Epigenetic reprogramming of Runx3 reinforces CD8 + T-cell function and improves the clinical response to immunotherapy.

BACKGROUND: Checkpoint blockade immunotherapy, represented by PD-1 or PD-L1 antibody treatment, has been of tremendous success in clinical practice. However, the low clinical response rate and lack of biomarkers for prediction of the immune response limit the clinical application of anti-PD-1 immunotherapy. Our recent work showed that a combination of low-dose decitabine and PD-1-ab significantly improved the complete response (CR) rate of cHL patients from 32 to 71%, which indicates that there is a significant correlation between epigenetic regulation and the clinical response to immunotherapy. METHODS: We recruited two groups of Hodgkin lymphoma patients who were treated with anti-PD-1 and DAC+anti-PD-1. CD8+ T cells were isolated from the patients' peripheral blood, DNA methylation was analyzed by EPIC, the expression profile was analyzed by RNA-seq, and multigroup analysis was performed with IPA and GSEA functional annotations. We explored the effect of DAC on the function of CD8+ T cells in the blood, spleen, tumor and lymph nodes using a mouse model. Furthermore, we explored the function of Tils in the tumor microenvironment. Then, we constructed Runx3-knockout mice to confirm the T-cell-specific function of Runx3 in CD8+ T cells and analyzed various subtypes of T cells and cytokines using mass cytometry (CyTOF). RESULTS: Multiomics analysis identified that DNA methylation reprogramming of Runx3 was a crucial mediator of CD8+ T-cell function. Multiomics data showed that reversal of methylation of the Runx3 promoter promoted the infiltration of CD8+ TILs and mitigated the exhaustion of CD8+ T cells. Furthermore, experiments on tissue-specific Runx3-knockout mice showed that Runx3 deficiency reduced CD8+ T infiltration and the differentiation of effector T and memory T cells. Furthermore, Runx3 deficiency significantly decreased CCR3 and CCR5 levels. Immunotherapy experiments in Runx3 conditional knockout mice showed that DAC could not reverse the resistance of anti-PD-1 in the absence of Runx3. Moreover, both our clinical data and data from TISIDB showed that Runx3 could be a potential biomarker for immunotherapy to predict the clinical response rate. CONCLUSION: We demonstrate that the DNA methylation of Runx3 plays a critical role in CD8+ T-cell infiltration and differentiation during decitabine-primed PD-1-ab immunotherapy, which provides a supporting mechanism for the essential role of epiregulation in immunotherapy.

Camrelizumab combined with apatinib and S-1 as second-line treatment for patients with advanced gastric or gastroesophageal junction adenocarcinoma: a phase 2, single-arm, prospective study.

BACKGROUND: The current second-line treatment of advanced gastric or gastroesophageal junction adenocarcinoma remains unsatisfactory. Anti-PD-1 monoclonal antibody combined with anti-angiogenic therapy shows anti-tumor activity and synergistic effect. We aimed to assess the efficacy and safety of the combination therapy of camrelizumab, apatinib, and S-1 in patients with gastric or gastroesophageal junction adenocarcinoma. METHODS: In this open-label, single-arm, phase 2 trial, in each 21-day cycle, eligible patients received 200 mg intravenous camrelizumab in the first day, 500 mg oral apatinib once daily continuously, and specific dose oral S-1 in the first 14 days until the trial was discontinued disease progression, development of intolerable toxicity, or withdrawal of consent. The primary endpoint was objective response rate. The secondary endpoints were disease control rate, progression-free survival and overall survival, and safety. This study was registered at ClinicalTrials.gov, NCT04345783. RESULTS: Between May 2019 and August 2020, we enrolled a total of 24 patients in this trial. At the data cutoff (December 1, 2020), the median follow-up duration was 8.13 months. Seven of 24 (29.2%, 95%CI 14.9-49.2%) patients reached objective response. The median-progression-free survival was 6.5 months (95%CI 6.01-6.99) and the median overall survival was not reached. Grade 3 or 4 adverse events occurred in 6 (25.0%) patients, including elevated transaminase, thrombocytopenia, fatigue, proteinuria, and intestinal obstruction. No serious treatment-related adverse events or treatment-related deaths occurred. CONCLUSIONS: In this trial, the combination of camrelizumab, apatinib, and S-1 showed promising anti-tumor activity and manageable toxicity as a second-line therapy in patients with advanced gastric or gastroesophageal junction adenocarcinoma, regardless of PD-L1 expression. CLINICAL TRIAL REGISTRATION: NCT04345783.

Analysis of Real-World Data to Investigate the Impact of Race and Ethnicity on Response to Programmed Cell Death-1 and Programmed Cell Death-Ligand 1 Inhibitors in Advanced Non-Small Cell Lung Cancers.

BACKGROUND: Racial disparities among clinical trial participants present a challenge to assess whether trial results can be generalized into patients representing diverse races and ethnicities. The objective of this study was to evaluate the impact of race and ethnicity on treatment response in patients with advanced non-small cell lung cancer (aNSCLC) treated with programmed cell death-1 (PD-1) or programmed cell death-ligand 1 (PD-L1) inhibitors through analysis of real-world data (RWD). MATERIALS AND METHODS: A retrospective cohort study of 11,138 patients with lung cancer treated at hospitals within the Mount Sinai Health System was performed. Patients with confirmed aNSCLC who received anti-PD-1/PD-L1 treatment were analyzed for clinical outcomes. Our cohort included 249 patients with aNSCLC who began nivolumab, pembrolizumab, or atezolizumab treatment between November 2014 and December 2018. Time-to-treatment discontinuation (TTD) and overall survival (OS) were the analyzed clinical endpoints. RESULTS: After a median follow-up of 14.8 months, median TTD was 7.8 months (95% confidence interval, 5.4-not estimable [NE]) in 75 African American patients versus 4.6 (2.4-7.2) in 110 White patients (hazard ratio [HR], 0.63). Median OS was not reached (18.4-NE) in African American patients versus 11.6 months (9.7-NE) in White patients (HR, 0.58). Multivariable Cox regression conducted with potential confounders confirmed longer TTD (adjusted HR, 0.65) and OS (adjusted HR, 0.60) in African American versus White patients. Similar real-world response rate (42.6% vs. 43.5%) and disease control rate (59.6% vs. 56.5%) were observed in the African American and White patient populations. Further investigation revealed the African American patient group had lower incidence (14.7%) of putative hyperprogressive diseases (HPD) upon anti-PD-1/PD-L1 treatment than the White patient group (24.5%). CONCLUSION: Analysis of RWD showed longer TTD and OS in African American patients with aNSCLC treated with anti-PD-1/PD-L1 inhibitors. Lower incidence of putative HPD is a possible reason for the favorable outcomes in this patient population. IMPLICATIONS FOR PRACTICE: There is a significant underrepresentation of minority patients in randomized clinical trials, and this study demonstrates that real-world data can be used to investigate the impact of race and ethnicity on treatment response. In retrospective analysis of patients with advanced non-small cell lung cancer treated with programmed cell death-1 or programmed cell death-ligand 1 inhibitors, African American patients had significantly longer time-to-treatment discontinuation and longer overall survival. Analysis of real-world data can yield clinical insights and establish a more complete picture of medical interventions in routine clinical practice.

KDM5 histone demethylases repress immune response via suppression of STING.

Cyclic GMP-AMP (cGAMP) synthase (cGAS) stimulator of interferon genes (STING) senses pathogen-derived or abnormal self-DNA in the cytosol and triggers an innate immune defense against microbial infection and cancer. STING agonists induce both innate and adaptive immune responses and are a new class of cancer immunotherapy agents tested in multiple clinical trials. However, STING is commonly silenced in cancer cells via unclear mechanisms, limiting the application of these agonists. Here, we report that the expression of STING is epigenetically suppressed by the histone H3K4 lysine demethylases KDM5B and KDM5C and is activated by the opposing H3K4 methyltransferases. The induction of STING expression by KDM5 blockade triggered a robust interferon response in a cytosolic DNA-dependent manner in breast cancer cells. This response resulted in resistance to infection by DNA and RNA viruses. In human tumors, KDM5B expression is inversely associated with STING expression in multiple cancer types, with the level of intratumoral CD8+ T cells, and with patient survival in cancers with a high level of cytosolic DNA, such as human papilloma virus (HPV)-positive head and neck cancer. These results demonstrate a novel epigenetic regulatory pathway of immune response and suggest that KDM5 demethylases are potential targets for antipathogen treatment and anticancer immunotherapy.

The KDM5 family is required for activation of pro-proliferative cell cycle genes during adipocyte differentiation.

The KDM5 family of histone demethylases removes the H3K4 tri-methylation (H3K4me3) mark frequently found at promoter regions of actively transcribed genes and is therefore generally considered to contribute to corepression. In this study, we show that knockdown (KD) of all expressed members of the KDM5 family in white and brown preadipocytes leads to deregulated gene expression and blocks differentiation to mature adipocytes. KDM5 KD leads to a considerable increase in H3K4me3 at promoter regions; however, these changes in H3K4me3 have a limited effect on gene expression per se. By contrast, genome-wide analyses demonstrate that KDM5A is strongly enriched at KDM5-activated promoters, which generally have high levels of H3K4me3 and are associated with highly expressed genes. We show that KDM5-activated genes include a large set of cell cycle regulators and that the KDM5s are necessary for mitotic clonal expansion in 3T3-L1 cells, indicating that KDM5 KD may interfere with differentiation in part by impairing proliferation. Notably, the demethylase activity of KDM5A is required for activation of at least a subset of pro-proliferative cell cycle genes. In conclusion, the KDM5 family acts as dual modulators of gene expression in preadipocytes and is required for early stage differentiation and activation of pro-proliferative cell cycle genes.

AID-associated DNA repair pathways regulate malignant transformation in a murine model of BCL6-driven diffuse large B-cell lymphoma.

Somatic hypermutation and class-switch recombination of the immunoglobulin (Ig) genes occur in germinal center (GC) B cells and are initiated through deamination of cytidine to uracil by activation-induced cytidine deaminase (AID). Resulting uracil-guanine mismatches are processed by uracil DNA glycosylase (UNG)-mediated base-excision repair and MSH2-mediated mismatch repair (MMR) to yield mutations and DNA strand lesions. Although off-target AID activity also contributes to oncogenic point mutations and chromosome translocations associated with GC and post-GC B-cell lymphomas, the role of downstream AID-associated DNA repair pathways in the pathogenesis of lymphoma is unknown. Here, we show that simultaneous deficiency of UNG and MSH2 or MSH2 alone causes genomic instability and a shorter latency to the development of BCL6-driven diffuse large B-cell lymphoma (DLBCL) in a murine model. The additional development of several BCL6-independent malignancies in these mice underscores the critical role of MMR in maintaining general genomic stability. In contrast, absence of UNG alone is highly protective and prevents the development of BCL6-driven DLBCL. We further demonstrate that clonal and nonclonal mutations arise within non-Ig AID target genes in the combined absence of UNG and MSH2 and that DNA strand lesions arise in an UNG-dependent manner but are offset by MSH2. These findings lend insight into a complex interplay whereby potentially deleterious UNG activity and general genomic instability are opposed by the protective influence of MSH2, producing a net protective effect that promotes immune diversification while simultaneously attenuating malignant transformation of GC B cells.

NTRK fusion oncogenes in pediatric papillary thyroid carcinoma in northeast United States.

BACKGROUND: An increase in thyroid cancers, predominantly papillary thyroid carcinoma (PTC), has been recently reported in children. METHODS: The histopathology of 28 consecutive PTCs from the northeast United States was reviewed. None of the patients (ages 6-18 years; 20 females, 8 males) had significant exposure to radiation. Nucleic acid from tumors was tested for genetic abnormalities (n = 27). Negative results were reevaluated by targeted next-generation sequencing. RESULTS: Seven of 27 PTCs (26%) had neurotrophic tyrosine kinase receptor (NTRK) fusion oncogenes (NTRK type 3/ets variant 6 [NTRK3/ETV6], n =5; NTRK3/unknown, n = 1; and NTRK type 1/translocated promoter region, nuclear basket protein [NTRK1/TPR], n = 1), including 5 tumors that measured >2 cm and 3 that diffusely involved the entire thyroid or lobe. All 7 tumors had lymphatic invasion, and 5 had vascular invasion. Six of 27 PTCs (22%) had ret proto-oncogene (RET) fusions (RET/PTC1, n = 5; RET/PTC3, n = 1); 2 tumors measured >2 cm and diffusely involved the thyroid, and 5 had lymphatic invasion, with vascular invasion in 2. Thirteen PTCs had the B-Raf proto-oncogene, serine/threonine kinase (BRAF) valine-to-glutamic acid mutation at position 600 (BRAF(V) (600E)) (13 of 27 tumors; 48%), 11 measured <2 cm, and 6 had lymphatic invasion (46%), with vascular invasion in 3. Fusion oncogene tumors, compared with BRAF(V) (600E) PTCs, were associated with large size (mean, 2.2 cm vs 1.5 cm, respectively; P = .05), solid and diffuse variants (11 of 13 vs 0 of 13 tumors, respectively; P < .001), and lymphovascular invasion (12 of 13 vs 6 of 13 tumors, respectively; P = .02); BRAF(V) (600E) PTCs were predominantly the classic variant (12 of 13 vs 1 of 13 tumors). Two tumors metastasized to the lung, and both had fusion oncogenes (NTRK1/TPR, n = 1; RET/PTC1, n = 1). CONCLUSIONS: Fusion oncogene PTC presents with more extensive disease and aggressive pathology than BRAF(V) (600E) PTC in the pediatric population. The high prevalence of the NTRK1/NTRK3 fusion oncogene PTCs in the United States is unusual and needs further investigation.

Histone demethylase jumonji AT-rich interactive domain 1B (JARID1B) controls mammary gland development by regulating key developmental and lineage specification genes.

The JmjC domain-containing H3K4 histone demethylase jumonji AT-rich interactive domain 1B (JARID1B) (also known as KDM5B and PLU1) is overexpressed in breast cancer and is a potential target for breast cancer treatment. To investigate the in vivo function of JARID1B, we developed Jarid1b(-/-) mice and characterized their phenotypes in detail. Unlike previously reported Jarid1b(-/-) strains, the majority of these Jarid1b(-/-) mice were viable beyond embryonic and neonatal stages. This allowed us to further examine phenotypes associated with the loss of JARID1B in pubertal development and pregnancy. These Jarid1b(-/-) mice exhibited decreased body weight, premature mortality, decreased female fertility, and delayed mammary gland development. Related to these phenotypes, JARID1B loss decreased serum estrogen level and reduced mammary epithelial cell proliferation in early puberty. In mammary epithelial cells, JARID1B loss diminished the expression of key regulators for mammary morphogenesis and luminal lineage specification, including FOXA1 and estrogen receptor α. Mechanistically, JARID1B was required for GATA3 recruitment to the Foxa1 promoter to activate Foxa1 expression. These results indicate that JARID1B positively regulates mammary ductal development through both extrinsic and cell-autonomous mechanisms.

[Correlation between type 2 diabetes and DNA methylation and mRNA expression of APN in abdominal adipose tissues in Xinjiang Uygur population].

To investigate the relationship between type 2 diabetes mellitus (T2DM) onset and development and mRNA expression and promoter methylation of adiponectin (APN) gene in abdominal adipose tissues of Xinjiang Uygur population, abdominal adipose tissues of omentum were collected and divided into control, obesity and T2DM groups. The status of APN promoter methylation was detected by denaturing high performance liquid chromatography (DHPLC), while the mRNA expression level of APN was detected by RT-PCR. Results show that methylation positive rate of APN was at the lowest level in control, middel in obesity and highest in T2DM groups, and the differences are statistically significant. Comparing the APN mRNA relative copy number of adipose tissue in each group, we found that the relative copy number of APN in control group is significantly higher than that of obesity and T2DM groups. There is a negative correlation between the mRNA expression level of APN in abdominal adipose tissue and fasting plasma glucose (FPG), glycosylated hemoglobin (HbA1c) and triglyceride (TG) level. There is a negative correlation in DNA promoter methylation and mRNA expression of APN gene. Relative copy number of APN in DNA methylation positive group is significantly lower than that of the negative group. In conclusion, increased APN promoter methylation results in decreased mRNA expression, which induces glucose and lipid metabolic disorder, thus contributing to the initiation and development of T2DM in Xinjiang Uygur population.

Association of the HTR2A gene with alcohol and heroin abuse.

Positive genetic associations of rs6313 (102T/C at exon 1) and rs6311 (-1438A/G) on the 5-hydroxytryptamine (serotonin) 2A receptor gene (HTR2A or 5-HT2A) were reported for alcohol and drug abuse; however, other association studies failed to produce consistent results supporting the susceptibility of the two single nucleotide polymorphisms (SNPs). To clarify the associations of the HTR2A gene with substance use disorders, we performed a meta-analysis based on the genotypes from the available candidate gene association studies of the two SNPs with alcohol and drug abuse from multiple populations. Evidence of association was found for HTR2A rs6313 in all the combined studies (e.g., allelic P = 0.0048 and OR 0.86, 95 % CI 0.77-0.95) and also in the combined studies of alcohol dependence (abuse) (e.g., allelic P = 0.0001 and OR 0.71, 95 % CI 0.59-0.85). The same association trend was also observed in the Study of Addiction: Genetics and Environment datasets. The meta-analysis supports a contribution of the HTR2A gene to the susceptibility to substance use disorders, particularly alcohol dependence.

RNASTAR: an RNA STructural Alignment Repository that provides insight into the evolution of natural and artificial RNAs.

Automated RNA alignment algorithms often fail to recapture the essential conserved sites that are critical for function. To assist in the refinement of these algorithms, we manually curated a set of 148 alignments with a total of 9600 unique sequences, in which each alignment was backed by at least one crystal or NMR structure. These alignments included both naturally and artificially selected molecules. We used principles of isostericity to improve the alignments from an average of 83%-94% isosteric base pairs. We expect that this alignment collection will assist in a wide range of benchmarking efforts and provide new insight into evolutionary principles governing change in RNA structural motifs. The improved alignments have been contributed to the Rfam database.

Let-7b-5p regulates proliferation and apoptosis in multiple myeloma by targeting IGF1R.

Multiple myeloma (MM) is the most common cause of death from hematological malignancy worldwide, and recent studies have revealed that let-7b-5p can play an inhibitory role in tumorigenesis. However, the role of let-7b-5p in MM still remains unclear. The aim of this study was to elucidate the molecular mechanisms by which let-7b-5p acts as a tumor suppressor in MM. Here, quantitative real-time polymerase chain reaction results showed that the expression of let-7b-5p was remarkably reduced in MM tissues and MM cell lines (RPMI-8226 and U266 cells). Furthermore, over-expression of let-7b-5p significantly suppressed RPMI-8226 cell proliferation and induced S/G2 cell cycle arrest and apoptosis. Luciferase reporter assay results demonstrated that insulin-like growth factor receptor 1 (IGF1R) was negatively regulated by let-7b-5p at the post-transcriptional level. The mRNA and protein levels of IGF1R in RPMI-8226 cells were down-regulated by let-7b-5p. Furthermore, the cell phenotype altered by let-7b-5p inhibitor can be rescued by IGF1R silencing (si-IGF1R). Taken together, our results demonstrated that let-7b-5p functions as a tumor suppressor in MM, suggesting that let-7b-5p may be a potential therapeutic target for MM.

Loss of the retinoblastoma binding protein 2 (RBP2) histone demethylase suppresses tumorigenesis in mice lacking Rb1 or Men1.

Aberrations in epigenetic processes, such as histone methylation, can cause cancer. Retinoblastoma binding protein 2 (RBP2; also called JARID1A or KDM5A) can demethylate tri- and dimethylated lysine 4 in histone H3, which are epigenetic marks for transcriptionally active chromatin, whereas the multiple endocrine neoplasia type 1 (MEN1) tumor suppressor promotes H3K4 methylation. Previous studies suggested that inhibition of RBP2 contributed to tumor suppression by the retinoblastoma protein (pRB). Here, we show that genetic ablation of Rbp2 decreases tumor formation and prolongs survival in Rb1(+/-) mice and Men1-defective mice. These studies link RBP2 histone demethylase activity to tumorigenesis and nominate RBP2 as a potential target for cancer therapy.

ASCL1 Drives Tolerance to Osimertinib in EGFR Mutant Lung Cancer in Permissive Cellular Contexts.

The majority of EGFR mutant lung adenocarcinomas respond well to EGFR tyrosine kinase inhibitors (TKI). However, most of these responses are partial, with drug-tolerant residual disease remaining even at the time of maximal response. This residual disease can ultimately lead to relapses, which eventually develop in most patients. To investigate the cellular and molecular properties of residual tumor cells in vivo, we leveraged patient-derived xenograft (PDX) models of EGFR mutant lung cancer. Subcutaneous EGFR mutant PDXs were treated with the third-generation TKI osimertinib until maximal tumor regression. Residual tissue inevitably harbored tumor cells that were transcriptionally distinct from bulk pretreatment tumor. Single-cell transcriptional profiling provided evidence of cells matching the profiles of drug-tolerant cells present in the pretreatment tumor. In one of the PDXs analyzed, osimertinib treatment caused dramatic transcriptomic changes that featured upregulation of the neuroendocrine lineage transcription factor ASCL1. Mechanistically, ASCL1 conferred drug tolerance by initiating an epithelial-to-mesenchymal gene-expression program in permissive cellular contexts. This study reveals fundamental insights into the biology of drug tolerance, the plasticity of cells through TKI treatment, and why specific phenotypes are observed only in certain tumors. SIGNIFICANCE: Analysis of residual disease following tyrosine kinase inhibitor treatment identified heterogeneous and context-specific mechanisms of drug tolerance in lung cancer that could lead to the development of strategies to forestall drug resistance. See related commentary by Rumde and Burns, p. 1188.

Microbial community resemblance methods differ in their ability to detect biologically relevant patterns.

High-throughput sequencing methods enable characterization of microbial communities in a wide range of environments on an unprecedented scale. However, insight into microbial community composition is limited by our ability to detect patterns in this flood of sequences. Here we compare the performance of 51 analysis techniques using real and simulated bacterial 16S rRNA pyrosequencing datasets containing either clustered samples or samples arrayed across environmental gradients. We found that many diversity patterns were evident with severely undersampled communities and that methods varied widely in their ability to detect gradients and clusters. Chi-squared distances and Pearson correlation distances performed especially well for detecting gradients, whereas Gower and Canberra distances performed especially well for detecting clusters. These results also provide a basis for understanding tradeoffs between number of samples and depth of coverage, tradeoffs that are important to consider when designing studies to characterize microbial communities.

GPHMM: an integrated hidden Markov model for identification of copy number alteration and loss of heterozygosity in complex tumor samples using whole genome SNP arrays.

There is an increasing interest in using single nucleotide polymorphism (SNP) genotyping arrays for profiling chromosomal rearrangements in tumors, as they allow simultaneous detection of copy number and loss of heterozygosity with high resolution. Critical issues such as signal baseline shift due to aneuploidy, normal cell contamination, and the presence of GC content bias have been reported to dramatically alter SNP array signals and complicate accurate identification of aberrations in cancer genomes. To address these issues, we propose a novel Global Parameter Hidden Markov Model (GPHMM) to unravel tangled genotyping data generated from tumor samples. In contrast to other HMM methods, a distinct feature of GPHMM is that the issues mentioned above are quantitatively modeled by global parameters and integrated within the statistical framework. We developed an efficient EM algorithm for parameter estimation. We evaluated performance on three data sets and show that GPHMM can correctly identify chromosomal aberrations in tumor samples containing as few as 10% cancer cells. Furthermore, we demonstrated that the estimation of global parameters in GPHMM provides information about the biological characteristics of tumor samples and the quality of genotyping signal from SNP array experiments, which is helpful for data quality control and outlier detection in cohort studies.

A negative regulatory loop between microRNA and Hox gene controls posterior identities in Caenorhabditis elegans.

MicroRNAs (miRNAs) have been found to regulate gene expression across eukaryotic species, but the function of most miRNA genes remains unknown. Here we describe how the analysis of the expression patterns of a well-conserved miRNA gene, mir-57, at cellular resolution for every minute during early development of Caenorhabditis elegans provided key insights in understanding its function. Remarkably, mir-57 expression shows strong positional bias but little tissue specificity, a pattern reminiscent of Hox gene function. Despite the minor defects produced by a loss of function mutation, overexpression of mir-57 causes dramatic posterior defects, which also mimic the phenotypes of mutant alleles of a posterior Hox gene, nob-1, an Abd homolog. More importantly, nob-1 expression is found in the same two posterior AB sublineages as those expressing mir-57 but with an earlier onset. Intriguingly, nob-1 functions as an activator for mir-57 expression; it is also a direct target of mir-57. In agreement with this, loss of mir-57 function partially rescues the nob-1 allele defects, indicating a negative feedback regulatory loop between the miRNA and Hox gene to provide positional cues. Given the conservation of the miRNA and Hox gene, the regulatory mechanism might be broadly used across species. The strategy used here to explore mir-57 function provides a path to dissect the regulatory relationship between genes.

CRISPR-Cas13d effectively targets SARS-CoV-2 variants, including Delta and Omicron, and inhibits viral infection.

The recent pandemic of variants of concern (VOC) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) highlights the need for innovative anti-SARS-CoV-2 approaches in addition to vaccines and antiviral therapeutics. Here, we demonstrate that a CRISPR-Cas13-based strategy against SARS-CoV-2 can effectively degrade viral RNA. First, we conducted a cytological infection experiment, screened CRISPR-associated RNAs (crRNAs) targeting conserved regions of viruses, and used an in vitro system to validate functional crRNAs. Reprogrammed Cas13d effectors targeting NSP13, NSP14, and nucleocapsid transcripts achieved >99% silencing efficiency in human cells which are infected with coronavirus 2, including the emerging variants in the last 2 years, B.1, B.1.1.7 (Alpha), D614G B.1.351 (Beta), and B.1.617 (Delta). Furthermore, we conducted bioinformatics data analysis. We collected the sequence information of COVID-19 and its variants from China, and phylogenetic analysis revealed that these crRNA oligos could target almost 100% of the SARS-CoV family, including the emerging new variant, Omicron. The reprogrammed Cas13d exhibited high specificity, efficiency, and rapid deployment properties; therefore, it is promising for antiviral drug development. This system could possibly be used to protect against unexpected SARS-CoV-2 variants carrying multiple mutations.

Analysis of real-world data to investigate evolving treatment sequencing patterns in advanced non-small cell lung cancers and their impact on survival.

Background: Although optimal sequencing of systemic therapy in cancer care is critical to achieving maximal clinical benefit, there is a lack of analysis of treatment sequencing in advanced non-small cell lung cancer (aNSCLC) in real-world settings. Methods: A retrospective cohort study of 13,340 lung cancer patients within the Mount Sinai Health System (MSHS) was performed. Systemic therapy data of aNSCLC in 2,106 patients was the starting point in our analysis to investigate how treatment sequencing has evolved, the impact of sequencing patterns on clinical outcomes, and the effectiveness of 2nd line chemotherapy after patients progressed on immune checkpoint inhibitor (ICI)-based therapy as the 1st line of therapy (LOT). Results: There is a significant shift to more ICI-based therapy and multiple lines of targeted therapy after 2015. We compared clinical outcomes of two patient populations with different treatment sequencing patterns, with the 1st group receiving chemotherapy as the 1st LOT followed by ICI-based treatment, and the 2nd group treated in the opposite order receiving a 1st line ICI-containing regimen followed by a 2nd line chemotherapy. No statistically significant difference in overall survival (OS) was observed between the two groups [group 2 vs. group 1, adjusted hazard ratio (aHR) =1.36, P=0.39]. We assessed the efficacy of the 2nd line chemotherapy in three patient populations given either 1st line ICI single agent, 1st line ICI-chemotherapy combination, or 1st line chemotherapy alone, there was no statistically significant difference in time-to-next treatment (TTNT) and in OS among the three patient groups. Conclusions: Analysis of real-world data has shown two treatment sequencing patterns in aNSCLC, ICI followed by chemotherapy or chemotherapy followed by ICI, achieved similar clinical benefit. The chemotherapies routinely used following platinum doublet 1st LOT, is effective as the 2nd line option after ICI-chemotherapy combination in the 1st line setting.

Detecting Ground Glass Opacity Features in Patients With Lung Cancer: Automated Extraction and Longitudinal Analysis via Deep Learning-Based Natural Language Processing.

BACKGROUND: Ground-glass opacities (GGOs) appearing in computed tomography (CT) scans may indicate potential lung malignancy. Proper management of GGOs based on their features can prevent the development of lung cancer. Electronic health records are rich sources of information on GGO nodules and their granular features, but most of the valuable information is embedded in unstructured clinical notes. OBJECTIVE: We aimed to develop, test, and validate a deep learning-based natural language processing (NLP) tool that automatically extracts GGO features to inform the longitudinal trajectory of GGO status from large-scale radiology notes. METHODS: We developed a bidirectional long short-term memory with a conditional random field-based deep-learning NLP pipeline to extract GGO and granular features of GGO retrospectively from radiology notes of 13,216 lung cancer patients. We evaluated the pipeline with quality assessments and analyzed cohort characterization of the distribution of nodule features longitudinally to assess changes in size and solidity over time. RESULTS: Our NLP pipeline built on the GGO ontology we developed achieved between 95% and 100% precision, 89% and 100% recall, and 92% and 100% F1-scores on different GGO features. We deployed this GGO NLP model to extract and structure comprehensive characteristics of GGOs from 29,496 radiology notes of 4521 lung cancer patients. Longitudinal analysis revealed that size increased in 16.8% (240/1424) of patients, decreased in 14.6% (208/1424), and remained unchanged in 68.5% (976/1424) in their last note compared to the first note. Among 1127 patients who had longitudinal radiology notes of GGO status, 815 (72.3%) were reported to have stable status, and 259 (23%) had increased/progressed status in the subsequent notes. CONCLUSIONS: Our deep learning-based NLP pipeline can automatically extract granular GGO features at scale from electronic health records when this information is documented in radiology notes and help inform the natural history of GGO. This will open the way for a new paradigm in lung cancer prevention and early detection.