CRISPR/Cas: Advances, Limitations, and Applications for Precision Cancer Research.

Cancer is one of the most leading causes of mortalities worldwide. It is caused by the accumulation of genetic and epigenetic alterations in 2 types of genes: tumor suppressor genes (TSGs) and proto-oncogenes. In recent years, development of the clustered regularly interspaced short palindromic repeats (CRISPR) technology has revolutionized genome engineering for different cancer research ranging for research ranging from fundamental science to translational medicine and precise cancer treatment. The CRISPR/CRISPR associated proteins (CRISPR/Cas) are prokaryote-derived genome editing systems that have enabled researchers to detect, image, manipulate and annotate specific DNA and RNA sequences in various types of living cells. The CRISPR/Cas systems have significant contributions to discovery of proto-oncogenes and TSGs, tumor cell epigenome normalization, targeted delivery, identification of drug resistance mechanisms, development of high-throughput genetic screening, tumor models establishment, and cancer immunotherapy and gene therapy in clinics. Robust technical improvements in CRISPR/Cas systems have shown a considerable degree of efficacy, specificity, and flexibility to target the specific locus in the genome for the desired applications. Recent developments in CRISPRs technology offers a significant hope of medical cure against cancer and other deadly diseases. Despite significant improvements in this field, several technical challenges need to be addressed, such as off-target activity, insufficient indel or low homology-directed repair (HDR) efficiency, in vivo delivery of the Cas system components, and immune responses. This study aims to overview the recent technological advancements, preclinical and perspectives on clinical applications of CRISPR along with their advantages and limitations. Moreover, the potential applications of CRISPR/Cas in precise cancer tumor research, genetic, and other precise cancer treatments discussed.

Golgi phosphoprotein-3 promotes invasiveness of gastric cancer cells through the mTOR signalling pathway.

PURPOSE: Golgi phosphoprotein-3 (GOLPH3) is an oncogene that is overexpressed in multiple cancers and is associated with poor prognosis. The aim of this study was to examine the impact of GOLPH3 on the migration and metastasis of gastric cancer cells. METHODS: Following the shRNA-mediated knockdown of GOLPH3, we analyzed cytoskeletal reorganization and cell invasion, migration and adhesion, and determined the impact of components of the mammalian target of the rapamycin (mTOR) signalling pathway. RESULTS: The GOLPH3 mRNA and protein expression were significantly lower in both SGC-7901 and MKN-28 cells as compared with poorly-differentiated BGC-823 cells. The GOLPH3 knockdown also significantly reduced cell invasion in all three cell lines through reduced migration as compared with the non-targeting control sequence group. The GOLPH3 knockdown also reduced F-actin in all three cell lines, and decreased cell adhesion in BGC-823 and SGC-7901 cells. Finally, p-mTOR, p70S6K, p-4EBP1 and RhoA protein levels were significantly downregulated in shGOLPH3-1-treated cells. CONCLUSIONS: In conclusion, GOLPH3 increased in poorly-differentiated gastric cancer cells, activating the mTOR-70S6K/4EBP1-RhoA signalling pathway to promote the migration and metastasis of gastric cancer cells.

Amyloid ß precursor protein silencing attenuates epithelial­mesenchymal transition of nasopharyngeal carcinoma cells via inhibition of the MAPK pathway.

Advances in the treatment of nasopharyngeal carcinoma (NPC) have significantly improved the local control rate; however, distant metastasis remains a principal cause of mortality. Previous studies have demonstrated that the expression levels of amyloid ß precursor protein (APP) are increased in NPC. The present study aimed to investigate the association between APP and the development of NPC. In order to knockdown APP expression, an APP­small interfering RNA vector was synthesized and transfected into SUNE­1 cells. Cell Counting Kit­8 assay was performed to assess cell viability. The migratory and invasive abilities of SUNE­1 cells were examined by wound healing and Transwell assays, respectively. Reverse transcription­quantitative polymerase chain reaction and western blotting were performed to measure the mRNA and protein expression levels of APP, and additional factors involved in epithelial­mesenchymal transition (EMT) and in the mitogen­activated protein kinase (MAPK) signaling pathway. APP silencing significantly suppressed cell viability, migration and invasion. In addition, APP interference downregulated the expression levels of metastasis­associated 1, matrix metalloproteinase (MMP)­2 and MMP­9; however, knockdown of APP led to upregulation of tissue inhibitor of metalloproteinases 2 and inhibited EMT. The phosphorylation levels of p38, extracellular signal­-regulated kinases 1/2 and c­Jun N­terminal kinases 1/2 were decreased following downregulation of APP. The present results suggested that APP knockdown may significantly inhibit the development of NPC by suppressing cell viability, migration and invasion, and by inhibiting the EMT process via downregulation of the MAPK signaling pathway. Therefore, APP may facilitate the development of a novel gene therapy for the treatment of NPC.

Knockdown of serpin peptidase inhibitor clade C member 1 inhibits the growth of nasopharyngeal carcinoma cells.

Nasopharyngeal carcinoma (NPC) is a type of cancer originating in the nasopharynx. There are no NPC­specific treatments available at present. Serpin peptidase inhibitor clade C member 1 (SERPINC1) serves roles in anticoagulation and anti­inflammation. The aim of the present study was to investigate the role of SERPINC1 in the proliferation and apoptosis of NPC cells. Tumor and adjacent healthy tissue samples were collected from patients with NPC. Additionally, the SERPINC1 gene was silenced in the HNE3 cell line using short interfering RNA targeted against SERPINC1 (SERPINC1­siRNA). Cell viability was determined via a Cell Counting Kit­8 assay; furthermore, proliferation and apoptosis were investigated via flow cytometry. Western blotting and reverse transcription­quantitative polymerase chain reaction analysis were performed to determine the expression levels of protein and mRNA. It was revealed that the expression levels of SERPINC1 mRNA and protein were increased in NPC tumor tissues compared with in adjacent healthy tissues. The expression of SERPINC1 mRNA and protein in HNE3 cells decreased following SERPINC1­siRNA transfection. Furthermore, knockdown of SERPINC1 promoted apoptosis and inhibited proliferation. It was also demonstrated that silencing SERPINC1 upregulated the expression of B­cell lymphoma-2 (Bcl­2)­associated X protein and p53 mRNA and protein, and downregulated that of Bcl­2, survivin and cyclin D1. Downregulation of SERPINC1 reduced the phosphorylation of phosphatidylinositol 3­kinase (PI3K), protein kinase B (Akt) and mammalian target of rapamycin (mTOR). Thus, SERPINC1 knockdown may promote the apoptosis of HNE3 cells and inhibit proliferation via the suppression of the PI3K/Akt/mTOR signaling pathway.

Male breast cancer and mantle cell lymphoma in a single patient: A case report and literature review.

RATIONALE: Although still relatively rare, multiple primary malignant neoplasms (MPMNs) have been increasingly reported in recent years. PATIENT CONCERNS AND DIAGNOSES: A 65-year-old man was referred to our hospital for a painless, incidental left axillary lump. Ultrasound showed enlarged left axillary lymph nodes. An excisional biopsy was conducted on 3 lymph nodes. The pathological diagnosis was determined to be metastatic adenocarcinoma and mantle cell lymphoma (MCL) in the lymph nodes. Further physical examination of the patient yielded a 1.5-cm hard, left subareolar mass. INTERVENTIONS AND OUTCOMES: The patient underwent modified radical mastectomy. The diagnosis was grade II invasive ductal carcinoma (stage IIA). The axillary lymph node showed MCL (stage I, group A), but not metastatic ductal carcinoma. The patient received chemotherapy, including 6 courses of CHOP (A chemotherapy protocol consists of cyclophosphamide 1.2 g day 1, doxorubicin 80 mg day 1, vindesine 4 mg day1, and prednisone 90 mg from day 1 to 5) for lymphoma and breast cancer. The patient was also administered endocrine therapy. After a 54-month follow-up, the patient was well with no evidence of disease. LESSONS: MPMNs are easily misdiagnosed as a primary and metastatic tumor, leading to delayed or erroneous treatment. Male breast cancer in a patient with MCL is rare. Early diagnosis and proper therapy are necessary for an optimal prognosis. Further studies are required to define the mechanisms and risk factors of MPMNs.