Traditional development of small protein scaffolds has relied on display technologies and mutation-based engineering, which limit sequence and functional diversity, thereby constraining their therapeutic and application potential. Protein design tools have significantly advanced the creation of novel protein sequences, structures, and functions. However, further improvements in design strategies are still needed to more efficiently optimize the functional performance of protein-based drugs and enhance their druggability. Here, we extended an evolution-based design protocol to create a novel minibinder, BindHer, against the human epidermal growth factor receptor 2 (HER2). It not only exhibits super stability and binding selectivity but also demonstrates remarkable properties in tissue specificity. Radiolabeling experiments with ^99mTc, ⁶⁸Ga, and ¹⁸F revealed that BindHer efficiently targets tumors in HER2-positive breast cancer mouse models, with minimal nonspecific liver absorption, outperforming scaffolds designed through traditional engineering. These findings highlight a new rational approach to automated protein design, offering significant potential for large-scale applications in therapeutic mini-protein development.

Objective:To evaluate the clinical efficacy of individualized thrombolysis-assisted comprehensive intervention for deep vein thrombosis (DVT) in the lower limbs.Methods:This study included 32 patients with acute lower limb DVT diagnosed by angiography who received treatment at the Jianhu Clinical Medical College of Yangzhou University from March 2012 to November 2021. These patients first received implantation of an inferior vena cava filter. Then they were divided into a control group and an observation group based on treatment methods. The control group received thrombolytic catheterization and a routine infusion of urokinase. In the observation group, balloon dilation was performed first, and a large lumen catheter was used to draw blood clots. Subsequently, urokinase at a dose based on fibrinogen measurement was injected through a thrombolytic catheter. Swelling reduction, venous patency, and complications of the affected limbs were monitored.Results:In the control group, the difference in thigh circumference before treatment was (4.65 ± 1.06) cm, and after treatment, it was (2.76 ± 1.25) cm. In the observation group, the difference in thigh circumference before treatment was (4.73 ± 1.03) cm, and it was (1.40 ± 0.83) cm after treatment. In the control group, the difference in calf circumference before treatment was (2.24 ± 0.90) cm, and it was (1.56 ± 0.86) cm after treatment. In the observation group, the difference in calf circumference before treatment was (2.40 ± 0.83) cm, and it was (0.80 ± 0.73) cm after treatment. After treatment, the differences in thigh circumference and calf circumference between the healthy and affected sides were statistically significant ( t = 3.58, 2.67, both P < 0.05). After treatment, there was a significant difference in venous patency between the control and observation groups (34.02% [33/97] vs. 68.18% [60/88], t = 3.44, P < 0.05). After 12 months of follow-up, the Villalta scale score, which was used to evaluate post-thrombotic syndrome, was (9.23 ± 4.07) points in the control group, which was significantly different from (5.73 ± 3.39) points in the observation group ( t = 2.62, P < 0.05). Conclusion:Individualized thrombolysis-assisted comprehensive intervention is highly effective in the treatment of DVT in the lower limbs and results in few complications.

Objective To investigate AXIN2 mRNA expression level in hepatocellular carcinoma (HCC) , and to analyze the effect of AXIN2 gene methylation status on its mRNA expression and HCC genesis and development. Methods Fifty-three surgical excised HCC specimens and paired adjacent non-cancerous specimens, seven normal liver specimens and five HCC cell lines were collected. The expression of AXIN2 at mRNA level and the methylation status of AXIN2 gene promoter were determined by quantitative PCR. Results The expression of AXIN2 mRNA was lower in HCC tissues (0.1629 + 0.0679) than that in adjacent non-cancerous tissues (0. 4155 + 0. 2330), and there was significant difference (Z= -2. 567, P = 0. 010). The methylation level of AXIN2 gene in HCC and adjacent non-cancerous tissues (39. 77% ±3. 89%, and 36. 92% ±2. 81%) was significantly higher than that in normal liver tissues (7. 38% ±2. 40% , t=-3. 663 ,P = 0. 009;t= -4. 591 ,P = 0. 007).AXIN2 gene was hypermethylated in all five HCC cell lines. There was a negative correlation between AXIN2 mRNA expression level and the degree of methylation ( r = -0. 458, P = 0. 032). The methylation level was higher in TNM Ⅲ patients of HCC than that in TNM Ⅰ and Ⅱ patients (P =0.008). Conclusion The down-regulation of AXIN2 gene mRNA expression is correlated with its hypermethylation status. The low expression of AXIN2 mRNA and the abnormal methylation of promoter may be one of the important mechanism of HCC genesis and development.