High Mutation Frequency Research Articles

Pan-cancer analysis from multi-omics data reveals AAMP as an unfavourable prognostic marker

ObjectivesAngio-associated migratory cell protein (AAMP) is a protein that participates in cell migration and is reported to be involved in cancer progression. However, the molecular mechanism of AAMP in pan-cancer is not known.MethodsWe used multi-omics data, such as TIMER, TCGA, GTEx, CPTAC, HPA, and cBioPortal to analyze AAMP expression, and gene alteration in pan-cancer. Univariate Cox regression and Kaplan–Meier were utilized to explore prognostic significance of AAMP expression level. We applied Spearman analysis to investigate the correlation between AAMP and TMB, MSI, immune cell infiltration, immune checkpoints. Moreover, we mainly studied liver hepatocellular carcinoma(LIHC) to explore AAMP expression, clinical significance, and prognosis. Cox regression analysis was used to study independent factor to predict prognosis for AAMP in LIHC. GSEA was utilized to investigate the biological function for AAMP in LIHC.ResultsAAMP was overexpressed in most cancers, and high AAMP expression was associated with worse overall survival (OS), disease-specific survival (DSS), and progress-free interval (PFI) for LIHC and adrenocortical carcinoma (ACC). Moreover, AAMP had the highest mutation frequency in uterine corpus endometrial carcinoma (UCEC). AAMP was correlated with TMB and MSI in esophageal carcinoma (ESCA), stomach adenocarcinoma (STAD), lung squamous cell carcinoma (LUSC), and thyroid carcinoma (THCA). Then, we focus on LIHC to investigate the expression and prognosis of AAMP. AAMP overexpression was related to histological grade and pathological stage in LIHC. Multivariate Cox regression analysis revealed that AAMP overexpression was an independent adverse prognostic marker for LIHC. AAMP expression was correlated with immune cell infiltration and immune checkpoints in LIHC. Function enrichment analysis indicated the participation of AAMP in the cell cycle and DNA replication.ConclusionsAAMP was a latent prognostic indicator for pan-cancer and had high potential as a biomarker for the diagnosis and prognosis of LIHC.

High frequency of Lamivudine and Telbivudine resistance mutations in hepatitis B virus isolates from human immunodeficiency virus co-infected patients on highly active antiretroviral therapy in Bucaramanga, Colombia.

Hepatitis B virus (HBV) antiviral Resistance-Associated Mutations (RAMs) in human immunodeficiency virus (HIV) coinfected patients undergoing highly active antiretroviral therapy (HAART) are complex and incompletely understood. We aimed to determine the prevalence of HBV coinfection, HBV genotypes, and RAMs in a cohort of people living with HIV (PLWH) in the northeastern region of Colombia. This cross-sectional study was carried out between February 2013 and February 2014. Virological, immunological and HAART data were collected from clinical records. In-house nested PCR and Sanger sequencing of the HBV pol gene were used to identify coinfections, genotypes, RAMs and HBV s antigen (HBsAg) escape mutants. Among 275 PLWH, HBV coinfection was confirmed in 32 patients (11.6%), of whom nine (28.2%) were HBsAg positive (active hepatitis B), and 23 (71.8%) were occult hepatitis B infections (OBI). All HBV sequences (n = 23) belonged to the genotype F3. Among HIV/HBV coinfections, 71.9% had CD4+ T cell counts above 200 cells/mm3 and 37.5% had undetectable HIV viral loads. The RAMs rtL80I, rtL180M, and rtM204V, which confer resistance to Lamivudine/Telbivudine and partially resistant to Entecavir, were found in all HBV isolates. An unknown rt236Y mutation to Tenofovir was also identified. Most patients under HAART received first-generation HBV antiviral therapy with a low genetic barrier to resistance. Antiviral Drug-associated Potential Vaccine-escape Mutations (ADAPVEMs) in the S gene were observed in all isolates ranging from 1-20 amino acid substitutions. However, no vaccine escape mutants were detected. In Conclusion, these findings highlight the importance of HBV molecular screening, antiviral resistance monitoring and new guidelines for PLWH to overcome RAMs and prevent HBV-related liver disease.

IMMUNOREACT 6: weak immune surveillance characterizes early-onset rectal cancer.

Colon cancer in young patients is often associated with hereditary syndromes; however, in early-onset rectal cancer, mutations of these genes are rarely observed. The aim of this study was to analyse the features of the local immune microenvironment and the mutational pattern in early-onset rectal cancer. Commonly mutated genes were analysed within a rectal cancer series from the University Hospital of Padova. Mutation frequency and immune gene expression in a cohort from The Cancer Genome Atlas ('TCGA') were compared and immune-cell infiltration levels in the healthy rectal mucosa adjacent to rectal cancers were evaluated in the IMMUNOlogical microenvironment in REctal AdenoCarcinoma Treatment 1 and 2 ('IMMUNOREACT') series. In the authors' series, the mutation frequency of BRAF, KRAS, and NRAS, as well as microsatellite instability frequency, were not different between early- and late-onset rectal cancer. In The Cancer Genome Atlas series, among the genes with the most considerable difference in mutation frequency between young and older patients, seven genes are involved in the immune response and CD69, CD3, and CD8β expression was lower in early-onset rectal cancer. In the IMMUNOlogical microenvironment in REctal AdenoCarcinoma Treatment 1 and 2 series, young patients had a lower rate of CD4+ T cells, but higher T regulator infiltration in the rectal mucosa. Early-onset rectal cancer is rarely associated with common hereditary syndromes. The tumour microenvironment is characterized by a high frequency of mutations impairing the local immune surveillance mechanisms and low expression of immune editing-related genes. A constitutively low number of CD4 T cells associated with a high number of T regulators indicates an imbalance in the immune surveillance mechanisms.

Construction of PANoptosis signature: Novel target discovery for prostate cancer immunotherapy

PANoptosis pathway gene sets encompassing pyroptosis, apoptosis, and necroptosis were identified from the MSigDB database. We analyzed the perturbations and crosstalk in the PANoptosis pathway in prostate adenocarcinoma (PRAD), including gene mutation, transcription, methylation, and clinical features. By constructing a PANoptosis signature, we accurately predicted the prognosis and immunotherapeutic response of PRAD patients. We further explored the molecular features and immunological roles of the signature, dividing patients into high- and low-score groups. Notably, the high-score group correlated with better survival outcomes and immunotherapeutic responses, as well as a higher mutation frequency and enrichment score in the PANoptosis and HALLMARK pathways. The PANoptosis signature also enhanced overall antitumor immunity, promoted immune cell infiltration, upregulated immune checkpoint regulators, and revealed the cold tumor characteristics of PRAD. We also identified potential drug targets based on the PANoptosis signature. These findings lead the way in identifying novel prognostic markers and therapeutic targets for patients with PRAD.

Mutant p53 in head and neck squamous cell carcinoma: Molecular mechanism of gain‑of‑function and targeting therapy (Review).

Head and neck squamous cell carcinoma (HNSCC) is one of the most widespread malignancies worldwide. p53, as a transcription factor, can play its role in tumor suppression by activating the expression of numerous target genes. However, p53 is one of the most commonly mutated genes, which frequently harbors missense mutations. These missense mutations are nucleotide substitutions that result in the substitution of an amino acid in the DNA binding domain. Most p53 mutations in HNSCC are missense mutations and the mutation rate of p53 reaches 65‑85%. p53 mutation not only inhibits the tumor suppressive function of p53 but also provides novel functions to facilitate tumor recurrence, called gain‑of‑function (GOF). The present study focused on the prevalence and clinical relevance of p53 mutations in HNSCC, and further described how mutant p53 accumulates. Moreover, mutant p53 in HNSCC can interact with proteins, RNA, and exosomes to exert effects on proliferation, migration, invasion, immunosuppression, and metabolism. Finally, several treatment strategies have been proposed to abolish the tumor‑promoting function of mutant p53; these strategies include reactivation of mutant p53 into wild‑type p53, induction of mutant p53 degradation, enhancement of the synthetic lethality of mutant p53, and treatment with immunotherapy. Due to the high frequency of p53 mutations in HNSCC, a further understanding of the mechanism of mutant p53 may provide potential applications for targeted therapy in patients with HNSCC.

Inference of selection pressures that drive insecticide resistance in Anopheles and Culex mosquitoes in Korea

Pyrethroids are primarily used for mosquito control in Korea. However, high frequencies of mutations conferring resistance to not only pyrethroids but also to other insecticides have been found in mosquito populations. This study aimed to examine the hypothesis that insecticides used outside of public health may play a role in selection. Briefly, the resistance mutation frequencies to three insecticide groups (pyrethroids, organophosphates, and cyclodienes) were estimated in two representative groups of mosquito species (Anopheles Hyrcanus Group and Culex pipiens complex). The relationship between these frequencies and the land-use status of the collection sites was investigated through multiple regression analysis. In the Anopheles Hyrcanus Group, the frequencies of both ace1 (organophosphate resistance) and rdl (cyclodiene resistance) mutations were positively correlated with ‘proximity to golf course’, possibly be due to the insecticides used for turf maintenance. They also showed positive correlations with field area and rice paddy area, respectively, suggesting the role of agricultural insecticides in the selection of these resistance traits. For the Cx. pipiens complex, the kdr (pyrethroid resistance), ace1, and rdl mutations were positively correlated with the residential area, field, and rice paddy, respectively. Therefore, pyrethroids used for public health could serve as a direct source of resistance selection pressure against kdr, whereas non-public health insecticides may pose primary selection pressure against the ace1 and rdl traits. The current findings suggest that the insecticides used in agriculture and the golf industry play a significant role in mosquito selection, despite variations in the extent of indirect selection pressure according to the mosquito groups and insecticide classes.

Construction and validation of a cuproptosis-related five-lncRNA signature for predicting prognosis, immune response and drug sensitivity in breast cancer

BackgroundDespite advances in treatment, recurrence and mortality rates from breast cancer (BrCa) continue to rise, clinical effectiveness is limited, and prognosis remains disappointing, especially for patients with HER2-positive, triple-negative, or advanced breast cancer. Based on cuproptosis-related long noncoding RNAs (CRLs), this study aims to create a predictive signature to assess the prognosis in patients with BrCa.MethodsThe related CRLs RNA-seq data clinicopathological data were collected from The Cancer Genome Atlas (TCGA) database, and the predictive model was constructed after correlation analysis. Subsequently, we examined and validated connections and changes in the CRLs model with prognostic features (including risk curves, ROC curves and nomograms), pathway and functional enrichment, tumor mutation (TMB), tumor immune dysfunction and exclusion (TIDE) and treatment sensitivity.ResultsA prediction model formula composed of 5 CRLs was obtained, and divided breast cancer patients into high and low risk subgroups according to the obtained risk scores. The results showed that the overall survival (OS) of patients in the high-risk group was lower than that in the low-risk group, and the AUC of all samples at 1, 3 and 5 years were 0.704, 0.668 and 0.647, respectively. It was indicated that CRLs prognostic model could independently predict prognostic indicators of BrCa patients. In addition, analysis of gene set enrichment, immune function, TMB, and TIDE showed that these differentially expressed CRLs had a wealth of related pathways and functions, and might be closely related to immune response and immune microenvironment. Additionally, TP53 was found to have the highest mutation frequency in high-risk group (40%), while PIK3CA was found to have the highest mutation frequency in low-risk group (42%), which might become new targets for targeted therapy. Finally, we compared susceptibility to anticancer agents to identify potential treatment options for breast cancer. Lapatinib, Sunitinib, Phenformin, Idelalisib, Ruxolitinib, Cabozantinib were more sensitive to patients in the low-risk group, while Sorafenib, Vinorelbine, Pyrimethamine were more sensitive to patients in high-risk group, namely, these drugs could potentially be used in the future to treat breast cancer patients grouped according to the risk model.ConclusionThis study identified CRLs associated with breast cancer and provided a tailored tool for predicting prognosis, immune response, and drug sensitivity in patients with BrCa.

Feasibility analysis of rapid gene detection using intraoperative frozen tissues: comparison of intraoperative frozen tissues with paraffin-embedded tissues in epidermal growth factor receptor gene mutation detection of lung adenocarcinoma.

Epidermal growth factor receptor (EGFR) is the driver gene with the highest frequency of mutations in lung adenocarcinoma and can guide the development of targeted therapies. The detection of routine gene mutations must be performed after the preparation of paraffin samples in a standard polymerase chain reaction (PCR) laboratory, which is time-consuming. The Idylla™ EGFR fully automatic PCR system for rapid detection requires no special detection environment and completes the process in only 2.5h. It has been applied to tissues embedded in paraffin. The Idylla™ EGFR automated PCR system was used to detect EGFR gene mutations in intraoperative frozen fresh tissues and paraffin-embedded tissues from 47 enrolled patients with lung adenocarcinoma. The gold standard amplification refractory mutation system (ARMS) method for gene mutation detection was used for verification, and the concordance between the three detection results was compared, to investigate the feasibility of detecting rapid gene mutations in intraoperative frozen samples. The EGFR mutation rate in 47 fresh samples of lung adenocarcinoma was 61.7% (29/47), which is consistent with the mutation level of lung adenocarcinoma in the Asian population (38.8-64.0%). The concordance rate between the Idylla™ frozen tissues and paraffin-embedded tissues was 91.4% (43/47) when compared to the ARMS method, while the coincidence rate between the two methods was 93.6% (44/47). The three methods had a total consistency rate of 89.4% (42/47). The Idylla™ EGFR fully automatic PCR system directly detects EGFR mutations in fresh tissues. The operation is simple, the detection time is short, and the accuracy is high. The detection time is reduced to 1/4-1/3 of the original time while meeting clinical standards for detecting the gene status of patients, thus saving crucial time for individualized and accurate treatment of patients. The method has promising clinical application prospects.

Biological effect of gamma and electron beam to target turning X-ray irradiation on two varieties of freesia in M3 generation

The study was carried out to explore the effects of 60Co-γ irradiation and electron beam to target turning X-ray (EBTTX) irradiation in the M3 generation to accelerate the breeding process of freesia. Bulbs of two varieties of freesia “Freesia armstrongii W. Watson” and “Freesia corymbosa (Burm.f.) N. E. Br” were exposed to 4 doses of 60Co-γ and EBTTX irradiation (25, 50, 75, and 100 Gy). The stimulatory effect on germination rate, survival rate, plant height, leaf number, leaf area, root number, root length, flowering plant rate, and pedicel length of two freesia cultivars at 25 Gy of 60Co-γ irradiation dose was better than EBTTX in M3 generation. High irradiation dose exhibited stronger inhibitory effects on these plants' growth parameters and even the survival rate of two freesia cultivars was 0.00% at 100 Gy irradiation dose for both types of irradiation. The total chlorophyll content of freesia “Freesia armstrongii W. Watson” and “Freesia corymbosa (Burm.f.) N. E. Br” reached the maximum under the radiation dose of 25 Gy. With the increase in radiation dose, the content of MDA, SOD, and POD in the leaves of both varieties of freesia increased gradually. In the M3 generation of the two freesia varieties, five new mutant plants such as dwarf plants, altered flower color, and increased petal numbers were found. A high frequency of mean mutations was found in EBTTX irradiated plants (4.16% and 3.3%) than in gamma-irradiated plants (2.4% and 2.5%) at 50 Gy as compared with control in Freesia armstrongii W. Watson and Freesia corymbosa (Burm.f.) N. E. Br.

The epigenetic modifier lysine methyltransferase 2C is frequently mutated in gastric remnant carcinoma.

Gastric remnant carcinoma (GRC), which occurs in the stomach after partial gastrectomy, is a rare and aggressive form of gastric adenocarcinoma (GAC). Comprehensive profiling of genomic mutations in GRC could provide the basis for elucidating the origin and characteristics of this cancer. Herein, whole-exome sequencing (WES) was performed on 36 matched tumor-normal samples from patients with GRC and identified recurrent mutations in epigenetic modifiers, notably KMT2C, ARID1A, NSD1, and KMT2D, in 61.11% of cases. Mutational signature analysis revealed a low frequency of microsatellite instability (MSI) in GRC, which was further identified by MSIsensor, MSI-polymerase chain reaction, and immunohistochemistry analysis. Comparative analysis demonstrated that GRC had a distinct mutation spectrum compared to that of GAC in The Cancer Genome Atlas samples, with a significantly higher mutation rate of KMT2C. Targeted deep sequencing (Target-seq) of an additional 25 paired tumor-normal samples verified the high mutation frequency (48%) of KMT2C in GRC. KMT2C mutations correlated with poor overall survival in both WES and Target-seq cohorts and were independent prognosticators in GRC. In addition, KMT2C mutations were positively correlated with favorable outcomes in immune checkpoint inhibitor-treated pan-cancer patients and associated with higher intratumoral CD3+ , CD8+ tumor-infiltrating lymphocyte counts, and PD-L1expression in GRC samples (p = 0.018, 0.092, 0.047, 0.010, and 0.034, respectively). Our dataset provides a platform for information and knowledge mining of the genomic characteristics of GRC and helps to frame new therapeutic approaches for this disease.

Lethal and mutagenic effects of different LET radiations on Bacillus subtilis spores

New, useful microorganism resources have been generated by ionizing radiation breeding technology. However, the mutagenic effects of ionizing radiation on microorganisms have not been systematically clarified. For a deeper understanding and characterization of ionizing radiation-induced mutations in microorganisms, we investigated the lethal effects of seven different linear energy transfer (LET) radiations based on the survival fraction (SF) and whole-genome sequencing analysis of the mutagenic effects of a dose resulting in an SF of around 1% in Bacillus subtilis spores. Consequently, the lower LET radiations (gamma [surface LET: 0.2 keV/µm] and 4He2+ [24 keV/µm]) showed low lethality and high mutation frequency (MF), resulting in the major induction of single-base substitutions. Whereas higher LET radiations (12C5+ [156 keV/µm] and 12C6+ [179 keV/µm]) showed high lethality and low MF, resulting in the preferential induction of deletion mutations. In addition, 12C6+ (111) ion beams likely possess characteristics of both low- and high-LET radiations simultaneously. A decrease in the relative biological effectiveness and an evaluation of the inactivation cross section indicated that 20Ne8+ (468 keV/µm) and 40Ar13+ (2214 keV/µm) ion beams had overkill effects. In conclusion, in the mutation breeding of microorganisms, it should be possible to regulate the proportions, types, and frequencies of induced mutations by selecting an ionizing radiation of an appropriate LET in accordance with the intended purpose.

Are the original SARS-CoV-2 novel mutants from in vitro culture able to escape the immune response?

Monitoring variations in the virus genome to understand the SARS-CoV-2 evolution and spread of the virus is extremely important. Seven early SARS-CoV-2 isolates in China were cultured in vitro and were analyzed for their viral infectivity through viral growth assay, tissue culture infectious dose(TCID50 ) assay, spike protein quantification, and next generation sequencinganalysis, and the resultant mutations in spike protein were used to generate the corresponding pseudoviruses for analysis of immune escape from vaccination and postinfection immunity. The results revealed that in vitro cultured SARS-CoV-2 virus had much higher mutation frequency (up to ~20 times) than that in infected patients, suggesting that SARS-CoV-2 diversify under favorable conditions. Monitoring viral mutations is not only helpful for better understanding of virus evolution and virulence change, but also the key to prevent virus transmission and disease progression. Compared with the D614G reference strain, a pseudovirus strain of SARS-CoV-2 was constructed with a high mutation rate site on the spike protein. We found some novel spike mutations during in vitro culture, such as E868Q, conferred further immune escape ability.

CSF3R-mutant chronic myelomonocytic leukemia is a distinct clinically subset with abysmal prognosis: a case report and systematic review of the literature

Chronic myelomonocytic leukemia (CMML) is a myelodysplastic syndrome/myeloproliferative neoplasm (MDS/MPN) chacaterized by persistent peripheral blood monocytosis, hypercellular bone marrow and dysplasia at least in one myeloid lineage. CMML shares much of its molecular landscape with other myeloid neoplasms, while differs from others such as chronic neutrophilic leukemia (CNL), given the high frequency of CSF3R mutations in the latter. In this article, we report a case of CSF3R-mutated CMML and dissect this rare entity by reviewing the medical literature, with the intent to understand how this rare mutation shapes CMML's clinical and morphological phenotype. CSF3R-mutated CMML emerges as a rare entity meeting the ICC/WHO diagnostic criteria for CMML and simultaneously showing clinical-pathological and molecular traits of CNL and atypical chronic myeloid leukemia, rising an important and difficult diagnostic and therapeutical issue.

Dose-dense neoadjuvant chemotherapy of operable and locally advanced inoperable triple negative breast cancer: First results of single-centre prospective trial

Introduction. Neoadjuvant chemotherapy (NACT) is the standard of care for II–III stages of TN BC. Complete pathomorphological response (pCR) is associated with a signifiant increase in event-free and overall survival. In addition, in the absence of pCR, post-neoadjuvant adjuvant therapy is prescribed, while if pCR is achieved, additional treatment is not carried out. Despite a large number of studies on NACT of TN BC, different NACT regimens in various clinical trials make it diffiult to interpret their results.Objective. To investigate the effiacy of 4ddAC followed by 12 weekly cycles of paclitaxel and carboplatin in TN BC (according to the RCB system and the frequency of pCR); determine the predictive factors for the effectiveness of this chemotherapy regimen, and the effect of dose intensity on the pCR rate.Methods. This prospective study included 154 patients with TN breast cancer who received dose-dence neoadjuvant chemotherapy from January 2017 to March 2022.Results. PCR rate was 53.25 % (n = 82), RCB 0–I was 66.88 % (n = 103), disease progression was observed in 3.25 % (n = 4). The frequency of germline mutations in the BRCA1/2 genes was 21.43 % (n = 33). The most common mutation was BRCA1 5382insC – 63.64 % (n = 21) of all BRCA1/2 mutations. Rare mutations detected by NGS accounted for 30.3 % (n = 10). The only factor associated with a signifiant increase in the pCR rate was age ≤50 years (p = 0.010), there was a trend towards an increase pCR rate in the subgroups T1–2 (p = 0.052) and BRCA1/2 mut (p = 0.080). There was no effect of the dose intensity on the achievement of pCR.Conclusions. This retrospective analysis is the largest study evaluating the effiacy of 4 ddAC followed by 12 PC in NACT TN BC. The regimen allows to achieve a high frequency of pCR, despite the large proportion of patients with locally advanced breast cancer. The high frequency of rare mutations in the BRCA1/2 genes and the potential therapeutic signifiance of this marker in residual disease treatment dictates the need for NGS in all patients with TN in breast cancer.

Joint analysis of WES and RNA-Seq identify signature genes related to metastasis in prostate cancer.

Prostate cancer (PCa) has a certain degree of heritability, and metastasis occurs as cancer progresses. However, its underlying mechanism remains largely unknown. We sequenced four cases of cancer without metastasis, four metastatic cancer, and four benign hyperplasia tissues as controls. A total of 1839 damaging mutations were identified. Pathway analysis, gene clustering, and weighted gene co-expression network analysis were employed to find characteristics associated with metastasis. Chr19 had the most mutation density and 1p36 had the highest mutation frequency across the genome. These mutations occurred in 1630 genes, including the most frequently mutated genes TTN and PLEC, and dozens of metastasis-related genes, such as FOXA1, NCOA1, CD34, and BRCA2. Ras signalling and arachidonic acid metabolism were uniquely enriched in metastatic cancer. Gene programmes 10 and 11 showed the signatures indicating the occurrence of metastasis better. A module (135 genes) was specifically associated with metastasis. Of them, 67.41% reoccurred in program 10, with 26 genes further retained as the signature genes related to PCa metastasis, including AGR3, RAPH1, SOX14, DPEP1, and UBL4A. Our study provides new molecular perspectives on PCa metastasis. The signature genes and pathways could be served as potential therapeutic targets for metastasis or cancer progression.

High Frequency of Antiviral Resistance Mutations in HBV Genotypes A2 and H: Multidrug Resistance Strains in Mexico.

Lamivudine (3TC), telbivudine (LdT), entecavir (ETV), adefovir (ADF), and tenofovir (TFV) are drugs used to treat hepatitis B virus (HBV) infection, but specific mutations allow some viruses to become resistant to antiviral drugs or to acquire immune escape capacities. These mutations have not been thoroughly investigated in Mexico. This study aimed to estimate the prevalence of HBV antiviral resistance and escape mutations. This cross-sectional study analyzed 158 samples. HBV DNA was extracted, amplified, and sequenced in serum samples using the spin column method, PCR assay, and Sanger's sequencing, respectively. HBV genotypes were determined, and HBV mutations were tested using the Geno2pheno tool. Overall, 68.4% (108/158) of HBV patients were infected with genotype H, followed by G (11.4%, 18/158), A2 (10.8%, 17/158), F1b (6.9.0%, 11/158), D (1.9%, 3/158), and E (0.6%, 1/158), and 5.1% (8/158) had evidence of recombination. The prevalence of resistance mutations was 8.2% (13/158) and the most common combined mutation was rt180M+rt204V. Notably, we found the combinations rt180M+rt204V+rt173L (n=2) and rt180M+rt204V+rt202G (n=1) that confer multidrug resistance to 3TC, LdT, and ETV. Resistance mutations were found in genotypes A2 (11.8%, 2/17), and H (10.2%, 11/108), and escape mutations were detected in HBV genotypes A2 (11.8%, 2/17), H (10.2%, 11/108), F1b (9.1%, 1/11) and G (5.6%, 1/18). The highest prevalence of antiviral resistance mutations or escape mutations was detected in HBV genotypes A2 and H. The earliest cases of HBV multidrug resistance were detected in Mexico.

Integrative pan-cancer analysis reveals the importance of PAQR family in lung cancer.

The progestin and adipoQ receptors (PAQRs) family contains 11 genes involved in the regulation of metabolism and cancer development. However, a comprehensive understanding of the role of PAQRs in cancer remains largely scarce, and the associations between their expression levels and immune signatures also need to be researched. Here, we applied pan-cancer analysis to explore the associations between PAQRs expression and survival, tumor microenvironment (TME), and drug sensitivity from the UCSC Xena and CellMiner databases. Besides, we further studied the expression, survival and somatic mutations of PAQRs in lung cancer (LC) from TCGA database. The results showed that PAQRs had significant heterogeneity with some upregulation and some downregulation in most tumors. Specifically, compared with PAQR3/5/6/9 and MMD2, ADIPOR1/2, PAQR4/7/8 and MMD had higher levels of average expression in all tumor types. PAQRs expression was greatly correlated with survival, immune subtypes, TME, and drug sensitivity. Furthermore, this research concentrated on analyzing the relationship of PAQRs expression with LC prognosis, and proved that ADIPOR2, PAQR4/9 and MMD were independent prognostic factors for LC patients. Finally, based on somatic mutation data, the genetic mutations in LC patients were majorly missense mutations, and TP53 and TTN had the top two highest mutation frequencies. Collectively, PAQRs may serve as robust biomarkers to predict the prognosis and guide immunotherapy of tumors, especially LC, which enables novel ways for improving cancer treatment.

Molnupiravir Inhibits Porcine Epidemic Diarrhea Virus Infection In Vitro.

Porcine epidemic diarrhea virus (PEDV) is a swine coronavirus that is highly infectious and prone to variation. Vaccines derived from traditional PEDV strains provide less protection against PEDV-variant strains. Furthermore; there is a complex diversity of sequences among various PEDV-variant strains. Therefore; there is an urgent need to develop alternative antiviral strategies to defend against PEDV. Molnupiravir is a nucleotide analogue that could replace natural nucleosides to restrain viral RNA replication. Our study provided evidence for the dose-dependent inhibition of PEDV replication by molnupiravir in Vero cells. Molnupiravir also exhibited a strong inhibitory effect on viral RNA and protein production. Our results demonstrated that molnupiravir inhibits PEDV RNA-dependent RNA polymerase (RdRp) activity and induces a high frequency of mutations in the PEDV genome. Further studies revealed that molnupiravir can reverse changes in the transcriptome caused by viral infection. In conclusion, our results indicated that molnupiravir has the potential to be an effective treatment for PEDV infection.

Intratumoral heterogeneity as a predictive biomarker in immunotherapy for advanced head and neck squamous cell carcinoma.

e18028 Background: Intratumoral heterogeneity (ITH) is associated with the poor prognosis in a variety of solid tumors, because of the therapeutic failure and drug resistance. However, we still lack understanding of the relationship between ITH and immunotherapy for advanced head and neck squamous cell carcinoma (HNSCC). Methods: We used the Bioconductor R package Maftools to analyze somatic variants of HNSCC from MSK-IMPACT-1611 study cohort (PMID: 30643254) with data from 68 patients. Then, a mutant-allele tumor heterogeneity (MATH) algorithm was used to measure ITH score. Finally, we analyzed the difference in the genomes, and the association with tumor mutational burden (TMB) between the ITH score high (ITH-H) and ITH score low (ITH-L) groups, and the effect of ITH on the prognosis of immunotherapy. Results: There was no significant difference in the high frequency mutated genes between the high ITH and low ITH groups. The genes with the highest mutation frequency in ITH-H group were TP53 (90.48%), TERT (47.62%), CDKN2A (38.1%), FAT1 (33.33%), NOTCH1 (28.57%), KMT2D (19.05%), APC (14.29%), and ASXL1 (14.29%), while the genes with the highest mutation frequency in ITH-L group were TP53 (44.68%), PIK3CA (38.3%), TRET (36.17%), FAT1 (21.28%), NOTCH1 (19.15%), KMT2C (17.02%), KMT2D (17.02%), and CDKN2A (14.89%). The median overall survival (mOS) of ITH-L group was significantly higher than that of ITH-H group (not reach versus 10 months, hazard ratio (HR) = 2.15, 95%CI 1.05−4.4, P = 0.032). There was no significant difference in median TMB levels between ITH-L and ITH-H groups (P = 0.55). The TMB high (TMB-H) patients had no significantly better mOS than the TMB low (TMB-L) group (HR = 0.54, 95%CI 0.25−1.16, P = 0.11). A pooled analysis of ITH and TMB show that the ITH-L+TMB-H group had significantly better improvement mOS than that in the ITH-L+TMB-L, ITH-H+TMB-H, and ITH-H+TMB-L group (HR = 0.71, 95%CI 0.13−1.35, P = 0.048). Conclusions: ITH may serve as a positive predictor of immunotherapy in patients with advanced HNSCC and is independent of TMB.

Intratumoral heterogeneity as a predictive biomarker in immunotherapy for advanced esophageal cancer.

e16059 Background: Previous studies have shown that the acquired resistance to anti-cancer therapy is largely due to intratumoral heterogeneity (ITH), thus leading to unfavourable prognosis. As known, there is no comprehensive research using ITH as a biomarker to predict the efficacy of immunotherapy in advanced esophageal cancer (ESCA) patients. Methods: Bioconductor R package Maftools was used to analyze somatic variants of advanced ESCA from MSK-IMPACT-1611 study cohort (PMID: 30643254) with data from 39 patients. We then used a mutant-allele tumor heterogeneity (MATH) algorithm to measure ITH score and analyzed the difference in the genetic profile between the ITH score high (ITH-H) and ITH score low (ITH-L) groups, the association with tumor mutational burden (TMB), and the effect of ITH on the prognosis of immunotherapy. Results: There were significant differences in gene mutation spectrum between ITH-H and ITH-L groups. The genes with the highest mutation frequency in ITH-H group were TP53 (93.75%), CDKN2A (37.5%), PREX2 (31.25%), PTPRD (25%), ALK (18.75%), ERBB4 (18.75%), and KMT2D (18.75%), while the genes with the highest mutation frequency in ITH-L group were TP53 (69.57%), CDKN2A (21.74%), PIK3CA (21.74%), APC (17.39%), ARID1A (17.39%), LATS1 (17.39%), and RNF43 (17.39%).There was significant difference in median overall survival (mOS) between ITH-L and ITH-H groups (not reach versus 5 months, hazard ratio (HR) = 3.96, 95%CI 1.02−15.39, P = 0.034). There was no significant difference in median TMB levels between ITH-L and ITH-H groups (7 Muts/Mb versus 8 Muts/Mb, P = 0.09). The TMB high (TMB-H) patients had no significantly better mOS than the TMB low (TMB-L) group (HR = 0.27, 95%CI 0.06−1.26, P = 0.072). A pooled analysis of ITH and TMB show that mOS in the ITH-L+ TMB-H group was significantly better than that in the ITH-L+TMB-L, ITH-H+TMB-H, and ITH-H+TMB-L group (HR = 2.03, 95%CI 0.06−7.69, P = 0.002). Conclusions: The results suggest ITH is a predictor of ESCA immunotherapy efficacy independent of TMB.