Base editing strategies to convert CAG to CAA diminish the disease-causing mutation in Huntington's disease.

An expanded CAG repeat in the huntingtin gene (HTT) causes Huntington's disease (HD). Since the length of uninterrupted CAG repeat, not polyglutamine, determines the age-at-onset in HD, base editing strategies to convert CAG to CAA are anticipated to delay onset by shortening the uninterrupted CAG repeat. Here, we developed base editing strategies to convert CAG in the repeat to CAA and determined their molecular outcomes and effects on relevant disease phenotypes. Base editing strategies employing combinations of cytosine base editors and guide RNAs (gRNAs) efficiently converted CAG to CAA at various sites in the CAG repeat without generating significant indels, off-target edits, or transcriptome alterations, demonstrating their feasibility and specificity. Candidate BE strategies converted CAG to CAA on both expanded and non-expanded CAG repeats without altering HTT mRNA and protein levels. In addition, somatic CAG repeat expansion, which is the major disease driver in HD, was significantly decreased in the liver by a candidate BE strategy treatment in HD knock-in mice carrying canonical CAG repeats. Notably, CAG repeat expansion was abolished entirely in HD knock-in mice carrying CAA-interrupted repeats, supporting the therapeutic potential of CAG-to-CAA conversion strategies in HD and potentially other repeat expansion disorders.

Personalized allele-specific CRISPR-Cas9 strategies for myofibrillar myopathy 6.

Myofibrillar myopathy 6 (MFM6) is a rare childhood-onset myopathy characterized by myofibrillar disintegration, muscle weakness, and cardiomyopathy. The genetic cause of MFM6 is p.Pro209Leu mutation (rs121918312-T) in the BAG3 gene, which generates the disease outcomes in a dominant fashion. Since the consequences of the BAG3 mutation are strong and rapidly progressing, most MFM6 patients are due to de novo mutation. There are no effective treatments for MFM6 despite its well-known genetic cause. Given p.Pro209Leu mutation is dominant, regenerative medicine approaches employing orthologous stem cells in which mutant BAG3 is inactivated offer a promising avenue. Here, we developed personalized allele-specific CRISPR-Cas9 strategies capitalizing on PAM-altering SNP and PAM-proximal SNP. In order to identify the disease chromosome carrying the de novo mutation in our two affected individuals, haplotype phasing through cloning-sequencing was performed. Based on the sequence differences between mutant and normal BAG3, we developed personalized allele-specific CRISPR-Cas9 strategies to selectively inactivate the mutant allele 1) by preventing the transcription of the mutant BAG3 and 2) by inducing nonsense-mediated decay (NMD) of mutant BAG3 mRNA. Subsequent experimental validation in patient-derived induced pluripotent stem cell (iPSC) lines showed complete allele specificities of our CRISPR-Cas9 strategies and molecular consequences attributable to inactivated mutant BAG3. In addition, mutant allele-specific CRISPR-Cas9 targeting did not alter the characteristics of iPSC or the capacity to differentiate into cardiomyocytes. Together, our data demonstrate the feasibility and potential of personalized allele-specific CRISPR-Cas9 approaches to selectively inactivate the mutant BAG3 to generate cell resources for regenerative medicine approaches for MFM6.

Optimal Hyperspectral Band Selection for Tissue Oxygenation Mapping with Generative Adversarial Network.

Tissue oxygenation assessment using hyperspectral imaging is an emerging technique for the diagnosis and pre- and post-treatment monitoring of ischemic patients. However, the high spectral resolution of hyperspectral imaging leads to large data sizes and a long imaging time. In this study, we propose a method that utilizes multi-objective evolutionary algorithms to determine the optimal hyperspectral band combination when developing a deep learning model for predicting tissue oxygenation from hyperspectral images. Our results confirm that the deep learning model effectively predicts tissue oxygenation images for various oxygenation states. Moreover, we demonstrate that a high-performance prediction model can be developed using only a small number of spectral bands, indicating the potential for more efficient non-contact tissue oxygenation mapping with the proposed method.Clinical Relevance- The proposed method allows for the non-contact and efficient acquisition of two-dimensional tissue oxygenation information in various oxygenation states.

Base editing strategies to convert CAG to CAA diminish the disease-causing mutation in Huntington's disease.

An expanded CAG repeat in the huntingtin gene ( HTT ) causes Huntington's disease (HD). Since the length of uninterrupted CAG repeat, not polyglutamine, determines the age-at-onset in HD, base editing strategies to convert CAG to CAA are anticipated to delay onset by shortening the uninterrupted CAG repeat. Here, we developed base editing strategies to convert CAG in the repeat to CAA and determined their molecular outcomes and effects on relevant disease phenotypes. Base editing strategies employing combinations of cytosine base editors and gRNAs efficiently converted CAG to CAA at various sites in the CAG repeat without generating significant indels, off-target edits, or transcriptome alterations, demonstrating their feasibility and specificity. Candidate BE strategies converted CAG to CAA on both expanded and non-expanded CAG repeats without altering HTT mRNA and protein levels. In addition, somatic CAG repeat expansion, which is the major disease driver in HD, was significantly decreased by a candidate BE strategy treatment in HD knock-in mice carrying canonical CAG repeats. Notably, CAG repeat expansion was abolished entirely in HD knock-in mice carrying CAA-interrupted repeats, supporting the therapeutic potential of CAG-to-CAA conversion base editing strategies in HD and potentially other repeat expansion disorders.

Loss of HRD functional phenotype impedes immunotherapy and can be reversed by HDAC inhibitor in ovarian cancer.

In recent years, homologous recombination deficiency (HRD) has not achieved the expected substantial promotion of immunotherapeutic efficacy in ovarian cancer. This study aims to explore the role of HRD functional phenotype as a powerful biomarker in identifying HRD patients who may benefit from immunotherapy. HRD functional phenotype, namely HRD-EXCUTE, was defined as the average level of the 15 hub genes upregulated in HRD ovarian cancer. A decision tree was plotted to evaluate the critical role of HRD-EXCUTE in HRD patients. Agents inducing HRD-EXCUTE were identified by CMAP web (Connectivity Map). The mechanisms and immunotherapeutic effect of PARPi and HDACi in promoting HRD-EXCUTE was examined in vitro and in vivo. The decision tree plotted on the basis of HRD and HRD-EXCUTE indicated the HRD patients without the HRD functional phenotype were largely unresponsive to immunotherapy, which was validated by the immunotherapeutic cohorts. Furthermore, loss of HRD-EXCUTE in the HRD patients attenuated immunogenicity and inhibited immune cells in tumor microenvironment. Moreover, Niraparib combined with Entinostat induced HRD-EXCUTE by activating the cGAS-STING pathway and increasing the histone acetylation. The combination therapy could enhance the cytotoxicity of immune cells, and promote pro-immune cells infiltrating into ascites, resulting in inhibited ovarian cancer growth. The HRD functional phenotype HRD-EXCUTE was set up as a potent biomarker to identify whether HRD patients can benefit from immunotherapy. Loss of HRD-EXCUTE in HRD patients were largely insensitive to immunotherapy. The combination of PARPi with HDACi could improve the efficacy of the PARPi-based immunotherapy in ovarian cancer by augmenting the HRD functional phenotype.

Effects of breast milk intake ratio during hospitalization on antibiotic therapy duration in preterm infants less than 34 gestational weeks: a multicenter retrospective cohort study / 中华围产医学杂志

Objective:To investigate the effects of breast milk to total milk intake ratio during hospitalization on the duration of antibiotic therapy in preterm infants less than 34 weeks of gestation.Methods:Clinical data of preterm infants ( n=1 792) less than 34 gestational weeks were retrospectively collected in 16 hospitals of Jiangsu Province Neonatal-Perinatal Cooperation Network from January 1, 2019, to December 31, 2021. The days of therapy (DOT) were used to evaluate the duration of antibiotic administration. The median DOT was 15.0 d (7.0-27.0 d). The patients were divided into four groups based on the quartiles of DOT: Q 1 (DOT≤7.0 d), Q 2 (7.0 d<DOT≤15.0 d), Q 3 (15.0 d<DOT≤27.0 d) and Q 4 (DOT>27.0 d) groups. According to the breast milk intake ratio (breast milk intake to total milk intake during hospitalization×100%), they were also divided into four groups: very-low-ratio breastfeeding group (breast milk intake ratio≤25%), low-ratio breastfeeding group (25%<breast milk intake ratio≤50%), medium-ratio breastfeeding group (50%<breast milk intake ratio≤75%) and high-ratio breastfeeding group (breast milk intake ratio>75%). Univariate analysis ( Chi-square test and Kruskal-Wallis rank-sum test) was used to analyze the factors influencing DOT. Spearman correlation analysis and trend Chi-square test were used to explore the relationship between breast milk intake ratio and DOT. After using multiple imputations to address missing data, two models were constructed after adjusting for different factors, and multinomial logistic regression model was applied to evaluate the effects of the breast milk intake ratio on DOT. Finally, sensitivity analysis was conducted to assess the stability of the models. Results:(1) Of the 1 792 preterm infants, there were 507 (28.3%) in the Q 1 group, 422 (23.5%) in the Q 2 group, 438 (24.4%) in the Q 3 group and 425 (23.7%) in the Q 4 group. (2) The median values of DOT in the very-low-ratio, low-ratio, medium-ratio and high-ratio breastfeeding groups were 20.0 d (11.0-31.0 d), 20.0 d (11.0-32.0 d), 13.0 d (6.0-25.8 d) and 10.0 d (4.0-21.0 d), respectively. Compared with the very-low-ratio and low-ratio breastfeeding groups, the medium-ratio and high-ratio breastfeeding groups had shorter DOT (all P<0.05). (3) After adjusting for factors with P<0.1 (prenatal glucocorticoid exposure, antimicrobial use within 24 h before delivery, gestational age at delivery, birth weight, Apgar score≤7 at 1 min, neonatal respiratory distress syndrome, infectious pneumonia and early-onset neonatal sepsis) between the DOT quartile groups, it showed that medium-ratio and high-ratio breastfeeding were protective factors in contrast to very-low-ratio breastfeeding in the Q 2, Q 3 and Q 4 groups as compared with the Q 1 group [Q 2 group: OR=0.50 (95% CI: 0.30-0.85) and OR=0.36 (95% CI: 0.26-0.51); Q 3 group: OR=0.31 (95% CI: 0.18-0.55) and OR=0.20 (95% CI: 0.14-0.29); Q 4 group: OR=0.22 (95% CI: 0.12-0.42) and OR=0.17 (95% CI: 0.12-0.26)]. Conclusion:Breast milk intake accounting for over 50% of total milk intake has a positive impact on reducing DOT in premature infants requiring antibiotics, which suggests that breastfeeding should be actively encouraged.

Analysis of disease burden and risk factors of diabetic kidney disease in China from 1990 to 2019 / 中华肾脏病杂志

Objective:To explore the changes of disease burden and risk factors of chronic kidney disease (CKD) due to type 1 and type 2 diabetes mellitus in China from 1990 to 2019, and to provide reference data for the prevention and control of diabetic kidney disease (DKD).Methods:The Chinese DKD data were obtained from the 2019 Global Burden of Disease (GBD) database. The morbidity, prevalence, mortality, years lived with disability (YLD), years of life lost (YLL), and disability-adjusted life year (DALY) were used to compare the disease burden of CKD due to type 1 and type 2 diabetes mellitus from 1990 to 2019. In addition, the risk factors of DKD were analyzed.Results:The numbers of CKD patients due to type 1 and type 2 diabetes mellitus in China were 574 (95% UI 495-665) and 31 076 (95% UI 28 152-33 909) thousand, and the numbers of new cases were 9 (95% UI 8-11) and 434 (95% UI 390-481) thousand in 2019, respectively. The numbers of death were 13 (95% UI 8-18) and 63 (95% UI 50-77) thousand, respectively. The age groups with the largest number of patients and new cases of CKD due to type 1 diabetes mellitus were 30-34 years old and <5 years old, respectively. The age group with the largest number of patients and new cases of CKD due to type 2 diabetes mellitus were 50-54 years old and 70-74 years old, respectively. From 1990 to 2019, the age-standardized prevalence rate of DKD patients in China was relatively stable, but the age-standardized incidence rate and YLD rate showed an upward trend, while the age-standardized mortality rate, YLL rate, and DALY rate showed a downward trend. The main risk factors associated with DKD death were high fasting plasma glucose, kidney dysfunction, high systolic blood pressure, high body mass index, high sodium diet, and lead exposure. The proportions of DKD death caused by high systolic blood pressure and high body mass index in the Chinese population were still increasing. Conclusions:From 1990 to 2019, the age-standardized incidence and YLD rate of DKD in China shows an upward trend, while the age-standardized prevalence rate is relatively stable, and the age-standardized mortality rate, YLL rate, and DALY rate show a decreasing trend. High fasting glucose, renal failure, high systolic blood pressure, high body mass index, high sodium diet, and lead exposure are risk factors associated with death in DKD patients. With the progress of aging, the disease burden of DKD in China will continuously increase. Future work should be focused on population-specific interventions, taking into consideration the risk factors identified within the study.

Impact of atrial fibrillation on in-hospital adverse outcomes in elderly patients with acute pulmonary thromboembolism / 中华老年医学杂志

Objective:To analyze the clinical characteristics of elderly acute pulmonary thromboembolism(APE)patients complicated with preexisting atrial fibrillation(AF)and the impact of preexisting AF on in-hospital adverse outcomes in elderly patients with APE.Methods:A retrospective analysis was performed on elderly APE patients with preexisting AF hospitalized in Beijing Anzhen Hospital, Capital Medical University between January 1, 2008 and December 31, 2021.We compared the comorbidities, symptoms, signs, laboratory test results and echocardiographic features, simplified pulmonary embolism severity index(sPESI)scores and adverse in-hospital outcomes between the preexisting AF group and the non-AF group.Logistic regression was used to analyze the risk factors of in-hospital adverse outcomes in elderly patients with APE.Results:A total of 240 patients diagnosed with APE were enrolled.There were 120 patients in the AF group and 120 patients in the non-AF group.For patients in the AF group and the non-AF group, the proportions with chronic heart failure were 38.3%(46/120)and 15.8%(19/120), the proportions with lower extremity deep vein thrombosis(DVT)were 36.7%(44/120)and 65.8%(79/120), the left ventricular ejection fractions(LVEF)were(59±10)% and(62±7)%, and hospital stays were(15±7)and(11±4)days, respectively, and the differences were statistically significant( χ2=15.381, 20.429, t=2.527, -4.710, all P<0.05). The incidences of in-hospital adverse outcomes in the AF group and the non-AF group were 4.2%(5/120)and 3.3%(4/120), respectively, with no significant difference( χ2=0.000, P=1.000). The overall incidence of in-hospital adverse outcomes was 3.8%(9/240). Multivariate Logistic regression analysis showed that elevated lactic acid was an independent risk factor for in-hospital adverse outcomes( OR=2.753, 95% CI: 1.367-5.542, P=0.005). However, AF( OR=2.880, 95% CI: 0.587-14.141, P=0.192)and sPESI score( OR=2.056, 95% CI: 0.904-4.673, P=0.086)were not associated with in-hospital adverse outcomes. Conclusions:Elderly APE patients with preexisting AF have a relatively low incidence of DVT, but a higher proportion have concurrent chronic heart failure and need a longer hospital stay.Elevated lactic acid is an independent risk factor for in-hospital adverse outcomes of elderly APE patients with preexisting AF.However, preexisting AF has no predictive value for in-hospital adverse outcomes in elderly patients with APE.

Research progress in emergency drug stockpile for nuclear and radiological accidents / 中华放射医学与防护杂志

In the current international situation, there is the possibility of nuclear war and nuclear terrorism. The nuclear and radiological emergency in the event of an accident should be enhanced. The World Health Organization (WHO) issued a publication on January 27, 2023, updating the list of drugs recommended for stockpiling in response to radiation and nuclear emergencies, which was the first update since 2007. The list of recommended drug stocks in the publication includes drugs to prevent or reduce the effects of radiation, as well as drugs used to treat injuries after exposure. Based on the list of drugs, this article reviews the emergency response to nuclear and radiological accidents, the general situation of emergency drug stockpiles, drug reserves in some countries, and current considerations, with a view to providing references for emergency medical rescue in nuclear and radiological accidents in China.

Radioactivity in marine organisms around Qinshan Nuclear Power Plant / 中国辐射卫生

@#<b>Objective</b> To determine the radioactive enrichment level in marine biological media around Qinshan Nuclear Power Plant and to evaluate the impact of effluent on marine biological media in the surrounding sea area based on the discharge of effluent from the nuclear power plant over the years and the monitoring level of surrounding marine media. <b>Methods</b> The monitoring data of ³H, ¹⁴C, ⁹⁰Sr, ¹³⁷Cs, and ^110mAg in the environmental media monitored in Qinshan Nuclear Power Plant from 2008 to 2017 were collected, and the data were processed and analyzed by IBM SPSS Statistics 26. <b>Results</b> The activity concentrations of tritium in mullet and oyster in the sea area around Qinshan Nuclear Power Plant were 0.5-2.9 Bq/kg and 0.55-8.2 Bq/kg, respectively, and the activity concentrations of tritium in both increased generally under the influence of effluent from the nuclear power plant. The activity concentrations of ¹⁴C, ¹³⁷Cs, and ⁹⁰Sr in the monitored organisms were lower than the background values. The results of correlation analysis showed that the effluent from the nuclear power plant had almost no effect on ¹⁴C. The activity level of ^110mAg decreased significantly year by year in oyster, and remained below the background level in mullet. <b>Conclusion</b> Qinshan Nuclear Power Plant is the longest operating nuclear power plant in mainland China. Most of the nuclides in biological media in the surrounding sea area are lower than the background levels, and the effluents from the nuclear power plant have little influence on marine biological media in the surrounding sea area.

Acupuncture combined with auricular point sticking for girls aged 3-8 years with incomplete precocious puberty: a randomized controlled trial / 中国针灸

OBJECTIVE@#To observe the efficacy and safety of acupuncture combined with auricular point sticking for girls aged 3-8 years with incomplete precocious puberty (IPP).@*METHODS@#Sixty girls with IPP were randomly divided into an observation group (30 cases, 2 cases dropped off) and a control group (30 cases, 2 cases were eliminated). The girls in the control group were treated with healthy diet and proper exercise for 12 weeks. On the basis of the treatment in the control group, the girls in the observation group were treated with acupuncture combined with auricular point sticking. The acupuncture was applied at Sanyinjiao (SP 6), Guanyuan (CV 4), Guilai (ST 29), etc., the needles were retained for 20 min, acupuncture was given twice a week (once every 3 days). The auricular point sticking was applied at Luanchao (TF2), Neishengzhiqi (TF2), Neifenmi (CO18), Yuanzhong (AT2,3,4i), etc., twice a week. The treatment was given for 12 weeks. Before treatment, after treatment and in follow-up after 12 weeks of treatment completion, the Tanner stage of breast, serum contents of sex hormone (luteinizing hormone [LH], follicle-stimulating hormone [FSH], estradiol [E2]) were observed. The ovarian volume, the number of follicles with diameter>4 mm, and the uterine volume were measured by abdominal color Doppler ultrasound. In addition, the safety of the observation group was evaluated.@*RESULTS@#Compared with before treatment, the Tanner stage of breast in the observation group was improved after treatment and in follow-up (P<0.05); after treatment and in follow-up, the Tanner stage of breast in the observation group was better than that in the control group (P<0.05). Compared with before treatment, the serum levels of LH and E2 in the observation group were increased (P<0.05), and the volume of bilateral ovaries was larger (P<0.05) in follow-up. Compared with before treatment, the serum contents of LH, FSH and E2 in the control group were increased (P<0.05), the volume of bilateral ovaries was larger (P<0.05), and the number of follicles was increased (P<0.05) after treatment and in follow-up. The serum levels of LH, FSH and E2 in the observation group were lower than those in the control group (P<0.05), the volume of bilateral ovaries was smaller than that in the control group (P<0.05), and the number of follicles was lower than that in the control group (P<0.05). Compared with before treatment, the uterine volume in the two groups was larger in follow-up (P<0.05). There was no statistically significant difference between the two groups after treatment and in follow-up (P>0.05). During the treatment, 3 cases in the observation group had slight abdominal pain and subcutaneous blood stasis, without serious adverse reactions.@*CONCLUSION@#Acupuncture combined with auricular point sticking could improve the Tanner stage of breast, reduce the level of sex hormone, slow down the development and maturation of ovary and follicle, and control the degree and speed of sexual development in girls aged 3-8 years with IPP.

Further progress of the etiology,diagnosis and treatment of peripheral precocious puberty / 中华预防医学杂志

Peripheral precocious puberty(PPP),also known as puberty independent from hypothalamic-pituitary axis activation,is stimulated by hormones from other sources, with only partial sexual characteristics development but without mature sexual function. The secondary sexual characteristics development occurs before 7.5 years of age in girls and before 9 years of age in boys. Clinical manifestations are diverse, and PPP has varied etiology including congenital adrenal hyperplasia, McCune-Albright syndrome, ovarian cyst, adrenal tumor, ovarian tumor, testicular tumor, human chorionic gonadotropin producing tumor, familial male precocious puberty, aromatase excess syndrome, and environmental estrogen. Early identification of etiology, accurate differential diagnosis and prenatal gene screening play a significant role in the prevention, diagnosis and treatment of the disease.

Research progresses on interventions of obesity in children and adolescents / 中华预防医学杂志

Childhood and adolescent obesity has become a global epidemic. The interventions mainly include lifestyle intervention, medication treatment and bariatric surgery. Among them, lifestyle intervention, especially intensive lifestyle intervention with participation of family members, is the first-line treatment for obesity in children and adolescents. Both medication and bariatric surgery are adjuvant treatments for severely obese children and adolescents. Currently, metformin is the most widely used drug for the treatment of obesity in children and adolescents in both China and other countries; orlistat and liraglutide are also the drugs that are safe and often used in other countries; other drugs are not recommended. As a tertiary prevention and treatment strategy for obesity, bariatric surgery should be carried out on the basis of good compliance from both the children and their family members, with the cooperation of multiple disciplines. Sleeve gastrectomy (SG) and Roux-en-Y gastric bypass (RYGB) are the most common types of procedure performed. Meanwhile, as a new treatment method, intra-gastric balloon procedure needs to be paid more attention to its efficacy and safety.

PAM-altering SNP-based allele-specific CRISPR-Cas9 therapeutic strategies for Huntington's disease.

Huntington's disease (HD) is caused by an expanded CAG repeat in huntingtin (HTT). Since HD is dominant and loss of HTT leads to neurological abnormalities, safe therapeutic strategies require selective inactivation of mutant HTT. Previously, we proposed a concept of CRISPR-Cas9 using mutant-specific PAM sites generated by SNPs to selectively inactivate mutant HTT. Aiming at revealing suitable targets for clinical development, we analyzed the largest HD genotype dataset to identify target PAM-altering SNPs (PAS) and subsequently evaluated their allele specificities. The gRNAs based on the PAM sites generated by rs2857935, rs16843804, and rs16843836 showed high levels of allele specificity in patient-derived cells. Simultaneous use of two gRNAs based on rs2857935-rs16843804 or rs2857935-rs16843836 produced selective genomic deletions in mutant HTT and prevented the transcription of mutant HTT mRNA without impacting the expression of normal counterpart or re-integration of the excised fragment elsewhere in the genome. RNA-seq and off-target analysis confirmed high levels of allele specificity and the lack of recurrent off-targeting. Approximately 60% of HD subjects are eligible for mutant-specific CRISPR-Cas9 strategies of targeting one of these three PAS in conjunction with one non-allele-specific site, supporting high applicability of PAS-based allele-specific CRISPR approaches in the HD patient population.

Allele-specific silencing of the gain-of-function mutation in Huntington's disease using CRISPR/Cas9.

Dominant gain-of-function mechanisms in Huntington's disease (HD) suggest that selective silencing of mutant HTT produces robust therapeutic benefits. Here, capitalizing on exonic protospacer adjacent motif-altering (PAM-altering) SNP (PAS), we developed an allele-specific CRISPR/Cas9 strategy to permanently inactivate mutant HTT through nonsense-mediated decay (NMD). Comprehensive sequence/haplotype analysis identified SNP-generated NGG PAM sites on exons of common HTT haplotypes in HD subjects, revealing a clinically relevant PAS-based mutant-specific CRISPR/Cas9 strategy. Alternative allele of rs363099 (29th exon) eliminates the NGG PAM site on the most frequent normal HTT haplotype in HD, permitting mutant-specific CRISPR/Cas9 therapeutics in a predicted ~20% of HD subjects with European ancestry. Our rs363099-based CRISPR/Cas9 showed perfect allele specificity and good targeting efficiencies in patient-derived cells. Dramatically reduced mutant HTT mRNA and complete loss of mutant protein suggest that our allele-specific CRISPR/Cas9 strategy inactivates mutant HTT through NMD. In addition, GUIDE-Seq analysis and subsequent validation experiments support high levels of on-target gene specificity. Our data demonstrate a significant target population, complete mutant specificity, decent targeting efficiency in patient-derived cells, and minimal off-target effects on protein-coding genes, proving the concept of PAS-based allele-specific NMD-CRISPR/Cas9 and supporting its therapeutic potential in HD.

HHLA2 predicts improved prognosis of anti-PD-1/PD-L1 immunotherapy in patients with melanoma.

Background: As a recognized highly immunogenic tumor, immune checkpoint blockades (ICB) have been widely used as a systemic treatment option for melanoma. However, only about half of treated patients could benefit from it in Caucasians, and only about 15% in Chinese melanoma patients. Robust predictive biomarkers are needed. HHLA2, a new-found member of B7 family, is generally expressed in kinds of tumors, such as melanoma. This study focuses on illustrating the prognostic value of HHLA2 in melanoma immunotherapy and its association with tumor-infiltrating lymphocytes. Methods: HHLA2 expression in pan-cancer and the association with prognosis and immune microenvironment were identified by analyzing gene expression profiles from TCGA database with selected bioinformatics tools and methods. Tumor tissues from 81 cases with advanced and unresectable melanoma were collected for detecting HHLA2 and CD8 levels by immunohistochemistry. Results: HHLA2 was found to be ubiquitously expressed in pan-cancer with high level and correlate with the prognosis of patients. Further comprehensive analysis from TCGA database demonstrated that the highly expressed HHLA2 was remarkably correlated with better prognosis, high infiltration status of various immune-active cells and immune activated pathways in skin cutaneous melanoma (SKCM). Moreover, immunohistochemistry (IHC) analyses of FFPE tissue from melanoma patients revealed that HHLA2 high expression was strongly related to improved response to ICB and indicated a longer progression-free survival (PFS) and overall survival (OS). Besides, HHLA2 expression was found to have a positive association with the density of CD8+ TILs. Conclusion: Our findings revealed that high expression of HHLA2 has important values in predicting the response to ICB and indicating improved PFS and OS in patients with advanced and unresectable melanoma, suggesting that HHLA2 may serve as a costimulatory ligand in melanoma, which renders it as an ideal biomarker for immunotherapy.

[Paneth Cell Carcinoma of the Stomach].

Paneth cell carcinoma is a rare carcinoma composed predominantly or purely of malignant Paneth cells. An 83-year-old woman presented for evaluation of an elevated lesion in the stomach. On endoscopy, a 15 mm, discolored, elevated lesion with a central depression was found on the greater curvature of the gastric lower body. Endoscopic forceps biopsy revealed chronic gastritis with intestinal metaplasia. Magnifying endoscopy revealed an irregularly oval/tubular microsurface pattern and an irregular loop microvascular pattern with a demarcation line, suggestive of early gastric cancer. Therefore, endoscopic submucosal dissection was performed. Histopathological examination revealed a well-differentiated tubular adenocarcinoma limited to the muscularis mucosae and the tumor cells contained coarse eosinophilic granules in the cytoplasm. These tumor cells were diffusely and strongly stained for lysozyme, confirming the tumor diagnosis as Paneth cell carcinoma. Herein, we report a rare case of Paneth cell carcinoma and its endoscopic and histopathologic findings.

N-glycosylation stabilizes MerTK and promotes hepatocellular carcinoma tumor growth.

Despite the evidences of elevated expression of Mer tyrosine kinase (MerTK) in multiple human cancers, mechanisms underlying the oncogenic roles of MerTK in hepatocellular carcinoma (HCC) remains undefined. We explored the functional effects of MerTK and N-Glycosylated MerTK on HCC cell survival and tumor growth. Here, we show that MerTK ablation increases reactive oxygen species (ROS) production and promotes the switching from glycolytic metabolism to oxidative phosphorylation in HCC cells, thus suppressing HCC cell proliferation and tumor growth. MerTK is N-glycosylated in HCC cells at asparagine 294 and 454 that stabilizes MerTK to promote oncogenic transformation. Moreover, we observed that nuclear located non-glycosylated MerTK is indispensable for survival of HCC cells under stress. Pathologically, tissue microarray (TMA) data indicate that MerTK is a pivotal prognostic factor for HCC. Our data strongly support the roles of MerTK N-glycosylation in HCC tumorigenesis and suggesting N-glycosylation inhibition as a potential HCC therapeutic strategy.

Haplotype-specific insertion-deletion variations for allele-specific targeting in Huntington's disease.

Huntington's disease (HD) is a dominantly inherited neurodegenerative disease caused by an expanded CAG repeat in huntingtin (HTT). Given an important role for HTT in development and significant neurodegeneration at the time of clinical manifestation in HD, early treatment of allele-specific drugs represents a promising strategy. The feasibility of an allele-specific antisense oligonucleotide (ASO) targeting single-nucleotide polymorphisms (SNPs) has been demonstrated in models of HD. Here, we constructed a map of haplotype-specific insertion-deletion variations (indels) to develop alternative mutant-HTT-specific strategies. We mapped indels annotated in the 1000 Genomes Project data on common HTT haplotypes, revealing candidate indels for mutant-specific HTT targeting. Subsequent sequencing of an HD family confirmed candidate sites and revealed additional allele-specific indels. Interestingly, the most common normal HTT haplotype carries indels of big allele length differences at many sites, further uncovering promising haplotype-specific targets. When patient-derived cells carrying the most common HTT diplotype were treated with ASOs targeting the mutant alleles of candidate indels (rs772629195 or rs72239206), complete mutant specificity was observed. In summary, our map of haplotype-specific indels permits the identification of allele-specific targets in HD subjects, potentially contributing to the development of safe HTT-lowering therapeutics that are suitable for early treatment in HD.

Fatality assessment and variant risk monitoring for COVID-19 using three new hospital occupancy related metrics

BackgroundThough case fatality rate (CFR) is widely used to reflect COVID-19 fatality risk, its use is limited by large temporal and spatial variation. Hospital mortality rate (HMR) is also used to assess the severity of COVID-19, but HMR data is not directly available except 35 states of USA. Alternative metrics are needed for COVID-19 severity and fatality assessment. MethodsNew metrics and their applications in fatality measurements and risk monitoring are proposed here. We also introduce a new mathematical model to estimate average hospital length of stay for death (Ldead) and discharges (Ldis). Multiple data sources were used for our analysis. FindingsWe propose three new metrics, hospital occupancy mortality rate (HOMR), ratio of total deaths to hospital occupancy (TDHOR) and ratio of hospital occupancy to cases (HOCR), for dynamic assessment of COVID-19 fatality risk. Estimated Ldead and Ldis for 501,079 COVID-19 hospitalizations in US 34 states between Aug 7, 2020 and Mar 1, 2021 were 14.0 and 18.2 days, respectively. We found that TDHOR values of 27 countries are less spatially and temporally variable and more capable of detecting changes in COVID-19 fatality risk. The dramatic changes in COVID-19 CFR observed in 27 countries during early stages of the pandemic were mostly caused by undiagnosed cases. Compared to the first week of November, 2021, the week mean HOCRs (mimics hospitalization-to-case ratio) for Omicron variant decreased 34.08% and 65.16% in the United Kingdom and USA respectively as of Jan 16, 2022. InterpretationThese new and reliable measurements for COVID-19 that could be expanded as a general index to other fatal infectious diseases for disease fatality risk and variant-associated risk monitoring. Research in contextO_ST_ABSEvidence before this studyC_ST_ABSWe searched PubMed, medRxiv, and bioRxiv for peer-reviewed articles, preprints, and research reports on risk and health care evaluation for COVID-19 using the search terms "hospital occupancy mortality rate", "ratio of total deaths to hospital occupancy", "ratio of hospital occupancy to case" up to Jan 20, 2022. No similar concepts or studies were found. No similar mathematical models based on "hospital occupancy mortality rate" for the estimation of hospital length of stay for deaths and discharges have been identified to date. Added value of the studyOur new metrics, HOMR and TDHOR, mimic HMR for COVID-19 fatality risk assessment but utilize readily available data for many US states and countries around the world. HOCR mimics hospitalization-to-case ratio for COVID-19. We also provide evidence that explains why COVID-19 CFR has such dramatic changes at the beginning of a COVID-19 outbreak. We have additionally provided new metrics for COVID-19 fatality risk dynamic monitoring including Omicron variant and showed that these metrics provided additional information. Implications of all the available evidenceThe results of this study, including average hospital length of stay for deaths and discharges for over 500,000 COVID-19 hospitalizations in the US, can aid county, state, and national leaders in making informed public health decisions related to the ongoing COVID-19 pandemic. This is the first study to provide quantitative evidence to address why CFR has a such a large variation at the beginning of the COVID-19 pandemic in most countries and will hopefully encourage more countries to release hospital occupancy data, which we show is both useful and easy information to collect. The new metrics introduced by our study are effective indicators for monitoring COVID-19 fatality risk, as well as potentially fatal COVID-19 variants, and could also be expanded to other fatal infectious diseases.