor Tyrosine Kinase Research Articles

Abstract C116: Oncogenic Kras signaling shapes the tumor microenvironment in lung adenocarcinoma

Abstract Despite advancements in targeted therapy and immunotherapy, lung cancer remains the leading cause of cancer-related deaths in both men and women worldwide, with an estimated 2.2 million deaths each year. At 85%, non-small cell lung cancer (NSCLC) is by far the most common subtype, comprising adenocarcinomas and lung squamous cell carcinoma. Mutations in Kirsten rat sarcoma viral oncogene homolog (KRAS), epidermal growth factor receptor (EGFR) and anaplastic lymphoma receptor tyrosine kinase (ALK) genes are common, with the worst overall survival in KRAS mutant adenocarcinoma patients. About 40% of lung tumors with KRAS mutations also harbor mutations in tumor protein p53 gene (TP53), a tumor suppressor gene. KrasG12C and KrasG12D are both prevalent mutations in lung cancer. Sotorasib, a KrasG12C inhibitor, was recently approved for treatment of NSCLC. However, only 37% of patients have an objective response to Sotorasib, with a median duration of just 11 months. The KrasG12D inhibitor MRTX1133 is entering clinical trials; elucidating the mechanism of action and potential resistance is imperative to design combination treatment approaches. While cell-intrinsic mechanisms of resistance in epithelial oncogenic KRASG12D (Kras*) are well studied, our group and others have shown that KRASG12D also extrinsically regulates the tumor microenvironment (TME) in pancreatic cancer. Whether and how this oncogene regulates the TME of lung cancer is less understood. To study the crosstalk between epithelial lung cells expressing oncogenic KRAS and the surrounding TME, we have established a triple transgenic murine model of Kras*-driven NSCLC, wherein both lung epithelial-specific expression of Kras* and expression of mutant TP53R172H (Tp53*) can be controlled in an inducible and reversible manner. Our preliminary data show that expression of Kras* drives tumor growth, while Kras* depletion results in regression, even in the presence of Tp53*. Here we interrogate how Kras* modulates the TME in lung adenocarcinomas. Harnessing scRNA sequencing, immunohistochemistry, and multiple biochemical approaches, we will provide insight into changes in stromal and immune contexture upon Kras* depletion in mice. Citation Format: Rachael Baliira, Emily Lasse Opsahl, Ivana Barravecchia, Marina Pasca di Magliano, Stefanie Galban. Oncogenic Kras signaling shapes the tumor microenvironment in lung adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr C116.

Downregulation of AP1S1 causes the lysosomal degradation of EGFR in non-small cell lung cancer.

Matrix stiffness has been shown to play a critical role in cancer progression by influencing various cellular processes, including epidermal growth factor (EGF) signaling. However, the underlying molecular mechanisms are not fully understood. Here, we investigated the role of adaptor-related protein complex 1 subunit sigma 1 (AP1S1), a component of adaptor protein complex-1, in the regulation of EGF receptor (EGFR) intracellular trafficking during cancer cell progression. We found that AP1S1 expression was upregulated under stiff matrix conditions, resulting in the regulation of EGFR trafficking in non-small cell lung adenocarcinoma cells. Knockout of AP1S1 caused the lysosomal degradation of EGFR, leading to suppressed EGF-induced anaplastic lymphoma receptor tyrosine kinase phosphorylation. In addition, the downregulation of AP1S1 increased the sensitivity of H1975 cancer cells, which are resistant to tyrosine kinase inhibitors, to erlotinib. Collectively, our results suggest that AP1S1 could regulate EGFR recycling under stiff matrix conditions, and AP1S1 inhibition could be a novel strategy for treating cancer cells resistant to EGFR-targeted anticancer drugs.

Correlation between ALK, ROS1 Biomarkers and EGFR Oncogene Mutations in Lung Tumours: Our Observations in an Apex Oncopathology Laboratory

Introduction: Analysis of Anaplastic lymphoma receptor tyrosine kinase gene (ALK), Repressor of Silencing 1 (ROS1) gene are determined by immunohistochemistry (IHC) and it is an easily applicable, cost-effective assay for potential treatment with crizotinib. Mutations of Epidermal growth factor receptor (EGFR) genes are evaluated by IHC/Multiplex RT-PCR. The purpose of this study is to assess the frequencies of ALK, ROS1 and their association with EGFR fusion gene mutations in a spectrum of lung tumours. Materials and methods: A total of 202 cases of lung tumours reported at our Center for Oncopathology from September 1st 2020 to 31st August 2021, were retrospectively analyzed for ALK, ROS1 and EGFR fusion genes based on Immunohistochemistry (IHC) and Multiplex PCR findings. ALK was tested using D5F3 clone, and ROS1 was analyzed using Cell Signalling’s D4D6 clone on the Ventana immunohistochemistry platform. EGFR status was analyzed using EGFR mutation test V2 real-time multiplex PCR assay on Roche Cobas Z480. Results: 202 biopsy samples and cellblocks of fluid aspirates were analyzed. 175/202 were histologically and immunologically proved as Non-small cell lung carcinoma (Primary pulmonary adenocarcinoma) and its metastases. 09/199 (5.23%) were Positive for ALK IHC and 03/199 (1.74%) cases had equivocal results. 06/179 (3.85%) cases were Positive for ROS1 IHC and 03/179 (1.92%) cases had equivocal results. Other histo-morphological diagnoses i.e., adenosquamous, squamous, small cell, mucinous carcinoma etc (27 cases) were all ALK and ROS1 Negative. 188/202 tumours were analyzed for EGFR mutation status, which showed 70/188 (37.23%) had specific EGFR mutations. 118/188 (62.76%) cases were EGFR wildtype. Conclusion: We observed that age related incidence of EGFR mutations was highest in elderly females, of 61 to 70 years. ALK gene mutations occurred in 6.03% and ROS1 gene mutations occurred in 5.02% of lung tumours and their metastases. EGFR-mutations were associated with ROS1 mutated lung adenocarcinomas. There are no coexistent ALK-EGFR or ALK-ROS1 mutations. All ALK IHC positive pulmonary adenocarcinomas are ROS1 negative and are mutually exclusive.

Abstract PR07: Oncogenic Kras signaling shapes the tumor microenvironment in lung adenocarcinoma

Abstract Despite advancements in targeted therapy and immunotherapy, lung cancer remains the leading cause of cancer-related deaths in both men and women worldwide, with an estimated 2.2 million deaths each year. At 85%, non-small cell lung cancer (NSCLC) is by far the most common subtype, comprising adenocarcinomas and lung squamous cell carcinoma. Mutations in Kirsten rat sarcoma viral oncogene homolog (KRAS), epidermal growth factor receptor (EGFR) and anaplastic lymphoma receptor tyrosine kinase (ALK) genes are common, with the worst overall survival in KRAS mutant adenocarcinoma patients. About 40% of lung tumors with KRAS mutations also harbor mutations in tumor protein p53 gene (TP53), a tumor suppressor gene. KrasG12C and KrasG12D are both prevalent mutations in lung cancer. Sotorasib, a KrasG12C inhibitor, was recently approved for treatment of NSCLC. However, only 37% of patients have an objective response to Sotorasib, with a median duration of just 11 months. The KrasG12D inhibitor MRTX1133 is entering clinical trials; elucidating the mechanism of action for potential resistance is imperative to design combination treatment approaches. While cell-intrinsic mechanisms of resistance in epithelial oncogenic KRASG12D (Kras*) are well studied, our group and others have shown that KRASG12D also extrinsically regulates the tumor microenvironment (TME) in pancreatic cancer. Whether and how this oncogene regulates the TME of lung cancer is less understood. To study the crosstalk between epithelial lung cells expressing oncogenic KRAS and the surrounding TME, we have established a triple transgenic murine model of Kras*-driven NSCLC, wherein both lung epithelial-specific expression of Kras* and expression of mutant TP53R172H (Tp53*) can be controlled in an inducible and reversible manner. Our preliminary data show that expression of Kras* drives tumor growth, while Kras* depletion results in regression, even in the presence of Tp53*. Here we interrogate how Kras* modulates the TME in lung adenocarcinomas. Harnessing scRNA sequencing, immunohistochemistry, and multiple biochemical approaches, we will provide insight into changes in stromal and immune contexture upon Kras* depletion in mice. Citation Format: Rachael K. Baliira, Emily Lasse Opsahl, Elyse McLintock, Katelyn L. Donahue, Ivana Barravacchia, Marina Pasca di Magliano, Stefanie Galbán. Oncogenic Kras signaling shapes the tumor microenvironment in lung adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference: Targeting RAS; 2023 Mar 5-8; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Res 2023;21(5_Suppl):Abstract nr PR07.

A kinase inhibitor screen reveals MEK1/2 as a novel therapeutic target to antagonize IGF1R-mediated antiestrogen resistance in ERα-positive luminal breast cancer

Antiestrogen resistance of breast cancer has been related to enhanced growth factor receptor expression and activation. We have previously shown that ectopic expression and subsequent activation of the insulin-like growth factor-1 receptor (IGF1R) or the epidermal growth factor receptor (EGFR) in MCF7 or T47D breast cancer cells results in antiestrogen resistance. In order to identify novel therapeutic targets to prevent this antiestrogen resistance, we performed kinase inhibitor screens with 273 different inhibitors in MCF7 cells overexpressing IGF1R or EGFR. Kinase inhibitors that antagonized antiestrogen resistance but are not directly involved in IGF1R or EGFR signaling were prioritized for further analyses. Various ALK (anaplastic lymphoma receptor tyrosine kinase) inhibitors inhibited cell proliferation in IGF1R expressing cells under normal and antiestrogen resistance conditions by preventing IGF1R activation and subsequent downstream signaling; the ALK inhibitors did not affect EGFR signaling. On the other hand, MEK (mitogen-activated protein kinase kinase)1/2 inhibitors, including PD0325901, selumetinib, trametinib and TAK-733, selectively antagonized IGF1R signaling-mediated antiestrogen resistance but did not affect cell proliferation under normal growth conditions. RNAseq analysis revealed that MEK inhibitors PD0325901 and selumetinib drastically altered cell cycle progression and cell migration networks under IGF1R signaling-mediated antiestrogen resistance. In a group of 219 patients with metastasized ER + breast cancer, strong pMEK staining showed a significant correlation with no clinical benefit of first-line tamoxifen treatment. We propose a critical role for MEK activation in IGF1R signaling-mediated antiestrogen resistance and anticipate that dual-targeted therapy with a MEK inhibitor and antiestrogen could improve treatment outcome.

A narrative review of predictive immuno-histo/cyto chemistry in lung cancer: focus on gene fusions and PD-L1 expression

Background and Objective: Personalized medicine has significant modified the way advanced stage non-small cell lung cancer (NSCLC) patients have been managed. In fact, the development of different target drugs has significantly improved the clinical outcomes of these patients. In this scenario, gene rearrangements play a crucial role. Gene fusions, including those involving anaplastic lymphoma receptor tyrosine kinase (ALK), ROS proto-oncogene 1, receptor tyrosine kinase (ROS1), rearranged during transfection (RET) and neurotrophic receptor tyrosine kinase (NTRK) genes, occur very rarely in advanced stage NSCLC patients. Remarkably, these genomic alterations represent an important target for treatment decision algorithm in these patients. In addition, programmed death-ligand 1 (PD-L1) protein expression evaluation is crucial for immune-checkpoint inhibitors (ICIs) administration.

Limited airway resection and reconstruction for paediatric tracheobronchial inflammatory myofibroblastic tumour.

OBJECTIVESThe paediatric tracheobronchial inflammatory myofibroblastic tumour (IMT) is a rare disease. Whether limited surgical resection is a feasible surgical approach for these patients remains controversial. The objectives of this study were to report the long-term prognosis after limited surgical resections on paediatric tracheobronchial IMT and provide a surgical management strategy for this rare disease.METHODSPaediatric tracheobronchial IMT patients who underwent limited surgical resection from 2012 to 2020 were enrolled in this study. The clinical characteristics, course of treatment and long-term outcomes of all participants were collated. We presented the accumulated data and analysed the feasibility of limited surgical resection on the paediatric tracheobronchial IMT.RESULTSA total of 9 children with tracheobronchial IMTs were enrolled in our study. Cough and shortness of breath were the most common symptoms. All 9 participants underwent surgical treatment, including 2 tracheal reconstructions, 4 carinal reconstructions and 3 bronchial sleeve resections. Among the participants, 6/9 (66%) were positive for the anaplastic lymphoma receptor tyrosine kinase gene in terms of immunohistochemistry. None of the participants died of short-term complications. The follow-up period was 5.4 (range, 1.1–9.3) years, during which all participants remained well.CONCLUSIONSLimited surgical resection is preferred for paediatrics with tracheobronchial IMTs. Meanwhile, patients with complete resection have an excellent long-term prognosis.

First report of an adult female patient with endobronchial inflammatory myofibroblastic tumor in Taiwan: A case report

An inflammatory myofibroblastic tumor (IMT) of the respiratory system is an uncommon disease. In Taiwan, there is a lack of previous studies on tracheobronchial IMT. The tumor is characterized by overexpression of anaplastic lymphoma receptor tyrosine kinase (ALK)‐1. Surgical resection is the standard treatment of choice nowadays.

Therapeutic Targeting of the Anaplastic Lymphoma Kinase (ALK) in Neuroblastoma-A Comprehensive Update.

Neuroblastoma (NBL) is an embryonic malignancy of the sympathetic nervous system and mostly affects children under the age of five. NBL is highly heterogeneous and ranges from spontaneously regressing to highly aggressive disease. One of the risk factors for poor prognosis are aberrations in the receptor tyrosine kinase anaplastic lymphoma kinase (ALK), which is involved in the normal development and function of the nervous system. ALK mutations lead to constitutive activation of ALK and its downstream signalling pathways, thus driving tumorigenesis. A wide range of steric ALK inhibitors has been synthesized, and several of these inhibitors are already in clinical use. Major challenges are acquired drug resistance to steric inhibitors and pathway evasion strategies of cancer cells upon targeted therapy. This review will give a comprehensive overview on ALK inhibitors in clinical use in high-risk NBL and on the potential and limitations of novel inhibitors. Because combinatory treatment regimens are probably less likely to induce drug resistance, a special focus will be on the combination of ALK inhibitors with drugs that either target downstream signalling pathways or that affect the survival and proliferation of cancer cells in general.

A novel break site of EML4-ALK report and a rare PRKAR1A-ALK report analyzed by different ALK detection platforms in non-small cell lung cancer patients.

Backgrounddetection of anaplastic lymphoma receptor tyrosine kinase gene (ALK) rearrangements in patients with non‐small‐cell lung cancer (NSCLC) has become a routine pathological diagnosis worldwide.Methodsthere are three major conventional diagnostic methods for ALK fusions: fluorescent in situ hybridization (FISH); immunohistochemistry (Ventana IHC (D5F3)); and polymerase chain reaction (PCR). Next‐generation sequencing (NGS) technology as is a new tool for ALK status detection with great potential. These four methods are highly consistent in detecting ALK status (coincidence rate >96%). However, discrepancies in ALK status have been found in some patients among these methods, which causes confusion for clinicians.Results and conclusionin this study, we analyzed two patients whose ALK statuses were not consistent using these four methods. We explored the potential reasons for deviation of the test results and found a novel EML4‐ALK break site, which had been not described previously.

Characterization of CD66b and its relationship between immune checkpoints and their synergistic impact in the prognosis of surgically resected lung adenocarcinoma

ObjectivesCD66b positive tumor-infiltrating neutrophils (TINs) are key immunity cells in the tumor microenvironment (TME). However, their relationship with clinicopathological features, immune checkpoints (ICs), and prognostic value remains undetermined in lung adenocarcinoma (LUAD). In this study, we aimed to characterize the infiltration by TINs and the prognostic significance in patients with surgically resected LUAD. Materials and methodsExpression of CD66b and ICs, including PD-L1, PD-1, CTLA4, LAG3, TIM3, TIGIT, VISTA, and BTLA, in both cancer cell and tumor-infiltrating lymphocytes (TILs) were estimated by immunohistochemistry in resected LUAD. The associations between CD66b expression and clinicopathological characteristics in patient prognoses were analyzed. We also verified results in another cohort from 85 patients with untreated LUAD and further analyzed the correlation between CD66b expression and EGFR and KRAS mutation status in addition to the rearrangement of the anaplastic lymphoma receptor tyrosine kinase gene (ALK). ResultsA total of 240 patients were included in this study. CD66b expression was observed in 87 (36.2%) samples. ICs including PD-L1, PD-1, CTLA4, LAG3, TIM3, TIGIT, VISTA, and BTLA were observed in percentages that ranged from 23.8% to 59.4%. Positive CD66b expression significantly correlated with smoking history (p = 0.029), pathological stage (p = 0.040), and the positive expression of LAG-3 (p < 0.001), PD-1 (p = 0.008), CTLA-4 (p = 0.013), TIM-3 (p = 0.025), TIGIT (p = 0.002), PD-L1 in TILs (p = 0.015), and PD-L1 in tumor cells (p = 0.010). CD66b positivity was significantly associated with worse recurrence-free survival (RFS) (hazard ratio, HR, 1.687; 95% confidence interval, CI, 1.058–2.690, p = 0.028) and overall survival (OS) (HR, 1.667; 95% CI, 1.097–2.534, p = 0.017). Subgroup analysis revealed that the CD66b+/LAG-3 + group had the worst RFS (5-year rate: 39.5%,) and OS (5-year rate: 53.7%,), while the CD66b−/LAG-3 − group had the best RFS (5-year rate: 65.6%) and OS (5-year rate: 78.8%). The p value in analysis of RFS and OS was 0.005 and 0.008, respectively. In the verification set, high expression of CD66b was also significantly correlated with the positive expression of LAG-3 (p < 0.001), PD-1 (p = 0.002), CTLA-4 (p = 0.034), TIM-3 (p = 0.049), PD-L1 in TILs (p = 0.003), and PD-L1 in tumor cells (p = 0.045). There was no correlation between CD66b expression and positive TIGIT expression (p = 0.077), EGFR mutation (p = 0.223), KRAS mutation (p = 0.151), and ALK fusion (p = 0.310). ConclusionCD66b had a relatively high positive expression rate and special clinicopathological features in patients with LUAD. CD66b + TINs were related to the expression of ICs and associated with poor prognoses in LUAD. A combination of CD66b and ICs, especially LAG-3 could further stratify patients into different groups with distinct prognoses.

SPP1 overexpression is associated with poor outcomes in ALK fusion lung cancer patients without receiving targeted therapy

The screening of non-small cell lung cancer (NSCLC) tumors for anaplastic lymphoma receptor tyrosine kinase (ALK) gene rearrangements is important because of the dramatically favorable therapy response to ALK inhibitor. However, the exact mechanism of poor survival in ALK fusion lung cancer patients without receiving targeted therapy is unclear. In this study, total of 521 tumor specimens from Chinese patients with lung cancer were screened for ALK fusion by immunohistochemistry (IHC) and confirmed by fluorescence in situ hybridization (FISH). As results, there were no cases of coexisting EGFR and ALK mutations identified. Fourteen cases (2.7%) harbored ALK fusion, including eight solid adenocarcinomas with signet ring cell features, four acinar adenocarcinomas with cribriform pattern containing mucin, one adenosquamous carcinoma and one micropapillary adenocarcinoma with mucin. Six (42.9%) of fourteen patients with ALK-positive lung cancer had stage IV disease, and five ALK-positive patients treated with platinum-based chemotherapy had poor outcome (all patients were dead and the mean survival time was 12 months), compared to 72 months for patients with ALK inhibitor therapy. Furthermore, Five ALK-positive cases were analyzed by whole exome sequencing (WES) and via direct transcript counting using a digital probe-base (NanoString) to explore the driver genes. Deregulation of PI3K/AKT signaling pathway in ALK-positive lung cancer was demonstrated by WES analysis, and significantly increased mRNA of ALK, ROS1, MET, SPP1 and PI3K signaling pathway was identified by NanoString assay. The concordance between NanoString, IHC and FISH methodologies for detecting ALK fusion was 100%. Significant overexpression of SPP1 protein in ALK-positive lung cancer was confirmed by IHC compared to paired adjacent normal tissues and ALK-negative cancers. Thus we concluded that SPP1 overexpression is associated with poor outcomes for patients with ALK fusion lung cancer without receiving targeted therapy and PI3K/AKT/SPP1 pathway may become the promising targets in patients with aggressive lung cancer.

Disparate Mutations in B-Acute Lymphoblastic Leukemia Among African American and European American Children

Background: B-acute lymphoblastic leukemia (B-ALL) is considered to be the most common malignancy diagnosed in childhood and leading cause of cancer-related death in children. There have been advances in the efficacy of multi-agent chemotherapy and stratification of treatment intensity according to clinical features of the patient, biologic features of leukemia cells, and early response to treatment. These advances herald the promise and challenges of precision medicine strategies that integrate leukemia genomics into contemporary therapy. Despite this progress, there are unmet needs related to the treatment of B-ALL. Among them the incidence varies according to race: African American (AA) children have significantly lower incidence of B-ALL compared to European American (EA). However, studies have shown that AA children have significantly lower survival rates compared to EA children. This disparity is not related to socioeconomic variables, suggesting a molecular basis for the lower survival rates of AA. Here we present a study showing race-specific genetic aberrations involved in leukemogenesis that may contribute to health disparities in B-ALL in AA and EA children.Aims: In a population of AA and EA pediatric B-ALL patients: 1) Discover the genomic susceptibility architecture at validated loci and determine whether genetic variants confer population-specific risks. 2) Define the landscape of molecular perturbation and genomic alterations. 3) Identify aberrant canonical pathways in leukemogenesis.Methods: A total of 20 newly diagnosed pediatric patients were utilized in our study (5 AA and 15 EA). Ages ranged between 1 and 18 years. Clinical history, hemogram, flow cytometry, immunophenotypic, morphologic, cytogenetic and prior molecular data were reviewed. None of the patients in our study had a relapse. Median percent cellularity present in the bone marrow aspirates was 100% (70% - 100%). Median percent of blasts was 94.8% (64.5% - 99.0%). Following manufacturer protocol (Qiagen, QIAmp DNA blood kit), 200µl of frozen bone marrow aspirates were used to extract the DNA. Following quality control on a Qubit fluorometer and RNA bioanalyzer, the samples were submitted to a translational genomics core for library preparation and whole-exome sequencing (TruSeq Exome, Illumina). Bioinformatics analysis was performed and genetic variants for each sample were identified.Results: We identified specific germ-line mutations within the most widely accepted cancer-related genes related to B-ALL. Most genetic aberrations (339) were shared between AA and EA, for example those present in the Anaplastic Lymphoma Receptor Tyrosine Kinase (ALK) gene. Some genes with genetic aberrations were specific of AA (58) such as, Lipoma Preferred Partner Gene (LPP) and others specific for EA (52) such as, Leukemia Inhibitory Factor Receptor (LIFR). The ingenuity pathway analysis revealed these genes clustered in race-specific canonical pathways. In AA, the specific pathways were related to telomerase signaling and cancer signaling. In EA, the specific pathways were related to stem cell pluripotency and hereditary cancers. Our findings suggest the value of whole exome sequencing as a tool for development of individual gene signatures and gene scores for AA and EA children afflicted by B-ALL.Conclusions: Overall, the prevalence of genetic aberrations in B-ALL may contribute to differences in incidences and outcomes between AA and EA children. Aberrant biological networks revealed by our study, provide information on distinguishing genetic aberrations and signaling networks that may be involved in race-specific leukemogenesis. Furthermore, our findings suggest that it may be possible to develop a whole-exome sequencing gene signature in B-ALL to help define a race-specific prognostic assay for B-ALL in bone marrow aspirates. In the future, the biomarker panel derived from this study will help guide risk stratification and treatments based on race and advance the prognosis of B-ALL in AA children. These findings may ultimately impact targeted disease management and contribute to the elimination of the disparate outcomes in B-ALL among AA children.Funding: Hyundai Hope on Wheels Scholar Grant, UMMC Division of Pediatric Hematology Oncology [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Binge-like ethanol drinking activates anaplastic lymphoma kinase signaling and increases the expression of STAT3 target genes in the mouse hippocampus and prefrontal cortex.

Alcohol use disorder (AUD) has a complex pathogenesis, making it a difficult disorder to treat. Identifying relevant signaling pathways in the brain may be useful for finding new pharmacological targets to treat AUD. The receptor tyrosine kinase anaplastic lymphoma kinase (ALK) activates the transcription factor STAT3 in response to ethanol in cell lines. Here, we show ALK activation and upregulation of known STAT3 target genes (Socs3, Gfap and Tnfrsf1a) in the prefrontal cortex (PFC) and ventral hippocampus (HPC) of mice after 4 days of binge-like ethanol drinking. Mice treated with the STAT3 inhibitor stattic drank less ethanol than vehicle-treated mice, demonstrating the behavioral importance of STAT3. To identify novel ethanol-induced target genes downstream of the ALK and STAT3 pathway, we analyzed the NIH LINCS L1000 database for gene signature overlap between ALK inhibitor (alectinib and NVP-TAE684) and STAT3 inhibitor (niclosamide) treatments on cell lines. These genes were then compared with differentially expressed genes in the PFC of mice after binge-like drinking. We found 95 unique gene candidates, out of which 57 had STAT3 binding motifs in their promoters. We further showed by qPCR that expression of the putative STAT3 genes Nr1h2, Smarcc1, Smarca4 and Gpnmb were increased in either the PFC or HPC after binge-like drinking. Together, these results indicate activation of the ALK-STAT3 signaling pathway in the brain after binge-like ethanol consumption, identify putative novel ethanol-responsive STAT3 target genes, and suggest that STAT3 inhibition may be a potential method to reduce binge drinking in humans.

Effect of Asbestos Exposure on the Frequency of EGFR Mutations and ALK/ROS1 Rearrangements in Patients With Lung Adenocarcinoma: A Multicentric Study.

The aim of this study is to investigate the effect of asbestos exposure on cancer-driver mutations. Between January 2014 and September 2018, epidermal growth factor receptor (EGFR), anaplastic lymphoma receptor tyrosine kinase (ALK), and c-ros oncogene 1 receptor tyrosine kinase gene (ROS1) alterations, demographic characteristics, asbestos exposure, and asbestos-related radiological findings of 1904 patients with lung adenocarcinoma were recorded. The frequencies of EGFR mutations, ALK, and ROS1 rearrangements were 14.5%, 3.7%, and 0.9%, respectively. The rates of EGFR mutations and ALK rearrangements were more frequent in asbestos exposed non-smokers (48.7% and 9%, respectively). EGFR mutation rate was correlated to female gender and not-smoking, ALK rearrangement rate was correlated to younger age, not-smoking, and a history of asbestos exposure. The higher rate of ALK rearrangements in asbestos-exposed lung adenocarcinoma cases shows that asbestos exposure may most likely cause genetic alterations that drive pulmonary adenocarcinogenesis.

Abstract 3855: Oncogenic Kras-mediated regulation of the tumor microenvironment in lung cancer

Abstract Lung cancer remains the leading cause of cancer-related deaths worldwide, with an estimated 1.6 million deaths each year. Non-small cell lung cancer (NSCLC) is with 85% by far the most common subtype of lung cancer, comprising adenocarcinomas and lung squamous cell carcinoma. Mutations in Kirsten rat sarcoma viral oncogene homolog (KRAS), epidermal growth factor receptor (EGFR) and anaplastic lymphoma receptor tyrosine kinase (ALK) genes are common with the worst overall survival for KRAS mutant adenocarcinoma patients. We have established a murine model of lung cancer, wherein expression of oncogenic Kras can be controlled genetically, allowing activation of oncogenic KrasG12D (Kras*) to initiate tumor growth, tumor eradication upon Kras* depletion and re-activation as a means to model relapse. Oncogenic Kras depletion (deactivation) has previously been reported to result in tumor cell apoptosis even in the presence of tumor suppressor loss. However, the mechanisms of apoptosis, the role of the immune system on these changes, and the mechanisms allowing some tumor cells to escape apoptosis, which typically results in tumor relapse, are unknown. Here, we interrogated the immune response in mediating tumor regression and relapse using this genetically engineered models. Multiplex immunohistochemistry as well as CyTOF provided insight into the changes in immune contexture upon Kras* depletion in mice haploinsufficient for tumor suppressor p53 or mutant for p53 (R172H). Interestingly, total number of T cells including cytotoxic T cells (CTLs) was elevated in lung tumors from p53 mutant mice supporting findings of heightened immune activation and overall response to immune therapy with an increased mutational burden. Kras* inactivation and thus inhibition of MAPK signaling resulted in an overall decrease in abundance of CTLs and antigen presenting cells (APC) as well as engagement of CTL with tumor cells and APCs indicating a decrease in immune presence likely due to proceeding tumor cell kill and immune recruitment. However, intracellular distance of CTL with tumor cells indicated active tumor cell kill of the CTLs to eradicate remaining tumor cells. In summary, these findings support recent observation of increased immune activation in tumors with higher mutational load as well as changes mediated by inhibition of MAPK signaling which both maybe harnessed for enhancing future immunotherapies. Citation Format: Nina Steele, Kristena Y. Abdelmalak, Sarah F. Ferris, Jennifer M. Lee, Carlos Espinoza, Yaqing Zhang, Sundaresh Ram, Craig Galban, Nithya Ramnath, Timothy L. Frankel, Marina Pasca di Magliano, Stefanie Galbán. Oncogenic Kras-mediated regulation of the tumor microenvironment in lung cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3855.

Metastazavusio nesmulkiųjų ląstelių plaučių vėžio su ROS1 geno translokacija gydymo galimybės tirozino kinazės inhibitoriais

ROS1 geno translokacija nustatoma apie 1–4 proc.nesmulkiųjų ląstelių plaučių vėžio (NSLPV) atvejų. ALK (anaplazinės limfomos receptoriaus tirozino kinazė) ir ROS1 (ROS 1 onkogeno tirozino kinazės receptorius) teigiamo metastazavusio NSLPV atveju rekomenduojamas pirmos eilės gydymas tirozino kinazės inhibitoriais (TKI). Krizotinibas – tai geriamasis pirmosios kartos ALK, ROS1 ir MET receptorių TKI, kuris tapo pirmuoju pirmaeilio ALK atžvilgiu teigiamo NSLPV gydymo standartu klinikinėje praktikoje. PROFILE 1001 tyrimo duomenimis, pacientams, sergantiems metastazavusiu ROS1 teigiamu NSLPV, bendro išgyvenamumo mediana, vartojant krizotinibą, siekia kiek daugiau nei ketverius metus. Svarbu tai, kad krizotinibas saugus vartoti ilgą laiką, o pasitaikantis nepageidaujamas poveikis yra trumpalaikis.

Novel ALK-specific mRNA in situ hybridization assay for non-small-cell lung carcinoma.

BackgroundA recent technical advance in mRNA in situ hybridization (mRNA-ISH) assays provides simultaneous signal amplification and background suppression with a unique probe design that enables single-molecule visualization. We assessed the utility of the mRNA-ISH assay as a diagnostic tool for detecting anaplastic lymphoma receptor tyrosine kinase (ALK) mRNA in non-small-cell lung carcinoma (NSCLC). We compared the mRNA-ISH assay with immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH).MethodsThe study included 279 surgically resected lung adenocarcinomas and 44 transbronchial-biopsied (TBB) adenocarcinomas. mRNA-ISH was conducted using the RNAscope 2.0 system, which includes pre-designed probes for detecting the tyrosine kinase domain encoded in ALK mRNA. IHC was conducted on all 323 samples using ALK-specific antibodies. mRNA-ISH was performed on 279 surgical samples and 6 TBB samples. Break-apart FISH was used to examine samples that were mRNA-ISH-positive or IHC-positive.ResultsALK protein expression was detected in 11 of 279 specimens (3.9%). ALK mRNA was also detected with mRNA-ISH in ALK-positive samples, and 9 of the 11 specimens (81%) were also positive for ALK using break-apart FISH. Using the IHC results as a reference, the sensitivity and specificity of mRNA-ISH was 100%. In the TBB cohort, ALK protein expression was observed in 3 of 44 specimens (6.8%), in which ALK mRNA expression was also detected.ConclusionsThe ALK mRNA-ISH data were highly correlated with the IHC data, and ALK mRNA-ISH detected ALK mRNA expression in every FISH-positive sample. We conclude that mRNA-ISH could serve as an alternative or complementary method for diagnosing ALK rearrangements in NSCLC.

Histopathological Type of Lung Cancer and Underlying Driver Mutations in Patients with Chronic Obstructive Pulmonary Disease (COPD) versus Patients with Asthma and COPD Overlap: A Single-Center Retrospective Study.

Chronic obstructive pulmonary disease (COPD) increases the risk of lung cancer. The relationships between COPD and Asthma COPD Overlap (ACO), and between the histopathological types of lung cancer and driver mutations remain unclear and need further study. The aim of this retrospective study was to examine the relationships between the histopathological type, frequency of epidermal growth factor receptor (EGFR) driver mutations, and anaplastic lymphoma receptor tyrosine kinase (ALK) rearrangements in the lung cancers of patients with COPD and ACO. Patients with pure COPD (n=198) or ACO (n=318) who were admitted to our hospital were reviewed retrospectively. Lung cancers were identified in 43 (21.7%) patients with pure COPD and 54 (17.0%) patients with ACO. The following lung cancers types were observed: patients with pure COPD had 19 (44.2%) adenocarcinomas, 13 (30.2%) squamous cell lung carcinomas (SCC), 8 (18.6%) small cell lung carcinomas (SCLC); patients with ACO had 23 (42.6%) adenocarcinomas, 23 (42.6%) SCC, 2 (3.70%) SCLC. SCLC was significantly more prevalent in patients with pure COPD (p<0.05) than in those with ACO. Differences between the numbers of other histological types of lung cancer and the numbers of driver mutations in the 2 groups of patients were not significant. The differences in the rate of lung cancer and prevalence of EGFR driver mutations between the patients with pure COPD and those with ACO were not significant.

Nucleophosmin1 (NPM1) abnormality in hematologic malignancies, and therapeutic targeting of mutant NPM1 in acute myeloid leukemia.

Nucleophosmin (NPM1) is an abundant nucleolar protein that is implicated in a variety of biological processes and in the pathogenesis of several human malignancies. For hematologic malignancies, approximately one-third of anaplastic large-cell non-Hodgkin’s lymphomas were found to express a fusion between NPM1 and the catalytic domain of anaplastic lymphoma receptor tyrosine kinase. About 50–60% of acute myeloid leukemia patients with normal karyotype carry NPM1 mutations, which are characterized by cytoplasmic dislocation of the NPM1 protein. Nevertheless, NPM1 is overexpressed in various hematologic and solid tumor malignancies. NPM1 overexpression is considered a prognostic marker of recurrence and progression of cancer. Thus, NPM1 abnormalities play a critical role in several types of hematologic malignancies. This has led to intense interest in the development of an NPM1 targeting strategy for cancer therapy. The aim of this review is to summarize present knowledge on NPM1 origin, pathogenesis, and therapeutic interventions in hematologic malignancies.