bluuebunny/crossref_metadata_2025_split huggingface dataset:
HDAC inhibitor Chidamide, but not romidepsin, synergizes with humanized PD1 antibody to enhance T-cell chemokine expression and augment IFN-gamma response in NK-T-cell related lymphoma: A preclinical study.
Authors: Wen Tingyu , Ma Fei , Liu Peng 
Published: June 2022

e14568 Background: The incidence of NK-T cell lymphoma (NKTL) in Asia is significantly higher than that in European and American countries. The median survival time of EKTL is only 8-12 months, and the 5-year overall survival rate is still less than 30%. New therapies or combinations are warranted. Methods: The murine NKTL cell line EL4 and RMA were used to establish cell line xenograft models in transgenic humanized PD1 mice. Different dosages or sequential HDAC inhibitors were combined with humanized PD1 antibody in vivo to test whether the synergistic effect is existing. RNA-seq, qPCR, and IHC were performed to illustrate the potential mechanisms. Three NTKL patients who were treated with hPD1 and Chidamide in clinical were recorded in this study. Results: Romidepsin combined with hPD1 antibody is superior to hPD1 antibody alone in the treatment of two NKTL CDX models (EL4 and RMA), but it is similar with romidepsin single agent, either in different dosages or sequence. However, Chidamide combined with hPD1 antibody significantly inhibits the tumor growth compared to single agents in these two pre-clinical models. RNA-seq reveals Chidamide synergizes with humanized PD1 antibody to enhance T-cell chemokine expression, augment IFN-gamma response and increase CD8 T cell infiltration. String PPI analysis and ingenuity pathway analysis suggest IFN-gamma is the hub upstream regulator in the combination. On the contrary, the romidepsin combinations decrease the activity of T cell immune responses and the function of immune cell migration. The expression of IFN-gamma gene set signatures is decreased in the romidepsin combination. Three NKTL patients who received PD1 antibody and Chidamide are showed promising efficacy, and the progression-free survival is 26+ months, 8 months, and 9+ months respectively. Conclusions: Different HDAC inhibitor shows different combination effects with immunotherapy. Chidamide combination shows a synergistic effect with immunotherapy by enhancing INF-gamma response, but romidepsin combination antagonizes this effect significantly.
A pan-cancer analysis of the granzyme family based on the TCGA database and single-cell RNA sequencing data.
Authors: Xu Bin , He Junju , Li Jing , Wu Jie , Zhang Huibo , Jin Yan , Yao Yi , Song Qibin 
Published: June 2022

e14580 Background: Human Granzymes (GZMs) are mainly regarded as protective cytotoxic proteases that block the development of cancer. This study aimed to systematically analyze the expression signature, immune infiltration pattern, predictive values of prognosis and immune response, and the biological process associated with GZMs in different cancers. Methods: We estimated the predictive values of GZMs in prognosis and drug sensitivities by Cox regression analysis, immunophenoscore (IPS) algorithm, etc. We identified potential factors associated with the expression of GZMs, involving the somatic mutation and copy number variation of driver genes. We further investigate the distributions of GZMs and their discrepancy in single-cell resolution. The package CellChat and SCENIC were used for cell-cell communications and regulatory network reconstruction respectively. The ssGSEA algorithm was used to calculate the GZMs score for a gene set of five granzymes and the enrichment score (ES) of regulons based on the bulk-seq data. We chose the specific regulons to cluster patients in pan-cancer. Results: In the immunotherapy context, the higher GZMs score was associated with better overall survival (OS) in urothelial carcinoma (HR = 0.31, p< 0.01). In melanoma, the pooled HR value calculated by meta-analysis assay was 0.33 (95%CI: 0.189-0.58). In the single-cell resolution, we found the three patterns of GZMs expression in CD8+T/NK cells. Furthermore, we analyzed the discrepancy among different GZMs subtype cells in functions, cell-cell communications, and cell-type-specific regulons. In CD8+ Effector memory T cells, cluster 1 (GZMB+) and cluster 2 (GZMK+) exhibited different enriched pathways in adhesion, anti-bacteria, chemotaxis, migration, and metabolism. In the cell-cell communications analysis, the IFNG − (IFNGR1+IFNGR2) between CD8+ T cells and malignant cells was only significant in the C1 and C2 clusters. Pan-cancer patients were divided into five regulons clusters based on the regulons. Cluster 5 had the worst OS (HR = 8.49, 95%CI = 6.67-10.81, p< 0.01) compared to cluster 1 which had the best prognosis. Cluster 5 had higher ES of regulons including Myb, Hmgb2, E2f7, E2f8, and lower ES of Egr3. Conclusions: In conclusion, the expressions of GZMs, GZMs score, and the specific regulons identified in this study were associated with the prognosis in pan-cancer patients. GZMs score was a valuable biomarker that can predict the response to immunotherapy. The patients who belong to different regulons subtypes have specific transcription factors, which may be potential targets for the precision medicine of anti-tumor strategies in pan-cancer.
Structure-based prediction of neoantigens paired with T cell receptors on pre-existing CD8+ tumor-infiltrating lymphocytes.
Authors: Lee Bo Ryeong , Kim Sungsik , Kim Sung-min , Park Keunchil , Park Woong-Yang 
Published: June 2022

e14583 Background: Tumor neoantigens have ability to expand tumor-specific T cell immunity, which could be boosted by anti-PD-1 immunotherapy. Therapeutically relevant neoantigens should be presented on patient HLAs and recognized by tumor-reactive T cells. Despite tumor-infiltrating lymphocytes (TILs) have enriched tumor-reactive T cell populations, most tumor-specific TILs exhibit exhausted phenotype and have limitations to in vitro screening. Here, we developed in silico structure-based prediction of neoantigen discovery pipeline by pairing with T cell receptors (TCRs) on pre-existing CD8+ exhausted tumor-infiltrating lymphocytes. Methods: Tumor resections from patients with liver, stomach, colorectal, and lung cancers are subject to whole exome and transcriptome. In addition, TILs from the patients are subject to scRNA/TCRseq to select TCRs of exhausted T cells, which are mostly thought to have tumor-reactivity. Neoantigen epitopes are prioritized from our in silico pipeline including HLA binding and TCR-peptide-HLA structure-based pairing score. To evaluate the immunogenicity of selected neoantigens, the neoantigens are tested for in vitro IFNg ELISPOT assay using peripheral blood mononuclear cells (PBMCs) from the same patients, which sensitively detects antigen-experienced T cells. Results: For 5 patients with T cell-reactive neoantigens, among 23 neoantigens selected by our in silico pipeline and tested in vitro, 10 neoantigens showed positive pre-existing T cell responses by matched patients. Then, we analyzed gene expression profiles of TILs paired in silico with in vitro T cell response-positive or negative neoantigens (pTILs and nTILs, respectively) combined with TILs from 23 solid cancer patients. Interestingly, pTILs were mostly mapped to the clusters which highly expressed markers of exhaustion and cytotoxicity, while nTILs showed random distribution across all the clusters. Conclusions: Neoantigen prediction using structure-based pairing of neoantigens and phenotype-selected TILs showed promising potential to better select therapeutically-relevant cancer vaccines. This data supports further investigation of our in silico pipeline into more patient samples and preclinical studies in mouse tumor models.
Validity of mutations as a cancer chronometer.
Authors: Kawsar Farhan , Safar Ahmad Mazen , Chen Jim Zhongning 
Published: June 2022

e15015 Background: Staging is devised to assess prognosis, and ultimately direct the management of cancer. Though staging systems don't necessarily confirm chronological progression from stage I to IV, there is near consensus that advanced stages are 'later' versions of a given cancer. This is reflected in the philosophy and efforts to identify 'early' cancer in screening programs. Tumor Mutational Burden (TMB) is one measure of the degree of proliferation, thus can be utilized as a molecular clock (Vogelstein), a surrogate for chronology. Since TMB is currently a clinical metric provided to oncologists in routine practice, proper interpretation of TMB is critical. We hypothesized that (i) metastatic disease is a ‘different’, rather than 'later' manifestation of a given cancer. Furthermore, we hypothesized that (ii) proliferation rates would be mirrored by the TMB, and lastly, that (iii) TMB will correlate with clinical virulence. Methods: (i) We mined the publicly-available TCGA database (cbioportal.org) for TMB from 558 samples spanning 5 whole genome & whole exome neoplasm datasets (Breast, Colon, Renal cell, Pancreatic, Cervical cancers). (ii) We compared the TMB in Burkitt's lymphoma, a highly proliferative neoplasm, with follicular lymphoma, a classically quiescent malignancy. (iii) We compared the TMB in skin squamous cell carcinoma (SSCC), a cancer with negligible impact on survival, with Acute Myelogenous Leukemia (AML), a highly lethal malignancy despite aggressive therapy. Results: (i) The 95% Confidence Intervals (CI) surrounding the median TMB were overlapping without statistical significance across ALL stages within EVERY histology examined. (ii) To our surprise, the TMB 95% CI were overlapping between the 2 subsets of B-cell lymphoma examined. (iii) TMB was substantially higher in SSCC (median of ̃800) compared to a mere 8 in AML. Conclusions: Assuming validity of Vogelstein’s molecular clock model, mutational probing does not support the concept that a stage IV malignancy is a chronologically delayed manifestation of a given neoplasm. Rather, we suggest an alternative approach where aggressive/metastatic cancers are inherently 'different' with unique abilities rather than a ‘later’ edition of a monodirectional succession. We also posit that cells that live for years (follicular lymphoma) can accumulate comparable levels of TMB as Burkitt's because the limited life span in Burkitt's (days to weeks) precludes the accumulation of high TMB. Finally, the clinical outcome cannot be discerned from the degree or mutability— which is rationally a predictor of immune susceptibility. It's noteworthy that the observed levels of mutations in SSCC, reflective of the sun mutagenicity, is more than 3-fold of that reported for melanoma (̃274) but this fact is rarely presented in cancer genomic landscapes, and may explain the rarity of metastatic SSCC owing to the immune clearance of metastases unlike the skin surface devoid of such immune access.
Dexamethasone and brain metastasis-infiltrating CD8+ T cells.
Authors: Sudmeier Lisa , Hoang Kimberly B. , Nduom Edjah , Goyal Subir , Liu Yuan , Olson Jeffrey J. , Ramalingam Suresh S. , Ahmed Rafi 
Published: June 2022

e15075 Background: Nearly all patients who develop large and/or symptomatic brain metastases receive corticosteroids such as dexamethasone (Dex) to treat or prevent symptoms like nausea, headaches, or focal neurologic deficits. Dex has immunosuppressive effects such as induction of T cell apoptosis, so it’s possible that this treatment may impair the anti-tumor immune response. To address this question, we performed a secondary analysis of our in-press data describing the phenotype of brain metastasis-infiltrating CD8+ T cells to determine whether the duration of Dex therapy prior to surgical resection correlated with phenotypes we described our cohort. Methods: Fresh brain metastases specimens were weighed, and immune cells were isolated. CD45+ and CD8+ lymphocytes were counted via flow cytometry from 35 and 34 samples, respectively. A subset of samples were analyzed by high-parameter spectral flow cytometry to gain phenotypic information. Dex treatment was later extracted from the electronic medical record, and Spearman’s correlation coefficients (CC) were calculated to evaluate the association between duration of Dex therapy and brain metastasis-infiltrating lymphocyte density or phenotype. Results: Patients were on Dex for a median of 3 days (4mg q6 hours) prior to surgery. On univariate analysis, the CC between CD45+ lymphocytes or CD8+ T cells per gram of tumor and duration of Dex treatment was -0.185 (P=0.288) and -0.122 (P=0.49), respectively. The CC between frequency of PD-1+ CD8+ T cells and duration of Dex was -0.064 (p=0.784, 21 patients). In our previous work, we used the FLOWSOM algorithm to identify distinct populations of CD8+ T cells that were enriched in brain metastases compared to blood. We described two such populations, one with a tissue residence phenotype, and the other with terminally-differentiated phenotype. The CC between frequency of these populations and duration of Dex was 0.043 (p=0.889) and 0.238 (p=0.433), respectively (13 patients). We also tested for correlation between frequency of individual PD-1+ CD8+ T cell populations and duration of Dex treatment (13-18 patients). The CC for Tim3+, CD39+, CTLA4+, CD127+, Tcf-1+, CD28+, and GZMB+ populations of PD-1+ CD8+ T cells were 0.037 (p=0.891), 0.096 (p=0.705), -0.094 (p=0.738), 0.011 (p=0.971), -0.026 (p=0.918), 0.384 (p=0.128), and 0.236 (p=0.358), respectively. Conclusions: Within this cohort of patients who were on Dex for a median of 3 days prior to surgery, duration of pre-surgery Dex did not significantly affect density of brain metastasis-infiltrating CD45+ lymphocytes or CD8+ T cells or frequency of PD-1+ CD8+ T cells.
Development of PLM-102, a novel dual inhibitor of FLT3 and RET as a new therapeutic option for acute myeloid leukemia.
Authors: Kim Myungjin , Oh Su-jin , Kim WooChan , Choi Yong June , Lee Jae-Seon , Lee So-Deok , Park Jin-Hee , Lee Eun Ji , Kang Keon Wook , Jang Soo Yeon , Kim Yong-Chul 
Published: June 2022

e15103 Background: Mutations of the FMS-like tyrosine kinase 3 (FLT3) gene occur in approximately 30% of all acute myeloid leukemia (AML) cases, with the internal tandem duplication (ITD) representing the most common type of FLT3 mutation (FLT3-ITD; approximately 25% of all AML cases). Although several FLT3 inhibitors have been developed, occurrence of secondary TKD mutations of FLT3 such as FLT3/D835Y and FLT3/F691L causes the acquired resistance to the current FLT3 inhibitors and eventually become a key area of unmet medical needs. Here, we have revealed that PLM-102, a novel, orally active FLT3 and RET dual inhibitor, has a potential to overcome the acquired resistance to current FLT3 inhibitors. Methods: 1. Kinase assay- Biochemical assays for FLT3 (WT and D835Y) and RET (WT and Mutants) were performed according to ADP-Glo kinase assay protocol(Promega). 2. Cell proliferation and Apoptosis- Human leukemia cell line MV4-11 and MOLM-14 were purchased from ATCC and DSMZ. Cells were seeded at a density of 2 X 103 cells per well and treated with the indicated concentrations of inhibitors for 72 hours at 37°C. Cell viability was determined by an Alamar Blue assay (Bio-Rad). Caspase-3/7 activity was measured by using the Caspase-Glo 3/7 assay (Promega). 3. Western blot analysis- Immunoblotting using MOLM-14 cells was performed using anti-phospho-FLT3 (Cell Signaling Technology #3461) and anti-FLT3 antibody (Cell Signaling Technology #3462). 4. In vivo mouse models- The MV4-11 and MOLM-14 cells are implanted into the subcutaneous space of the left flank of the mice. Resulting tumors are monitored by calipering twice weekly. Treatment started after randomization when tumor volumes had reached a size of approximately 100-150 mm3. For statistical analysis, analysis of variance (ANOVA) was performed using Prism 9.0 to examine statistical differences. Results: In compared to FDA-approved Gilteritinib, PLM-102 showed stronger sub-nanomolar IC50 values in FLT3 kinases regardless of wild-type, ITD, TKD and ITD/TKD mutants in both kinase- and cell-based assays. PLM-102 inhibited phosphorylation of FLT3 and its downstream signaling pathways, and induced apoptosis as evidenced by PARP-cleavage and caspase-3 activation. Moreover, PLM-102 showed an excellent anti-tumor activity in mouse xenograft models implanted with MV4-11 and MOLM-14 AML cells. Conclusions: Taken together, PLM-102 showing potent anti-cancer activity against various in vitro and in vivo AML models could be developed as a valuable agent overcoming the acquired resistance to the current FLT3 inhibitor.
Clinical pharmacokinetic/pharmacodynamic relationship of DZD2269, a potent and selective A2aR antagonist designed to overcome high levels of adenosine induced immunosuppression.
Authors: Liu Ziyi , Bai Yu , Hu Dongdong , Zhang Lin , Yong Tanyada , Yang Zhenfan 
Published: June 2022

e15106 Background: Extracellular adenosine suppresses immune cells’ function through binding to its receptor A2aR. In the tumor microenvironment, adenosine concentration could be several hundred-fold higher than in blood. DZD2269 was designed to fully block A2aR in a wide range of adenosine concentrations. In preclinical studies, DZD2269 could most effectively restore immune cell function inhibited by high levels of adenosine among A2aR antagonists currently in clinical studies. In mouse syngeneic prostate cancer model, DZD2269 could significantly enhance the antitumor activity of anti-PD1 antibody as well as radiotherapy. Two ongoing clinical studies of DZD2269 in healthy volunteers (PAN-GU 2, NCT04932005) and patients with metastatic castration resistant prostate cancer (PAN-GU 1, NCT04634344) are being conducted to characterize its safety, PK, PD and to assess its preliminary anti-tumor efficacy. Here, we report clinical data from the PAN-GU 2 study. Methods: The phase 1 healthy volunteer study is a double-blind, placebo-controlled study to evaluate safety, PK, and effect on biomarkers. It consists of three parts: single ascending dose (SAD): 5, 10, 20, 40, 80 and 160 mg; multiple ascending dose (MAD): from 10 mg BID for 7 consecutive days; and food effect (FE): fasted, low-fat or high-fat meal condition. A2aR pathway blockade by DZD2269 is evaluated by analyzing pCREB inhibition on whole blood T cells following ex vivo stimulation with 10 µM NECA. Results: In SAD part of the healthy volunteer study, a total of 48 subjects received a single oral dose of DZD2269 (n = 36) from 5 mg to 160 mg or placebo (n = 12). There were 7 TEAEs reported by 4 subjects and no ≥ grade 3 TEAE or SAE. After a single oral dose, the rate of absorption of DZD2269 was moderate (median tmax: 1.0 to 2.75 hr). DZD2269 exhibited low to moderate clearance (averaged value: 16.1 to 45.3 L/h) and extensive volume of distribution (averaged value: 209 to 738 L), resulting in a mean t1/2 of 7 ̃ 10 hr; a trend of increased t1/2 was observed at higher dose levels. Systemic exposure of DZD2269 (AUC and Cmax) increased in an approximately dose-dependent manner. Based on PK/PD analysis, approximately 90% pCREB inhibition could be achieved by a single dose of 80mg for 24 hours. Conclusions: Single oral dose of DZD2269 ranging from 5 mg to 160 mg was safe and well tolerated. Good PK/PD correlation was demonstrated in the study. DZD2269 could effectively reverse high adenosine induced immunosuppression. A phase1/2 study evaluating safety and efficacy of DZD2269 in patients is ongoing. Clinical trial information: NCT04932005.
A potential mechanism of ferroptosis modulation for immunotherapy in LUAD: ceRNAs networks of OTUD6B-AS1 modulate the balance of ubiquitination.
Authors: Yao Lintong , Zhou HaiYu 
Published: June 2022

e15111 Background: Ferroptosis plays a pivotal role in cancer progression and therapeutic response of immunotherapy in lung adenocarcinoma (LUAD). Recent studies on ferroptosis regulation by lncRNAs were conducted sporadically. Our study explored FrlncRNAs in LUAD comprehensively, exploring functional mechanisms and verified the correlation between ferroptosis and ubiquitination. Methods: Data of LUAD samples were download from GEO and TCGA databases. Annotated ferroptosis-related genes were retrieved from FerrDb and GeneCards databases. Ferroptosis-related lncRNA (FrlncRNAs) was identified by correlation analysis. The LASSO analysis and cox regression analysis were performed to filtrate significant prognostic FrlncRNAs, followed with model construction and patient stratification. Tumor immune microenvironment, functional pathways were investigated. Subcellular location and potential competing endogenous RNAs (ceRNAs) mechanisms were predicted by GDCRNATools algorithm. Results: Ferroptosis-related prognostic model constructed by 14 FrlncRNAs signature showed high prognostic efficacy with significant survival difference and robust area under the curve (AUC for 5 years survival: 0.89 in training cohort and 0.87 in validation cohort). Higher proportion of myeloid-derived suppressor cells were detected in high-risk group, of which ferroptotic status were supressed. Ferroptosis-enhanced samples harboured higher expression of immune checkpoints such as PD-L1, CTLA-4, TIM3 and CD47 and immune-recruiting interleukins such as IL2, IL4, IL7. Reasonably, more potential responders to immunotherapy were predicted in ferroptosis-enhanced samples. Considering the subcellular location, potential mechanism of ceRNA network was predicted that OTUD6B-AS1 may compete with mir-144 to regulate or modify NUDCD1 and MARCH7 through miRNA sponge. OTUD6B-AS1 encodes the protein of ovarian tumor domain deubiquitinase 6B, while MARCH7 contributes to a RING domain of membrane bound E3 ubiquitin ligases. GO, KEGG and GSEA pathway analyses focused on ubiquitin-protein transferase activity, polyubiquitination, mTOR, Hippo and NOTCH pathways. Protein-protein interaction identified hub genes in ferroptotic regulation such as NEDD8 and HERC2, which were coding genes of ubiquitin ligase E3 as well. Hence, the balance of ubiquitination and deubiquitination induced by the interplay of ceRNA network could be pivotal process in ferroptosis modulation in LUAD. Conclusions: In summary, we reveal potential mechanisms that OTUD6B-AS1 may compete with mir-144 to regulate the expression NUDCD1 and MARCH7 through miRNA sponge or modifying protein structures, modulating ferroptosis by balance of ubiquitination and deubiquitination. Ferroptosis may participate in the recruitment of immune system and portrait of tumor immune microenvironment.
The landscape of STK11 mutations and hotspots in Chinese patients with solid tumors.
Authors: Han Yun , Su Junyan 
Published: June 2022

e15112 Background: STK11 is a tumor suppressor gene, and loss-of-function mutations are oncogenic, due to loss of AMPK regulation of mTOR and HIF-1-α. STK11 has been identified as a tumor suppressor in several types of cancers. Emerging evidence suggests that STK11 mutations may influence clinical outcome and response to immunotherapy in cancer. However, the genomic features of STK11 are obscured in Chinese solid tumors. In this study, we investigated the landscape of STK11 in Chinese patients with solid tumors. Methods: Next-generation targeted sequencing of major cancer-related 599 genes in a large cohort of 5420 Chinese solid tumors. Results: STK11 mutations were identified in 184 (3.39%, 184/5420) Chinese solid tumors patients. Although STK11 mutations are negligible in solid tumors, endometrial cancer patients (15.79%), esophagus and esophagogastric junction cancer (9.09%), ovarian cancer (4.92%), and lung cancer (4.92%) possess relatively high rates of STK11 mutation. However, STK11 alterations were not found in head and neck cancer, kidney cancer, and CNS tumor cases. A total of 194 mutation sites have been identified in STK11, including 43 pathogenic/ likely pathogenic mutations, the most prominent mutational hotspots are P281fs, D194Y, W308C, S216F, D53fs, and Y60, occupying almost 48.84% of all mutations in STK11. A total of 17 patients harbored copy number variations (CNV) of STK11 in Chinese solid tumors, mainly occurring in the neuroendocrine tumor, breast cancer, and gastric cancer. Conclusions: We found a unique genomic feature of STK11 in Chinese solid tumor patients. Our results show that STK11 may be a potential therapeutic target.