Sucrose synthase (SuS), which catalyzes the reversible conversion of sucrose and uridine diphosphate (UDP) into fructose and UDP-glucose, is a key enzyme in sucrose metabolism in higher plants. SuS ...belongs to family 4 of the glycosyltransferases (GT4) and contains an E-X₇-E motif that is conserved in members of GT4 and two other GT families. To gain insight into the roles of this motif in rice sucrose synthase 3 (RSuS3), the two conserved glutamate residues (E678 and E686) in this motif and a phenylalanine residue (F680) that resides between the two glutamate residues were changed by site-directed mutagenesis. All mutant proteins maintained their tetrameric conformation. The mutants E686D and F680Y retained partial enzymatic activity and the mutants E678D, E678Q, F680S, and E686Q were inactive. Substrate binding assays indicated that UDP and fructose, respectively, were the leading substrates in the sucrose degradation and synthesis reactions of RSuS3. Mutations on E678, F680, and E686 affected the binding of fructose, but not of UDP. The results indicated that E678, F680, and E686 in the E-X₇-E motif of RSuS3 are essential for the activity of the enzyme and the sequential binding of substrates. The sequential binding of the substrates implied that the reaction catalyzed by RSuS can be controlled by the availability of fructose and UDP, depending on the metabolic status of a tissue.
Models that recapitulate the complexity of human tumors are urgently needed to develop more effective cancer therapies. We report a bank of human patient-derived xenografts (PDXs) and matched ...organoid cultures from tumors that represent the greatest unmet need: endocrine-resistant, treatment-refractory and metastatic breast cancers. We leverage matched PDXs and PDX-derived organoids (PDxO) for drug screening that is feasible and cost-effective with in vivo validation. Moreover, we demonstrate the feasibility of using these models for precision oncology in real time with clinical care in a case of triple-negative breast cancer (TNBC) with early metastatic recurrence. Our results uncovered a Food and Drug Administration (FDA)-approved drug with high efficacy against the models. Treatment with this therapy resulted in a complete response for the individual and a progression-free survival (PFS) period more than three times longer than their previous therapies. This work provides valuable methods and resources for functional precision medicine and drug development for human breast cancer.
Human electroencephalography (EEG) is a brain monitoring modality that senses cortical neuroelectrophysiological activity in high-temporal resolution. One of the greatest challenges posed in ...applications of EEG is the unstable signal quality susceptible to inevitable artifacts during recordings. To date, most existing techniques for EEG artifact removal and reconstruction are applicable to offline analysis solely, or require individualized training data to facilitate online reconstruction. We have proposed CLEEGN, a novel convolutional neural network for plug-and-play automatic EEG reconstruction. CLEEGN is based on a subject-independent pre-trained model using existing data and can operate on a new user without any further calibration. The performance of CLEEGN was validated using multiple evaluations including waveform observation, reconstruction error assessment, and decoding accuracy on well-studied labeled datasets. The results of simulated online validation suggest that, even without any calibration, CLEEGN can largely preserve inherent brain activity and outperforms leading online/offline artifact removal methods in the decoding accuracy of reconstructed EEG data. In addition, visualization of model parameters and latent features exhibit the model behavior and reveal explainable insights related to existing knowledge of neuroscience. We foresee pervasive applications of CLEEGN in prospective works of online plug-and-play EEG decoding and analysis.
Cellular detoxification is important for the routine removal of environmental and dietary carcinogens. Glutathione S-transferases (GST) are major cellular phase II detoxification enzymes. MRC-5 cells ...have been found to exhibit significantly higher GST activity than human H1355 cells. This study investigates whether GST-M2 activity acts as a critical determinant of the target dose of carcinogenic benzoapyrene-diolepoxide (BPDE) and whether it has an effect on
MDM2 splicing in the two cell lines. We used RT-PCR to clone Mu-class GST cDNA. Two forms of GST coming from the cell lines were characterized as GST-M2 (from MRC-5 cells) and GST-M4 (from H1355 cells). Nested-PCR showed that BPDE-induced
MDM2 splicing had occurred in the H1355 cell line but not in normal MRC-5 cells. Furthermore, using nested-PCR and competitive ELISA, we found that in H1355 cells modified to stably overexpress GST-M2, splicing was abolished and BPDE adducts appeared in low abundance. In conclusion, exogenously overexpressed GST-M2 was effective in reducing BPDE-induced DNA damage in H1355 cells. The catalytic activity of GST-M2 may play an important future role in lowering the incidence of BPDE-induced DNA damage.
Dipeptidyl peptidase-4 (DPP-4) inhibitors are oral anti-hyperglycemic drugs enabling effective glycemic control in type 2 diabetes (T2D). Despite DPP-4 inhibitors' advantages, the patients' ...therapeutic response varies. In this retrospective cohort study, 171 Taiwanese patients with T2D were classified as sensitive or resistant to treatment based on the mean change in HbA1c levels. Using an assumption-free genome-wide association study, 45 single nucleotide polymorphisms (SNPs) involved in the therapeutic response to DPP-4 inhibitors (P < 1 × 10-4) were identified at or near PRKD1, CNTN3, ASK, and LOC10537792. A SNP located within the fourth intron of PRKD1 (rs57803087) was strongly associated with DPP-4 inhibitor response (P = 3.2 × 10-6). This is the first pharmacogenomics study on DPP-4 inhibitor treatment for diabetes in a Taiwanese population. Our data suggest that genes associated with β-cell function and apoptosis are involved in the therapeutic effect of DPP-4 inhibitors, even in the presence of additional oral anti-diabetic drugs. Our findings provide information on how genetic variants influence drug response and may benefit the development of personalized medicine.
The incidence rate of endometrial hyperplasia (EH) and endometrial cancer (EC) have doubled over the last two decades going along with a drop in average diagnostic age and a statistically significant ...reduction in survival rate. The exploration of four genomic clusters by The Cancer Genome Atlas (TCGA) Network provides molecular insights and opens up the opportunities to improve the current management strategies. However, the advancement of genomic-guided treatment is hampered by the lack of appropriate disease models to obtain prospective validation of the relationship between cluster-association and treatment response. Work in this dissertation attempts to develop predictive, clinically relevant mouse models for treatment evaluation through three approaches: 1) providing a new mouse model for EH, 2) establishing and characterizing endometrial cancer patient-derived xenografts (EC-PDXs) for drug screening, 3) refining the current treatment strategy based on genomic classification using PDX preclinical trials. The doubled incidence rate suggests there is a need to develop a reliable, clinically relevant precancer model to study tumorigenesis and identify prevention strategies. In the first approach, an estrogen-induced EH mouse model was developed. Histological analysis of endometrial tissues demonstrated that this mouse model is capable of reflecting different stages of disease development, hormonal receptors status, and genetic alterations as seen in the human disease. In order to provide improved treatment strategies for EC patients based on genomic classification, a predictive model that can provide a broad spectrum of preclinical efficacy is required. In the second approach, a panel of EC-PDXs was developed by orthotopically transplanting patient cancer tissue specimens into murine uteri and propagating them over multiple generations. Established tumor grafts retain crucial histo-pathological characteristics, the capacity to form distant metastasis following known clinical patterns, as well as recapitulating molecular features of the original human tumor specimens. This model serves as a tool to investigate the need for reclassifying EC and feasibility of directing genomic-guided treatment-based on TCGA recommendations. The last approach aims to validate the predictive capabilities of utilizing genomic classification and cluster affiliation in the refinement of current treatment strategies conducting PDX preclinical trials. The results demonstrate that selecting therapeutics based on affiliation to genomic clusters indeed results in improved treatment response and accuracy of prediction, and genomic similarities across various cancer types can be used to direct treatment. The endeavor described in this dissertation aims to provide a more predictive, clinically relevant model for EH and EC. The results demonstrated the utility of genomic classification to enable the prediction of drug response, and thus may facilitate the refinement of current management in a more precise and personalized way.
Abstract
Recurrence of metastatic breast cancer stemming from acquired endocrine and chemotherapy resistance remains a health burden for women with luminal (ER+) breast cancer. Disseminated ER+ tumor ...cells can remain viable but quiescent for years to decades. Contributing factors to metastatic spread include the maintenance and expansion of breast cancer stem cells (CSCs). Breast CSCs are poorly proliferative and frequently exist as a minority population in therapy resistant tumors. Our objective is to define novel signaling pathways that govern therapy resistance in ER+ breast cancer. In this study, we show that cytoplasmic complexes composed of steroid receptor (SR) co-activators, PELP1 and SRC-3, modulate breast CSC expansion through upregulation of the HIF-activated metabolic target genes PFKFB3 and PFKFB4. Seahorse metabolic assays demonstrated that cytoplasmic PELP1 influences cellular metabolism by increasing both glycolysis and mitochondrial respiration. PELP1 interacts with PFKFB3 and PFKFB4 proteins, and inhibition of PFKFB3 and PFKFB4 kinase activity blocks PELP1-induced tumorspheres and protein-protein interactions with SRC-3. PFKFB4 knockdown inhibited in vivo emergence of circulating tumor cell (CTC) populations in ER+ mammary intraductal (MIND) xenografts. Application of PFKFB inhibitors in combination with ER targeted therapies blocked tumorsphere formation in multiple models of advanced breast cancer, including tamoxifen (TamR) and paclitaxel (TaxR) resistant models and ER+ patient-derived organoids (PDxO). Together, our data suggest that PELP1, SRC-3, and PFKFBs cooperate to drive ER+ tumor cells that include CSCs and CTCs. Identifying non-ER pharmacological targets offers a useful approach to blocking metastatic escape from standard of care ER/estrogen (E2)-targeted strategies to overcome endocrine and chemotherapy resistance in ER+ breast cancer.
A swept-source optical coherence tomography system is used to clinically scan oral precancer and cancer patients for statistically analyzing the effective indicators of diagnosis including the signal ...standard deviation, spatial-frequency spectral shape, and epithelium thickness.
PDF
![Alleviation of benzo[a]pyre...](http://api.summon.serialssolutions.com/2.0.0/image/issn/XR4PS5YY4K/1568-7864/small "Alleviation of benzo[a]pyrene-diolepoxide-DNA damage in human lung carcinoma by glutathione S-transferase M2")
