Predicting Ki-67 expression levels in breast cancer using radiomics-based approaches on digital breast tomosynthesis and ultrasound.

Purpose: To construct and validate radiomics models that utilize ultrasound (US) and digital breast tomosynthesis (DBT) images independently and in combination to non-invasively predict the Ki-67 status in breast cancer. Materials and methods: 149 breast cancer women who underwent DBT and US scans were retrospectively enrolled from June 2018 to August 2023 in total. Radiomics features were acquired from both the DBT and US images, then selected and reduced in dimensionality using several screening approaches. Establish radiomics models based on DBT, and US separately and combined. The area under the receiver operating characteristic curve (AUC), accuracy, specificity, and sensitivity were utilized to validate the predictive ability of the models. The decision curve analysis (DCA) was used to evaluate the clinical applicability of the models. The output of the classifier with the best AUC performance was converted into Rad-score and was regarded as Rad-Score model. A nomogram was constructed using the logistic regression method, integrating the Rad-Score and clinical factors. The model's stability was assessed through AUC, calibration curves, and DCA. Results: Support vector machine (SVM), logistic regression (LR), and random forest (RF) were trained to establish radiomics models with the selected features, with SVM showing optimal results. The AUC values for three models (US_SVM, DBT_SVM, and merge_SVM) were 0.668, 0.704, and 0.800 respectively. The DeLong test indicated a notable disparity in the area under the curve (AUC) between merge_SVM and US_SVM (p = 0.048), while there was no substantial variability between merge_SVM and DBT_SVM (p = 0.149). The DCA curve indicates that merge_SVM is superior to unimodal models in predicting high Ki-67 level, showing more clinical values. The nomogram integrating Rad-Score with tumor size obtained the better performance in test set (AUC: 0.818) and had more clinical net. Conclusion: The fusion radiomics model performed better in predicting the Ki-67 expression level of breast carcinoma, but the gain effect is limited; thus, DBT is preferred as a preoperative diagnosis mode when resources are limited. Nomogram offers predictive advantages over other methods and can be a valuable tool for predicting Ki-67 levels in BC.

Deep learning-based dose map prediction for high-dose-rate brachytherapy.

Background. Creating a clinically acceptable plan in the time-sensitive clinic workflow of brachytherapy is challenging. Deep learning-based dose prediction techniques have been reported as promising solutions with high efficiency and accuracy. However, current dose prediction studies mainly target EBRT which are inappropriate for brachytherapy, the model designed specifically for brachytherapy has not yet well-established.Purpose. To predict dose distribution in brachytherapy using a novel Squeeze and Excitation Attention Net (SE_AN) model.Method. We hypothesized the tracks of192Ir inside applicators are essential for brachytherapy dose prediction. To emphasize the applicator contribution, a novel SE module was integrated into a Cascaded UNet to recalibrate informative features and suppress less useful ones. The Cascaded UNet consists of two stacked UNets, with the first designed to predict coarse dose distribution and the second added for fine-tuning 250 cases including all typical clinical applicators were studied, including vaginal, tandem and ovoid, multi-channel, and free needle applicators. The developed SE_AN was subsequently compared to the classic UNet and classic Cascaded UNet (without SE module) models. The model performance was evaluated by comparing the predicted dose against the clinically approved plans using mean absolute error (MAE) of DVH metrics, includingD2ccandD90%.Results. The MAEs of DVH metrics demonstrated that SE_AN accurately predicted the dose with 0.37 ± 0.25 difference for HRCTVD90%, 0.23 ± 0.14 difference for bladderD2cc, and 0.28 ± 0.20 difference for rectumD2cc. In comparison studies, UNet achieved 0.34 ± 0.24 for HRCTV, 0.25 ± 0.20 for bladder, 0.25 ± 0.21 for rectum, and Cascaded UNet achieved 0.42 ± 0.31 for HRCTV, 0.24 ± 0.19 for bladder, 0.23 ± 0.19 for rectum.Conclusion. We successfully developed a method specifically for 3D brachytherapy dose prediction. Our model demonstrated comparable performance to clinical plans generated by experienced dosimetrists. The developed technique is expected to improve the standardization and quality control of brachytherapy treatment planning.

Epidemiological survey of avian adenovirus in China from 2015 to 2021 and the genetic variability of highly pathogenic Fadv-4 isolates.

The prevalence of poultry adenovirus in China is determined using clinical diagnosis, molecular biological testing, serological testing, and LMH cell virus isolation. These methods can track and test key poultry and waterfowl breeding areas across the country. From 2015 to 2021, 9613 suspected adenovirus samples were collected from 28 provinces. After the first generation of gene sequencing, a total of 2210 hexo gene fragments were obtained. Among them, FAdV-1 type accounted for 7.65%, FAdV-2 type accounted for 5.34%, FAdV-3 type accounted for 2.04%, FAdV-4 type accounted for 38.24%, FAdV-5 type accounted for 2.17%, FAdV-6 type accounted for 0.32%, FAdV-7 type accounted for 0.77%, FAdV-8a type accounted for 10.63%, FAdV-8b type accounted for 11.58%, FAdV-9 type accounted for 0.50%, FAdV-10 type accounted for 8.10%, and FAdV-11 type accounted for 12.67%. A total of 877 FAdV strains were isolated from FAdV suspected samples by seeding LMH cells, and there were 475 FAdV-4 strains among them. A total of 473 isolates were highly pathogenic FAdV-4, and the percentage of amino acid homology with the highly pathogenic FAdV-4 reference strains was >99.1%. Two isolates were non-pathogenic, and the amino acid homology with the ON1 reference strain was >99.6%. Part of the amino acid positions of the hexon gene have mutations, including positions 188, 193, 195, 238, and 240.

Genome Editing with AAV-BR1-CRISPR in Postnatal Mouse Brain Endothelial Cells.

Brain endothelial cells (ECs) are an important component of the blood-brain barrier (BBB) and play key roles in restricting entrance of possible toxic components and pathogens into the brain. However, identifying endothelial genes that regulate BBB homeostasis remains a time-consuming process. Although somatic genome editing has emerged as a powerful tool for discovery of essential genes regulating tissue homeostasis, its application in brain ECs is yet to be demonstrated in vivo. Here, we used an adeno-associated virus targeting brain endothelium (AAV-BR1) combined with the CRISPR/Cas9 system (AAV-BR1-CRISPR) to specifically knock out genes of interest in brain ECs of adult mice. We first generated a mouse model expressing Cas9 in ECs (Tie2Cas9). We selected endothelial ß-catenin (Ctnnb1) gene, which is essential for maintaining adult BBB integrity, as the target gene. After intravenous injection of AAV-BR1-sgCtnnb1-tdTomato in 4-week-old Tie2Cas9 transgenic mice resulted in mutation of 36.1% of the Ctnnb1 alleles, thereby leading to a dramatic decrease in the level of CTNNB1 in brain ECs. Consequently, Ctnnb1 gene editing in brain ECs resulted in BBB breakdown. Taken together, these results demonstrate that the AAV-BR1-CRISPR system is a useful tool for rapid identification of endothelial genes that regulate BBB integrity in vivo.

Loss of Smad4 promotes aggressive lung cancer metastasis by de-repression of PAK3 via miRNA regulation.

SMAD4 is mutated in human lung cancer, but the underlying mechanism by which Smad4 loss-of-function (LOF) accelerates lung cancer metastasis is yet to be elucidated. Here, we generate a highly aggressive lung cancer mouse model bearing conditional KrasG12D, p53fl/fl LOF and Smad4fl/fl LOF mutations (SPK), showing a much higher incidence of tumor metastases than the KrasG12D, p53fl/fl (PK) mice. Molecularly, PAK3 is identified as a downstream effector of Smad4, mediating metastatic signal transduction via the PAK3-JNK-Jun pathway. Upregulation of PAK3 by Smad4 LOF in SPK mice is achieved by attenuating Smad4-dependent transcription of miR-495 and miR-543. These microRNAs (miRNAs) directly bind to the PAK3 3'UTR for blockade of PAK3 production, ultimately regulating lung cancer metastasis. An inverse correlation between Smad4 and PAK3 pathway components is observed in human lung cancer. Our study highlights the Smad4-PAK3 regulation as a point of potential therapy in metastatic lung cancer.

Brain endothelial PTEN/AKT/NEDD4-2/MFSD2A axis regulates blood-brain barrier permeability.

The low level of transcytosis is a unique feature of cerebrovascular endothelial cells (ECs), ensuring restrictive blood-brain barrier (BBB) permeability. Major facilitator superfamily domain-containing 2a (MFSD2A) is a key regulator of the BBB function by suppressing caveolae-mediated transcytosis. However, the mechanisms regulating MFSD2A at the BBB have been barely explored. Here, we show that cerebrovascular EC-specific deletion of Pten (phosphatase and tensin homolog) results in a dramatic increase in vesicular transcytosis by the reduction of MFSD2A, leading to increased transcellular permeability of the BBB. Mechanistically, AKT signaling inhibits E3 ubiquitin ligase NEDD4-2-mediated MFSD2A degradation. Consistently, cerebrovascular Nedd4-2 overexpression decreases MFSD2A levels, increases transcytosis, and impairs BBB permeability, recapitulating the phenotypes of Pten-deficient mice. Furthermore, Akt deletion decreases phosphorylated NEDD4-2 levels, restores MFSD2A levels, and normalizes BBB permeability in Pten-mutant mice. Altogether, our work reveals the essential physiological function of the PTEN/AKT/NEDD4-2/MFSD2A axis in the regulation of BBB permeability.

The diversity of adult lung epithelial stem cells and their niche in homeostasis and regeneration.

The adult lung, a workhorse for gas exchange, is continually subjected to a barrage of assaults from the inhaled particles and pathogens. Hence, homeostatic maintenance is of paramount importance. Epithelial stem cells interact with their particular niche in the adult lung to orchestrate both natural tissue rejuvenation and robust post-injury regeneration. Advances in single-cell sequencing, lineage tracing, and living tissue imaging have deepened our understanding about stem cell heterogeneities, transition states, and specific cell lineage markers. In this review, we provided an overview of the known stem/progenitor cells and their subpopulations in different regions of the adult lung, and explored the regulatory networks in stem cells and their respective niche which collectively coordinated stem cell quiescence and regeneration states. We finally discussed relationships between dysregulated stem cell function and lung disease.

Dysregulated Tgfbr2/ERK-Smad4/SOX2 Signaling Promotes Lung Squamous Cell Carcinoma Formation.

Lung squamous cell carcinoma (SCC) is a common type of lung cancer. There is limited information on the genes and pathways that initiate lung SCC. Here, we report that loss of TGFß type II receptor (Tgfbr2), frequently deleted in human lung cancer, led to predominant lung SCC development in KrasG12D mice with a short latency, high penetrance, and extensive metastases. Tgfbr2-loss-driven lung SCCs resembled the salient features of human lung SCC, including histopathology, inflammatory microenvironment, and biomarker expression. Surprisingly, loss of Smad4, a key mediator of Tgfbr2, failed to drive lung SCC; instead, low levels of phosphorylated ERK1/2, a Smad-independent downstream effector of Tgfbr2, were tightly associated with lung SCC in both mouse and human. Mechanistically, inhibition of phosphorylated ERK1/2 significantly upregulated the expression of SOX2, an oncogenic driver of lung SCC, and cooperated with SMAD4 repression to elevate SOX2. Inhibition of ERK1/2 in Smad4fl/fl ;KrasG12D mice led to extensive lung SCC formation that resembled the SCC phenotype of Tgfbr2-deficient mice. Overall, we reveal a key role of ERK1/2 in suppressing SCC formation and demonstrate that dysregulated Tgfbr2/ERK-Smad4/SOX2 signaling drives lung SCC formation. We also present a mouse model of metastatic lung SCC that may be valuable for screening therapeutic targets. SIGNIFICANCE: This study sheds new light on the mechanisms underlying lung SCC formation driven by mutated Kras.

TGF-ß/SMAD4-Regulated LncRNA-LINP1 Inhibits Epithelial-Mesenchymal Transition in Lung Cancer.

Lung cancer is the leading cause of cancer related deaths worldwide. TGF-ß-induced epithelial-mesenchymal transition (EMT) is a key cell-intrinsic identity for tumor cell migration, invasion, and stemness acquisition in cancer metastasis. Long noncoding RNAs (lncRNAs) have not been fully investigated for their involvement in regulating TGF-ß-induced EMT and metastasis in lung cancer. Here, we demonstrated that the transcription of lncRNA in nonhomologous end joining (NHEJ) pathway 1 (LINP1) was inhibited by TGF-ß1 in a SMAD4-dependent manner. LINP1 suppressed EMT of lung cancer cells, thereby controlling cancer cell migration, invasion, and stemless. Moreover, LINP1 inhibited TGF-ß-induced EMT and cell invasion in lung cancer cells. Our study reveals the role of LINP1 in the regulation of TGF-ß-induced EMT in human lung cancer.

Ezh2 Acts as a Tumor Suppressor in Kras-driven Lung Adenocarcinoma.

Previous studies have suggested that enhancer zeste homolog 2 (Ezh2), a histone methyltransferase subunit of polycomb repressive complex 2 (PRC2), acts as an oncogene in lung adenocarcinoma (ADC) development. However, we found that in human lung ADC samples, deletion and mutations of EZH2 were also frequently present, with 14% of patients harboring loss-of-function EZH2 alterations. To explore the effect of Ezh2 loss on lung tumor formation, lung epithelial Ezh2 gene was deleted in Kras-driven lung ADC mouse model. Unexpectedly, Ezh2 loss dramatically promoted Kras-driven ADC formation. KrasG12D/+;Ezh2fl/fl mice exhibited shorter lifespan, more tumor lesions and higher tumor burden than KrasG12D/+ mice, suggesting the tumor-suppressive role of Ezh2 in Kras-driven ADCs. Mechanistically, Ezh2 loss amplified Akt and ERK activation through de-repressing its target insulin-like growth factor 1 (Igf1). Additionally, Ezh2 loss cooperated with Kras mutation to exacerbate the inflammatory response, as shown by massive macrophage and neutrophil infiltrates, as well as a marked increase in tumor-associated cytokines such as IL-6 and TNF-α. Taken together, our findings revealed the tumor suppressive function of Ezh2 in Kras-driven ADCs, underlining the importance of revaluating the application of EZH2 inhibitors in a variety of cancers.

Loss of p53 attenuates the contribution of IL-6 deletion on suppressed tumor progression and extended survival in Kras-driven murine lung cancer.

Interleukin-6 (IL-6) is involved in lung cancer tumorigenesis, tumor progression, metastasis, and drug resistance. Previous studies show that blockade of IL-6 signaling can inhibit tumor growth and increase drug sensitivity in mouse models. Clinical trials in non-small cell lung cancer (NSCLC) reveal that IL-6 targeted therapy relieves NSCLC-related anemia and cachexia, although other clinical effects require further study. We crossed IL-6(-/-) mice with Kras(G12D) mutant mice, which develop lung tumors after activation of mutant Kras(G12D), to investigate whether IL-6 inhibition contributes to tumor progression and survival time in vivo. Kras(G12D); IL-6(-/-) mice exhibited increased tumorigenesis, but slower tumor growth and longer survival, than Kras(G12D) mice. Further, in order to investigate whether IL-6 deletion contributes to suppression of lung cancer metastasis, we generated Kras(G12D); p53(flox/flox); IL-6(-/-) mice, which developed lung cancer with a trend for reduced metastases and longer survival than Kras(G12D); p53(flox/flox) mice. Tumors from Kras(G12D); IL-6(-/-) mice showed increased expression of TNFα and decreased expression of CCL-19, CCL-20 and phosphorylated STAT3(pSTAT3) than Kras(G12D) mice; however, these changes were not present between tumors from Kras(G12D); p53(flox/flox); IL-6(-/-) and Kras(G12D); p53(flox/flox) mice. Upregulation of pSTAT3 and phosphorylated AKT(pAKT) were observed in Kras(G12D) tumors with p53 deletion. Taken together, these results indicate that IL-6 deletion accelerates tumorigenesis but delays tumor progression and prolongs survival time in a Kras-driven mouse model of lung cancer. However, these effects can be attenuated by p53 deletion.