914 Loss of LKB1 is associated with resistance to IFN-gamma and T cell killing in non-small cell lung cancer

BackgroundKRAS-mutant non-small cell lung cancers (NSCLC) have exhibited unique response patterns to immunotherapy based on their co-occurring mutations. Patients harboring KRAS & STK11/LKB1 co-mutations (KL) have experienced shorter progression-free and overall survival compared to those with only KRAS mutations (K). Despite their limited responses, KL tumors exhibit a tumor mutational burden comparable to their K counterparts, suggesting the presence of additional mechanisms impairing antigen-specific responses. Accordingly, here we investigated the role of the MHC I antigen processing and presentation pathway in KL tumors.MethodsTCGA lung adenocarcinoma (LUAD) data were investigated for changes in expression of HLA molecules and chaperones involved in antigen processing and presentation. In mice, we performed single cell RNA sequencing of resected LKR13 K and KL tumors to evaluate changes in the tumor microenvironment and intrinsic differences in tumor antigen processing machinery. In vitro experiments were performed using the ovalbumin antigen to evaluate changes in antigen-specific T cell responses.ResultsExpression of HLA-A (p<0.0001), -B (p<0.0001), -C (p<0.0001), and beta2-microglobulin (B2M, p<0.0002) was downregulated in KL tumors from TCGA, as were expression of the TAP1 (p<0.001) and TAP2 (p<0.001) transporter associated with antigen processing subunits. LKR13 KL tumors exhibited similar patterns with lower H2-k1 (p<0.0001), H2-d1 (p<0.0001), B2m (p<0.0001), Tap1 (p<0.0001) and Tap2 (p<0.0001). As a result, LKR13 KL were resistant to recognition (p<0.005) and killing (56.9% K versus 7.8% KL) by OT-I T cells. Decreased expression of IFN-gamma-regulated genes such as PSMB8 (p<0.001), PSMB9 (p<0.0001), PSMB10 (p<001), CIITA (p<0.0001), NLRC5 (p<0.0001), IFNGR1 (p<0.0001), and IFNGR2 (p<0.0001) was also noted in KL tumors. Accordingly, KL tumors were unresponsive to exogenous IFN-gamma stimulation, maintaining repression of surface H2-Kb and resistance to T cell recognition (p<0.05) and killing (12.8% K versus 4% KL). Expression of T cell chemokines and receptors CXCR3 (p<0.0001), CXCL9 (p<0.0001), and CXCL10 (p<0.0001) was also repressed, potentially contributing to the lack of T cell infiltration in KL tumors.ConclusionsKRAS-mutant tumors harboring STK11/LKB1 alterations have an immunosuppressed phenotype and resistance to PD-1/PD-L1 inhibitors. Our findings provide evidence that these alterations are associated with markedly reduced antigen presentation and resistance to T cell killing, responsiveness to IFN-gamma stimulation, and impaired production of T cell chemokines, providing mechanistic insights into this immunosuppressed phenotype that could help guide the development of new therapeutic strategies for enhancing anti-tumor immunity.

Download Full-text

P60.05 Influence of Different Concentrations of Nivolumab on T Cell Activation in Patients with Non Small Cell Lung Cancer: in Vitro Study

Journal of Thoracic Oncology ◽

10.1016/j.jtho.2021.01.964 ◽

2021 ◽

Vol 16 (3) ◽

pp. S542

Author(s):

I. Wieleba ◽

K. Wojas-Krawczyk ◽

P. Krawczyk ◽

I. Chmielewska ◽

J. Milanowski

Keyword(s):

Lung Cancer ◽

T Cell ◽

Small Cell Lung Cancer ◽

T Cell Activation ◽

Cell Lung Cancer ◽

Cell Activation ◽

In Vitro Study ◽

Small Cell ◽

Small Cell Lung

Download Full-text

The Inhibition of Wnt Restrain KRASG12V-Driven Metastasis in Non-Small-Cell Lung Cancer

Cancers ◽

10.3390/cancers12040837 ◽

2020 ◽

Vol 12 (4) ◽

pp. 837

Author(s):

Pei-Shan Hung ◽

Ming-Hung Huang ◽

Yuan-Yeh Kuo ◽

James Chih-Hsin Yang

Keyword(s):

Small Cell Lung Carcinoma ◽

Small Cell ◽

Dominant Negative ◽

Gene Set Enrichment Analysis ◽

Dominant Negative Mutant ◽

Small Cell Lung ◽

Kras Mutations ◽

Cell Lung Carcinoma ◽

Wnt Inhibitor

The KRAS mutations have been an obstacle to identify therapeutic targets in cancer treatment. In this work, we clarified the distinct metastasis pattern of non-small-cell lung carcinoma (NSCLC) induced by KRASG12V/KRASG12D mutations and inhibited the KRASG12V mediated metastasis by Wnt inhibitor. First, we found that KRASG12V induced more aggressive phenotype in vitro and in vivo experiments. The Gene Set Enrichment Analysis (GSEA) results of H838 KRASG12V cells showed a significant negative correlation with RhoA-related signaling. Following this clue, we observed KRASG12D induced higher activation of RhoA and suppressed activation of Wnt/β-catenin in H838KRASG12D cells. The restored activation of Wnt/β-catenin in H838KRASG12D cells could be detected when expression with a dominant-negative mutant of RhoA or treatment with RhoA inhibitor. Furthermore, the Wnt inhibitor abolished the KRASG12V-induced migration. We elucidated the importance of the axis of RhoA/Wnt in regulatory NSCLC metastasis driven by KRAS mutations. Our data indicate that KRASG12V driven NSCLC metastasis is Wnt-dependent and the mechanisms of NSCLC metastasis induced by KRASG12V/KRASG12D is distinct.

Download Full-text

Gene signature of antigen processing and presentation machinery predicts response to checkpoint blockade in non-small cell lung cancer (NSCLC) and melanoma

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2020-000974 ◽

2020 ◽

Vol 8 (2) ◽

pp. e000974

Author(s):

Jeffrey C Thompson ◽

Christiana Davis ◽

Charuhas Deshpande ◽

Wei-Ting Hwang ◽

Seth Jeffries ◽

...

Keyword(s):

Lung Cancer ◽

Small Cell Lung Cancer ◽

Cell Lung Cancer ◽

Antigen Processing ◽

Small Cell ◽

Checkpoint Blockade ◽

Small Cell Lung ◽

Free Survival ◽

Antigen Processing And Presentation ◽

Inflammation Score

BackgroundLimited data exist on the role of alterations in HLA Class I antigen processing and presentation machinery in mediating response to immune checkpoint blockade (ICB).MethodsThis retrospective cohort study analyzed transcriptional profiles from pre-treatment tumor samples of 51 chemotherapy-refractory advanced non-small cell lung cancer (NSCLC) patients and two independent melanoma cohorts treated with ICB. An antigen processing machinery (APM) score was generated utilizing eight genes associated with APM (B2M, CALR, NLRC5, PSMB9, PSME1, PSME3, RFX5, and HSP90AB1). Associations were made for therapeutic response, progression-free survival (PFS) and overall survival (OS).ResultsIn NSCLC, the APM score was significantly higher in responders compared with non-responders (p=0.0001). An APM score above the median value for the cohort was associated with improved PFS (HR 0.34 (0.18 to 0.64), p=0.001) and OS (HR 0.44 (0.23 to 0.83), p=0.006). The APM score was correlated with an inflammation score based on the established T-cell-inflamed resistance gene expression profile (Pearson’s r=0.58, p<0.0001). However, the APM score better predicted response to ICB relative to the inflammation score with area under a receiving operating characteristics curve of 0.84 and 0.70 for PFS and OS, respectively. In a cohort of 14 high-risk resectable stage III/IV melanoma patients treated with neoadjuvant anti-PD1 ICB, a higher APM score was associated with improved disease-free survival (HR: 0.08 (0.01 to 0.50), p=0.0065). In an additional independent melanoma cohort of 27 metastatic patients treated with ICB, a higher APM score was associated with improved OS (HR 0.29 (0.09 to 0.89), p=0.044).ConclusionOur data demonstrate that defects in antigen presentation may be an important feature in predicting outcomes to ICB in both lung cancer and melanoma.

Download Full-text

E3 ligase MKRN3 is a tumor suppressor regulating PABPC1 ubiquitination in non–small cell lung cancer

Journal of Experimental Medicine ◽

10.1084/jem.20210151 ◽

2021 ◽

Vol 218 (8) ◽

Author(s):

Ke Li ◽

Xufen Zheng ◽

Hua Tang ◽

Yuan-Sheng Zang ◽

Chunling Zeng ◽

...

Keyword(s):

Lung Cancer ◽

Tumor Suppressor ◽

E3 Ligase ◽

Central Precocious Puberty ◽

Small Cell ◽

Missense Mutations ◽

Small Cell Lung ◽

Kras Mutations

Central precocious puberty (CPP), largely caused by germline mutations in the MKRN3 gene, has been epidemiologically linked to cancers. MKRN3 is frequently mutated in non–small cell lung cancers (NSCLCs) with five cohorts. Genomic MKRN3 aberrations are significantly enriched in NSCLC samples harboring oncogenic KRAS mutations. Low MKRN3 expression levels correlate with poor patient survival. Reconstitution of MKRN3 in MKRN3-inactivated NSCLC cells directly abrogates in vitro and in vivo tumor growth and proliferation. MKRN3 knockout mice are susceptible to urethane-induced lung cancer, and lung cell–specific knockout of endogenous MKRN3 accelerates NSCLC tumorigenesis in vivo. A mass spectrometry–based proteomics screen identified PABPC1 as a major substrate for MKRN3. The tumor suppressor function of MKRN3 is dependent on its E3 ligase activity, and MKRN3 missense mutations identified in patients substantially compromise MKRN3-mediated PABPC1 ubiquitination. Furthermore, MKRN3 modulates cell proliferation through PABPC1 nonproteolytic ubiquitination and subsequently, PABPC1-mediated global protein synthesis. Our integrated approaches demonstrate that the CPP-associated gene MKRN3 is a tumor suppressor.

Download Full-text

N6-methyladenosine-modified circIGF2BP3 inhibits CD8+ T-cell responses to facilitate tumor immune evasion by promoting the deubiquitination of PD-L1 in non-small cell lung cancer

Molecular Cancer ◽

10.1186/s12943-021-01398-4 ◽

2021 ◽

Vol 20 (1) ◽

Author(s):

Zhenchuan Liu ◽

Tingting Wang ◽

Yunlang She ◽

Kaiqing Wu ◽

Shaorui Gu ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Mouse Model ◽

Lung Carcinoma ◽

Immune Evasion ◽

Antitumor Immunity ◽

Immune Escape ◽

Small Cell ◽

Small Cell Lung

Abstract Background An in-depth understanding of immune evasion mechanisms in tumors is crucial to overcome resistance and enable innovative advances in immunotherapy. Circular RNAs (circRNAs) have been implicated in cancer progression. However, much remains unknown regarding whether circRNAs impact immune escape in non-small-cell lung carcinoma (NSCLC). Methods We performed bioinformatics analysis to profile and identify the circRNAs mediating immune evasion in NSCLC. A luciferase reporter assay, RNA immunoprecipitation (RIP), RNA pulldown assays and fluorescence in situ hybridization were performed to identify the interactions among circIGF2BP3, miR-328-3p, miR-3173-5p and plakophilin 3 (PKP3). In vitro T cell-mediated killing assays and in vivo syngeneic mouse models were used to investigate the functional roles of circIGF2BP3 and its downstream target PKP3 in antitumor immunity in NSCLC. The molecular mechanism of PKP3-induced PD-L1 upregulation was explored by immunoprecipitation, RIP, and ubiquitination assays. Results We demonstrated that circIGF2BP3 (hsa_circ_0079587) expression was increased in NSCLC and negatively correlated with CD8+ T cell infiltration. Functionally, elevated circIGF2BP3 inactivated cocultured T cells in vitro and compromised antitumor immunity in an immunocompetent mouse model, and this effect was dependent on CD8+ T cells. Mechanistically, METTL3 mediates the N6-methyladenosine (m6A) modification of circIGF2BP3 and promotes its circularization in a manner dependent on the m6A reader protein YTHDC1. circIGF2BP3 competitively upregulates PKP3 expression by sponging miR-328-3p and miR-3173-5p to compromise the cancer immune response. Furthermore, PKP3 engages with the RNA-binding protein FXR1 to stabilize OTUB1 mRNA, and OTUB1 elevates PD-L1 abundance by facilitating its deubiquitination. Tumor PD-L1 deletion completely blocked the impact of the circIGF2BP3/PKP3 axis on the CD8+ T cell response. The inhibition of circIGF2BP3/PKP3 enhanced the treatment efficacy of anti-PD-1 therapy in a Lewis lung carcinoma mouse model. Collectively, the PKP3/PD-L1 signature and the infiltrating CD8+ T cell status stratified NSCLC patients into different risk groups. Conclusion Our results reveal the function of circIGF2BP3 in causing immune escape from CD8+ T cell-mediated killing through a decrease in PD-L1 ubiquitination and subsequent proteasomal degradation by stabilizing OTUB1 mRNA in a PKP3-dependent manner. This work sheds light on a novel mechanism of PD-L1 regulation in NSCLC and provides a rationale to enhance the efficacy of anti-PD-1 treatment in NSCLC.

Download Full-text

Designing of cytotoxic and helper T cell epitope map provides insights into the highly contagious nature of the pandemic novel coronavirus SARS-CoV-2

Royal Society Open Science ◽

10.1098/rsos.201141 ◽

2020 ◽

Vol 7 (9) ◽

pp. 201141 ◽

Cited By ~ 2

Author(s):

Seema Mishra

Keyword(s):

T Cell ◽

Antigen Processing ◽

Cell Epitope ◽

Computational Tools ◽

Human Contact ◽

Novel Coronavirus ◽

Antigen Processing And Presentation ◽

Clear Information ◽

Ranked List

Novel coronavirus, SARS-CoV-2, has emerged as one of the deadliest pathogens of this century, creating an unprecedented pandemic. Belonging to the betacoronavirus family, it primarily spreads through human contact via symptomatic and asymptomatic transmission. Despite several attempts since it emerged, there is no known treatment in the form of drugs or vaccines. Hence, work on developing a potential multi-subunit vaccine is the need of the hour. In this study, attempts have been made to find globally conserved epitopes from the entire set of SARS-CoV-2 proteins as there is as yet, no clear information on the immunogenicity of these proteins. Using diverse computational tools, a ranked list of probable immunogenic, promiscuous epitopes generated through all the three main stages of antigen processing and presentation pathways has been prioritized. Moreover, several useful insights were gleaned during these analyses. One of the most important insights is that all of the proteins in this pathogen present unique epitopes, so that the targeting of a few specific viral proteins is not likely to result in an effective immune response in humans. Due to the presence of these unique epitopes in all of the SARS-CoV-2 proteins, stronger immune responses generated by T cell hyperactivation may lead to cytokine storm and immunopathology and consequently, remote chances of human survival. These epitopes, after due validation in vitro , may thus need to be presented to the human body in that form of multi-subunit epitope-based vaccine that avoids such immunopathologies.

Download Full-text