Combination of AAV-delivered tumor suppressor PTEN with anti-PD-1 loaded depot gel for enhanced antitumor immunity

Recent clinical studies have shown that mutation of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) gene in cancer cells may be associated with immunosuppressive tumor microenvironment (TME) and poor response to immune checkpoint blockade (ICB) therapy. Therefore, efficiently restoring PTEN gene expression in cancer cells is critical to improving the responding rate to ICB therapy. Here, we screened an adeno-associated virus (AAV) capsid for efficient PTEN gene delivery into B16F10 tumor cells. We demonstrated that intratumorally injected AAV6-PTEN successfully restored the tumor cell PTEN gene expression and effectively inhibited tumor progression by inducing tumor cell immunogenic cell death (ICD) and increasing immune cell infiltration. Moreover, we developed an anti-PD-1 loaded phospholipid-based phase separation gel (PPSG), which formed an in situ depot and sustainably release anti-PD-1 drugs within 42 days in vivo. In order to effectively inhibit the recurrence of melanoma, we further applied a triple therapy based on AAV6-PTEN, PPSG@anti-PD-1 and CpG, and showed that this triple therapy strategy enhanced the synergistic antitumor immune effect and also induced robust immune memory, which completely rejected tumor recurrence. We anticipate that this triple therapy could be used as a new tumor combination therapy with stronger immune activation capacity and tumor inhibition efficacy.

The Clinical Study on Short-Term Efficacy of Pelvic Magnetic Stimulation Combined with Pelvic Muscle Biofeedback on Female Idiopathic Overactive Bladder.

Objective: To evaluate the short-term efficacy of pelvic magnetic stimulation combined with pelvic muscle biofeedback on female Idiopathic Overactive Bladder (IOAB). Methods: 96 cases of IOAB females were randomly divided into control group (magnetic stimulation treatment n=48) and observation group (magnetic stimulation with biofeedback n=48). All the patients were dealt with the sacralneuromagnetic stimulation (once, qod), with 5 times in total. Then the patients in observation group were processed with biofeedback (10 times). Overactive Bladder Symptom Score (OABSS), Patients Perception Bladder Condition (PPBC) and Incontinence Quality of Life Scale (I-QOL) were evaluated as the indexes. Results: The effective rate of control group and observation group respectively were 89.58% and 93.75%. There were significant differences (P=0.67). The OABSS and PPBC of two groups were decreased, I-QOL were increased after treatment (P<0.05). Difference was statistically significant in OABSS three months after treatment between the two groups (P=0.00).The recurrence rate of three months after treatment of the two groups were 18.75% and 6.38% (P=0.04). Conclusions: Both magnetic stimulation alone and magnetic stimulation with biofeedback were effective and safe in female patients with IOAB. Magnetic stimulation with biofeedback may reduce the recurrence rate and continue to improve the symptoms in a certain extent.

Clinical efficacy and pharmacoeconomic evaluation of bevacizumab or cetuximab combined with chemotherapy in the treatment of advanced colorectal cancer / 中国医师进修杂志

Objective:To compare the clinical efficacy and pharmacoeconomic evaluation of bevacizumab or cetuximab combined with chemotherapy in the treatment of advanced colorectal cancer.Methods:The clinical data of 68 patients with advanced colorectal cancer from January 2018 to December 2020 in Baotou Tumor Hospital were retrospectively analyzed. Among them, 40 patients with treated with bevacizumab combined with chemotherapy (bevacizumab group), 28 patients were treated with cetuximab combined with chemotherapy (cetuximab group), and the chemotherapy of two group was FOLFOX/FOLFIRI program. The short-term clinical efficacy, adverse reactions and pharmacoeconomic evaluation result were compared between two groups.Results:There were no statistical differences in effective rate and disease control rate between bevacizumab group and cetuximab group: 30.00% (12/40) vs. 28.57% (8/28) and 67.5% (27/40) vs. 60.71% (17/28), P>0.05. The incidence of Ⅲ to Ⅳ grade erythra in bevacizumab group was significantly lower than that in cetuximab group: 2.50% (1/40) vs. 71.43% (20/28), and there was statistical difference ( P<0.01); there were no statistical differences in the incidences of Ⅲ to Ⅳ grade bone marrow suppression, nausea vomiting, hepatic functional lesion and diarrhea between two groups ( P>0.05). The pharmacoeconomic evaluation result showed that the cost of monoclonal antibody and total cost in bevacizumab group were significantly lower than those in cetuximab group: (9 009 ± 1 500) yuan vs. (27 840 ± 2 202) yuan and (11 242 ± 1 731) yuan vs. (29 867 ± 3 002) yuan, and there were statistical differences ( P<0.01); the cost-effectiveness ratio in bevacizumab group was 37 473.3, and it in cetuximab group was 104 430.1, the incremental cost-effectiveness ratio of two programs was 11 640.6. Conclusions:In the treatment of advanced colorectal cancer, the efficacy of bevacizumab combined with chemotherapy is similar to that of cetuximab combined with chemotherapy, but bevacizumab combined with chemotherapy has lower costs and fewer adverse reactions, so bevacizumab is more economical and applicable.

Mechanism of ciRS-7 silencing to reverse cisplatin resistance in gastric cancer cells by targeting miR-944 / 中国药理学通报

Aim To investigate the effect of silencing circRNA ciRS-7 on cisplatin resistance of gastric cancer cells and its related mechanism. Methods Cisplatin-resistant gastric cancer cells HGC-27/DDP were constructed, ciRS-7 was silenced and HGC-27/DDP cells were treated with cisplatin. Real-time quantitative PCR (qRT-PCR) was used to detect the expression levels of ciRS-7 and microrNA-944 (miR-944). Cell counting kit-8 (CCK-8) was used to detect cell survival rate. Clone formation assay was used to detect the number of clones. Western blot was used to detect the protein expression levels of CyclinD1, proliferating cell nuclear antigen (PCNA), B-lymphocytoma-2-associated X protein (Bax), B-lymphocytoma-2 (Bcl-2) and cleaved aspartate-specific cysteine proteinase 3 (cleaved-caspase-3). Flow cytometry was used to detect cell apoptosis rate. In situ end labeling (TUNEL) was used to detect apoptosis index. Dual luciferase assay was used to verify the targeting relationship between ciRS-7 and miR-944. Results CiRS-7 was highly expressed in cisplatin-resistant gastric cancer tissues and cells, while miR-944 was low expressed in cisplatin-resistant gastric cancer tissues and cells. The survival rate, clonal formation number, protein expression levels of CyclinD1, PCNA and Bcl-2 in cisplatin-treated HGC-27/DDP cells were significantly reduced by silencing ciRS-7, while ciRS-7 expression level, apoptosis rate, apoptosis index, Bax, cleaved-caspase-3 protein expression levels of cisplatin-treated HGC-27/DDP cells were significantly raised. CiRS-7 targetedly inhibited the expression of miR-944. Conclusions Silencing ciRS-7 can reverse cisplatin resistance of gastric cancer cells, and the mechanism may be related to the promotion of miR-944 expression by silencing ciRS-7.

Single-shot AAV-vectored vaccine against SARS-CoV-2 with fast and long-lasting immunity

Due to the insufficient long-term protection and significant efficacy reduction to new variants of current COVID-19 vaccines, the epidemic prevention and control are still challenging. Here, we employ a capsid and antigen structure engineering (CASE) strategy to manufacture an adeno-associated viral serotype 6-based vaccine (S663V-RBD), which expresses trimeric receptor binding domain (RBD) of spike protein fused with a biological adjuvant RS09. Impressively, the engineered S663V-RBD could rapidly induce a satisfactory RBD-specific IgG titer within 2 weeks and maintain the titer for more than 4 months. Compared to the licensed BBIBP-CorV (Sinopharm, China), a single-dose S663V-RBD induced more endurable and robust immune responses in mice and elicited superior neutralizing antibodies against three typical SARS-CoV-2 pseudoviruses including wild type, C.37 (Lambda) and B.1.617.2 (Delta). More interestingly, the intramuscular injection of S663V-RBD could overcome pre-existing immunity against the capsid. Given its effectiveness, the CASE-based S663V-RBD may provide a new solution for the current and next pandemic.

Activin A overexpression promotes rat follicular development through SCF-kit-mediated cell signals.

We explored the possible signaling pathway by which activin A induces oocyte maturation. Inhba-overexpressing lentivirus vectors were constructed and transfected into primary granulosa cells in vitro and ovary tissues in vivo in rats. The granulosa cell growth curve was drawn, and antibodies for phospho-Smad2, phospho-Erk5, phospho-Nur77, and stem cell factor (SCF) were prepared for western blot analysis. Protein expression of SCF and C-kit in the rat ovaries was detected by immunohistochemical staining. The rate of granulosa cell proliferation was higher in the Inhba gene overexpression group (INH) than in the control groups (CON group and GFP group) in vitro. Protein expression of SCF and C-kit was higher in the INH group than in the other two groups. phospho-Smad2, phospho-ERK5, P-nur77, and SCF proteins showed positive expression in rat ovarian granulosa cells in each group and were obviously increased in the INH group. Activin A overexpression may promote rat granulosa cell proliferation through Smad2/ERK5/nur77 signaling pathways, and rat granulosa cells overexpressing activin A in vitro showed increased levels of SCF and c-kit proteins.

Application of cyberknife in treatment of retroperitoneal lymph node metastatic tumors in 81 cases / 中华放射肿瘤学杂志

Objective@#To evaluate the clinical efficacy and side effects of cyberknife therapy in the treatment of retroperitoneal lymph node metastatic tumor.@*Methods@#Among the 81 patients presenting with postoperative retroperitoneal lymph node metastases, 33 cases suffered from abdominal pain or low back pain, 7 had unilateral hydronephrosis and ureteral dilatation, and 8 developed unilateral or bilateral lower limb swelling. Using stereotactic radiotherapy with a cyberknife, DT was delivered at 33-45 Gy/3-6 F. The improvement of symptoms, objective tumor response rate, and irradiation-induced side effects were observed.@*Results@#At 4 weeks after treatment, pain and swelling of the lower extremities were completely mitigated, and hydronephrosis was fully healed in all patients. Enhanced CT or MRI was performed every 2 to 3 months. After 6-month follow-up, the complete response (CR) rate was calculated as 77%(62/81), 21%(17/81) for the partial response (PR) rate, 3%(2/81) for the stable disease (SD) rate and the effective rate (CR+ PR) was 98%.No case progressed. The main side effect was intestinal injury, including grade 1 in 16%(13/81), grade 2 in 9%(7/81). Multivariate analysis demonstrated the CR rate was not correlated with the type of the primary tumor or the size of the tumor (P>0.05), but was significantly associated with the distance of>0.5 cm between the tumor and intestine and the BED (α/β=10)>70 Gy of the tumor irradiation (P<0.01). The incidence of irradiation-related side effects was correlated with the maximum diameter of tumor>5 cm and the distance of<0.5 cm between the tumor and intestine (P<0.01).@*Conclusion@#Cyberknife is an efficacious and safe approach in the treatment of retroperitoneal lymph node metastatic tumors.

Comparative analysis of the incidence and predictors of chronic subdural hematoma after surgical clipping between unruptured intracranial aneurysms and ruptured intracranial aneurysms / 中国医师杂志

Objective To analyze the incidence and predictors of chronic subdural hematoma (CSDH) after surgical clipping between unruptured intracranial aneurysms (UIAs) and ruptured intracranial aneurysms (RIAs).Methods A retrospective cohort study was adopted to collect 486 cases of aneurysm patients (102 cases of UIAs patients and 384 RIAs patients) closed by aneurysm surgery who were admitted to the department of neurosurgery of The Fifth People's Hospital of Chengdu from October 2009 to December 2017.The clinical data,preoperative and postoperative imaging data and postoperative follow-up results were collected.The incidence of CSDH after operation in UIAs patients and RIAs patients was compared.The risk factors of CSDH after UIAs and RIAs patients were analyzed by multivariate Logistic regression model.Results The incidence of CSDH in UIAs and RIAs patients (10.78％ vs 3.13％,x2 =10.487,P =0.001) and the reoperation rate after CSDH (3.92％ vs 0.78％;x2 =5.599,P =0.018) were all statistically different,all of which showed that the patients with UIAs were higher than those of the patients with RIAs.Brain atrophy of grade 3-4 (OR =1.978,95％ CI:1.939-2.030,P ＜ 0.001),subdural effusion CT value ≥40 (OR =3.394,95％ CI:2.908-3.867,P ＜ 0.001) and subdural effusion (OR =2.872,95％ CI:2.648-3.019,P ＜0.001 grade) of Ⅰ B are independent risk factors of CSDH in patients with UIAs after aneurysm clipping (P ＜0.05).Subdural effusion CT value ≥ 40 (OR =3.442,95％ CI:2.918-3.8769,P ＜ 0.001) and grade Ⅰ B subdural effusion (OR =2.329,95％ CI:2.011-2.564,P ＜ 0.001) are independent risk factors for CSDH in patients with RIAs after aneurysm clipping (P ＜0.05).Conclusions The incidence of CSDH after aneurysm clipping in UIAs patients was significantly higher than that of RIAs patients.The risk factors for CSDH in the two groups were not the same.

Efficacy of high-intensity focused ultrasound ablation combined with gestrinone for treating adenomyosis / 重庆医学

Objective To investigate the efficacy of high-intensity focused ultrasound(HIFU) ablation combined with gestrinone for treating adenomyosis.Methods Sixty patients with adenomyosis were divided into the HIFU group(n=30) and HIFU combined medication group(n =30).The change of dysmenorrheal,menstrual volume,volumes of uterus and lesion in the two group were compared between before and after treatment.Results The menstrual volume and dysmenorrhea scores at 6,12 months after treatment showed a statistical difference between the two groups(P＜0.05);the volumes of and adenomyotic lesion and uterus after treatment in the combined medication group were lower than those in the HIFU group,but the difference was not statistically significant(P＞0.05).The hemoglobin level after 6-month treatment had statistical difference between the combined medication group and HIFU group(P＜0.05).During the follow-up period,there were 5 cases of pregnancy without recurrence cases in the combine medication group,8 cases had slight abnormality in the liver function transaminase and recovered to normal in re-examination at 1 month after drug withdrawal;1 case had pregnancy and 4 cases had recurrence in the HIFU group.Conclusion HIFU ablation combined with gestrinone can effectively alleviate the clinical symptoms in the patients with adenomyosis.

The complete mitochondrial genome of Grumgzimailo's toad-headed agama, Phrynocephalus grumgrizimailoi (Reptilia, Squamata, Agamidae).

Polymerase chain reaction (PCR), long-and-accurate PCR and directly sequencing by primer walking was used to sequenced he complete mitochondrial genome sequence of Grumgzimailo's toad- headed agama, Phrynocephalus grumgrizimailoi. The Genbank accession was KM093859. There was 16,301 bp in length of the entire mitochondrial genome of P. grumgrizimailoi and the content of A, T, C, and G were 36.4%, 26.5%, 25.0% and 12.1%, respectively, that was similar to most vertebrate. The complete mitochondrial genome of P. grumgrizimailoi contain 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes, plus 2 control regions and was similar to those of other Phrynocephalus sand lizards in gene arrangement and composition, except P. przewalskii and P. versicolor. The complete mitochondrial genome of P. grumgrizimailoi provided fundamental data for resolving phylogenetic relationship problems related to Agaimidae and genus Phrynocephalus.

Pregnancy outcome after high intensity focused ultrasound (HIFU) treatment for patients with adenomyosis / 重庆医学

Objective To investigate the reproductive outcome after high intensity focused ultrasound (HIFU ) ablation for pa-tients with adenomyosis who have fertility requirements .Methods From October 2010 to June 2013 ,38 patients with adenomyosis who have fertility requirements treated by HIFU in our hospital were analyzed .Results In 38 cases ,the menorrhagia severity score ,the dysmenorrhea pain score and adenomyosis lesions after HIFU ablation were significantly different from the levels before HIFU treatment(P<0 .05) .Successful pregnancy occurred in 15 patients ,full-term birth was seen in 7 patients(three of Cesarean section ,four of vaginal delivery);the remaining patients accept routine prenatal examination .All pregnant patients showed no ab-normality .Conclusion Although large sample size study with long-term follow up is need ,based on our results ,HIFU ablation for patients with adenomyosis might improve the pregnancy rate safely .

Changes in the reproductive endocrine function in rat following intraovary microinjection of inhba overexpression lentivirus vectors.

OBJECTIVE: To investigate the reproductive endocrine changes after inhba overexpression into rat ovary. METHOD: Thirty Sprague-Dawley rats were randomly assigned to three groups. Inhba overexpression lentivirus vectors (LV-eGFP-inhba) were microinjected into rat ovary (INH group); Control animals received the same amount lentivirus vector empty (CON group) or LV-eGFP (GFP group). Antral follicle amount and diameter were counted and serum level of activin A, E2, P, FSH, and LH and the expression of ER-α, ER-ß, PR, FSHR, and LHR were measured. RESULTS: There was no significant difference among three groups in antral follicle amount; antral follicle diameter was increased in INH group rats compared with the other group rats. Serum levels of activin A, E2, P, and FSH were increased and LH was decreased in INH group rats compared with the other group rats. The mRNA and protein expression of ER-α, ER-ß, FSHR was higher in INH group rats than that in the other group rats. There was no significant difference in mRNA and protein expression of PR among the three group rats, LHR expression was decreased in INH group rats compared with the other two group rats. CONCLUSION: inhba overexpression in rat ovary in vivo may change reproductive endocrine function.

A Retrospective Study:High Intensity Focused Ultrasound (HIFU) Associated A Microbubble-enhancing Contrast (“SonoVue”) Therapy for Uterine Fibroids with Pelvic Operation History / 昆明医科大学学报

Objective To evaluate the feasibility of high intensity focused ultrasound (HIFU) associated microbubble-enhancing contrast (“SonoVue”) therapeutic ablation of uterine fibroids in patients with pelvic operation history. Methods A total of 100 patients with uterine fibroid from October 2010 to December 2012 in Suining Central Hospital of Sichuan were treated with HIFU associated SonoVue. Among them, 50 patients had pelvic operation history. The no-perfused volume, median fractional ablation, ablation power rate, ablation dose, treatment time and adverse reaction were recorded. Results There was no statistical significance between patients with operation history and without in no-perfused volume and ablation does. The ablation power was lower in patients with operation than without. The treatment time and room time were longer in patients with operation than without. The adverse reactions were higher in patients with operation than without, especially skin pain and lower abdominal pain ( <0.05) . According to short- and long-term follow-up results, no major complications were observed in any patients. Conclusion Pelvic operation history increases the difficulty in treatment of HIFU ablation uterine fibroids. HIFU associated contrast agent SonoVue ablation can be considered a safe and feasible approach to the management of uterine fibroids in patients with pelvic operation history after grasping the indications and contraindications.

Genes encoding Pir51, Beclin 1, RbAp48 and aldolase b are up or down-regulated in human primary hepatocellular carcinoma.

AIM: To reveal new tumor markers and target genes from differentially expressed genes of primary tumor samples using cDNA microarray. METHODS: The (33 )P labeled cDNAs were synthesized by reverse transcription of message RNA from the liver cancerous tissue and adjacent non-cancerous liver tissue from the same patient and used to hybridize to LifeGrid 1.0 cDNA microarray blot containing 8400 known and unique human cDNA gene targets, and an expression profile of genes was produced in one paired human liver tumor tissue. After a global analysis of gene expression of 8400 genes, we selected some genes to confirm the differential expression using Northern blot and RT-PCR. RESULTS: Parallel analysis of the hybridized signals enabled us to get an expression profile of genes in which about 500 genes were differentially expressed in the paired liver tumor tissues. We identified 4 genes, the expression of three (Beclin 1, RbAp48 and Pir51) were increased and one (aldolase b) was decreased in liver tumor tissues. In addition, the expression of these genes in 6 hepatoma cell lines was also showed by RT-PCR analysis. CONCLUSION: cDNA microarray permits a high throughput identification of changes in gene expression. The genes encoding Beclin 1, RbAp48, Pir51 and aldolase b are first reported that may be related with hepatocarcinoma.

Cloning and Characterization of fup1, A Gene Highly Expressed in Hepatocellular Carcinoma.

Primary hepatocellular carcinoma(HCC) is one of the common malignant tumors in China. In our previous work, a gene named fup1(function-unknown protein 1) was isolated that was expressed differently in HCC and in normal liver. We assumed that it might be a candidate oncogene for the HCC. The fup1 gene had a ORF of 1 233 bp, encoding a protein with M(r) of 46 kD and isoelectric point of 5.48. The sequence characteristics showed its possible localization in nuclei. Northern blots showed that this gene was weakly expressed in many types of human tissues, except in the heart, implying its tissue-specific expression pattern. MTT assay of the NIH 3T3 cells transfected with this gene in the form of recombinant eukaryotic expression plasmid showed its enhancing role to cellular proliferation.

Detection of Lis1 Gene Frame Shift Mutation in Human Hepatocarcinoma.

GST fusion protein expression system combined with protein truncation test(PTT) protocol was used to detect gene frame shift mutation. The RT-PCR products of Lis1 genes from hepatocarcinoma samples were respectively cloned into a GST fusion protein expression vector pGEX-1, then expressed in E.coli. The results showed a truncated 33 kD fusion protein in SDS-PAGE, although the full-translated product of Lis1 gene should be of 71 kD. Sequencing revealed insertion of an A residue, causing the premature termination of translation, between the 163th and 164th nucleotide of Lis1 gene. This improved PTT assay was proved to be a fast and effective way in detecting gene frame shift mutation.