Human 8-cell embryos enable efficient induction of disease-preventive mutations without off-target effect by cytosine base editor

Approximately 140 million people worldwide are homozygous carriers of APOE4 (ε4), a strong genetic risk factor for late onset familial and sporadic Alzheimer's disease (AD), 91% of whom will develop AD at earlier age than heterozygous carriers and noncarriers. Susceptibility to AD could be reduced by targeted editing of APOE4, but a technical basis for controlling the off-target effects of base editors is necessary to develop low-risk personalized gene therapies. Here, we first screened eight cytosine base editor variants at four injection stages (from 1- to 8-cell stage), and found that FNLS-YE1 variant in 8-cell embryos achieved the comparable base conversion rate (up to 100%) with the lowest bystander effects. In particular, 80% of AD-susceptible ε4 allele copies were converted to the AD-neutral ε3 allele in human ε4-carrying embryos. Stringent control measures combined with targeted deep sequencing, whole genome sequencing, and RNA sequencing showed no DNA or RNA off-target events in FNLS-YE1-treated human embryos or their derived stem cells. Furthermore, base editing with FNLS-YE1 showed no effects on embryo development to the blastocyst stage. Finally, we also demonstrated FNLS-YE1 could introduce known protective variants in human embryos to potentially reduce human susceptivity to systemic lupus erythematosus and familial hypercholesterolemia. Our study therefore suggests that base editing with FNLS-YE1 can efficiently and safely introduce known preventive variants in 8-cell human embryos, a potential approach for reducing human susceptibility to AD or other genetic diseases.

Expression of CK19 in sentinel lymph node for breast cancer and its clinical significance / 中国综合临床

Objective To detect the expression of CK19 in sentinel lymph node for breast cancer and to evaluate its clinical significance.Methods SLNs of 30 patients with breast cancer were successfully detected with methylene blue,both SLN and ALN were examined by hematoxylin and eosin staining ( HK staining). The expression level of CK19 in SLNs were assessed by RT-PCR and Western Blot for mRNA and Protein respectively. Results Detection positive rate of SLN melastases was 26. 67% (8/30) by HE staining,and 1 case of false negative. Through RT-PCR,12 cases,including all the positive cases detected by routine pathological examination,were detected to be positive in the expression of CK19mRNA, with a positive rate of 40. 0% ( 12/30) , and a positive rate of 36. 67% (11/30) for CK19 was observed by Western Blot There was a significant difference between RT-PCR or Western Blot methods and routine pathological examination ( P < 0.05 ) for the detection efficiency of SI.Ns melastases. Conclusions CK19rnRNA may be a potential tumor marker for detecting micrometastasis in SLNs of breast cancer. The combined utilization of locating SLNs and detecting CK19 mRNA expression by RT-PCR instead of histopathological examination can greatly enhance the detection efficiency of SLN micrometastasis of breast cancer.