Browsing by Subject "Proto-oncogene proteins"

Now showing 1 - 5 of 5
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    Development and evaluation of ActSeq: A targeted next-generation sequencing panel for clinical oncology use
    (PLOS, 2022-04-21) Shi, Zonggao; Lopez, Jacqueline; Kalliney, William; Sutton, Bobbie; Simpson, Joyce; Maggert, Kevin; Liu, Sheng; Wan, Jun; Stack, M. Sharon; Medical and Molecular Genetics, School of Medicine
    Purpose: The demand for high-throughput genetic profiling of somatic mutations in cancer tissues is growing. We sought to establish a targeted next generation sequencing (NGS) panel test for clinical oncology practice. Methods: Customized probes were designed to capture exonic regions of 141 genes selected for the panel, which was aimed for the detection of clinically actionable genetic variations in cancer, including KRAS, NRAS, BRAF, ALK, ROS1, KIT and EGFR. The size of entire targeted regions is 0.8 Mb. Library preparation used NEBNext Ultra II FS kit coupled with target enrichment. Paired-end sequencing was run on Illumina NextSeq 500 at a read length of 150 nt. A bioinformatics workflow focusing on single nucleotide variant and short insertions and deletions (SNV/indel) discovery was established using open source, in-house and commercial software tools. Standard reference DNA samples were used in testing the sensitivity and precision and limit of detection in variant calling. Results: The general performance of the panel was observed in pilot runs. Average total reads per sample ranged from 30 million to 48 million, 73% ~82% unique reads. All runs had more than 99% average mapping rate. Mean target coverage ranged from 727x to 879x. Depth of coverage at 50x or more reached 87% of targeted region and 60% of targeted region received 500x or more coverage depth. Using OncoSpan HD827 DNA, which bears 144 variants (SNV/indel) from 80 genes that are within the targeted region on the panel, our somatic variant calling pipeline reached 97% sensitivity and 100% precision respectively, with near 48 million reads. High concordance with orthogonal approaches in variant detection was further verified with 7 cancer cell lines and 45 clinical specimens. Conclusion: We developed a NGS panel with a focus on clinically actionable gene mutations and validated the performance in library construction, sequencing and variant calling. High concordance with reference materials and orthogonal mutation detection was observed.
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    MYC and HSF1 Cooperate to Drive Sensitivity to Polo-like Kinase 1 Inhibitor Volasertib in High-grade Serous Ovarian Cancer
    (American Association for Cancer Research, 2025) Williams, Imade; O’Malley, Matthew; DeHart, Haddie; Walker, Bobby; Ulhaskumar, Vrushabh; Jothirajah, Pranav; Ray, Haimanti; Landrum, Lisa M.; Delaney, Joe R.; Nephew, Kenneth P.; Carpenter, Richard L.; Obstetrics and Gynecology, School of Medicine
    We show that HSF1 and MYC genes are co-amplified in more than 30% of HGSOC and demonstrate that HSF1 and MYC functionally cooperate to drive the growth of HGSOC cells. This work provides the foundation for HSF1 and MYC co-amplification as a biomarker for treatment efficacy of the polo-like kinase 1 inhibitor volasertib in HGSOC.
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    PLK1 Inhibition Induces Synthetic Lethality in Fanconi Anemia Pathway-Deficient Acute Myeloid Leukemia
    (American Association for Cancer Research, 2025) Sheth, Aditya S.; Chan, Ka-Kui; Liu, Sheng; Wan, Jun; Angus, Steve P.; Rhodes, Steven D.; Mitchell, Dana K.; Davis, Christopher; Ridinger, Maya; Croucher, Peter J.; Zeidan, Amer M.; Wijeratne, Aruna; Qian, Shaomin; Tran, Ngoc Tung; Sierra Potchanant, Elizabeth A.; Pediatrics, School of Medicine
    This work demonstrates that FA pathway mutations, which are frequently observed in sporadic AML, induce hypersensitivity to PLK1 inhibition, providing rationale for a novel synthetic lethal therapeutic strategy for this patient population.
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    Quantitative phosphoproteomic analysis identifies novel functional pathways of tumor suppressor DLC1 in estrogen receptor positive breast cancer.
    (PLOS, 2018-10-02) Gökmen-Polar, Yesim; True, Jason D.; Vieth, Edyta; Gu, Yuan; Gu, Xiaoping; Qi, Guihong D.; Mosley, Amber L.; Badve, Sunil S.; Pathology and Laboratory Medicine, School of Medicine
    Deleted in Liver Cancer-1 (DLC1), a member of the RhoGAP family of proteins, functions as a tumor suppressor in several cancers including breast cancer. However, its clinical relevance is unclear in breast cancer. In this study, expression of DLC1 was correlated with prognosis using publicly available breast cancer gene expression datasets and quantitative Reverse Transcription PCR in cohorts of Estrogen Receptor-positive (ER+) breast cancer. Low expression of DLC1 correlates with poor prognosis in patients with ER+ breast cancer with further decrease in metastatic lesions. The Cancer Genome Atlas (TCGA) data showed that down regulation of DLC1 is not due to methylation or mutations. To seek further insights in understanding the role of DLC1 in ER+ breast cancer, we stably overexpressed DLC1-full-length (DLC1-FL) in T-47D breast cancer cells; this inhibited cell colony formation significantly in vitro compared to its control counterpart. Label-free global proteomic and TiO2 phosphopeptide enrichment assays (ProteomeXchange identifier PXD008220) showed that 205 and 122 phosphopeptides were unique to DLC1-FL cells and T-47D-control cells, respectively, whereas 6,726 were quantified by phosphoproteomics analysis in both conditions. The top three significant clusters of differentially phosphopeptides identified by DAVID pathway analysis represent cell-cell adhesion, mRNA processing and splicing, and transcription regulation. Phosphoproteomics analysis documented an inverse relation between DLC1 expression and several phosphopeptides including epithelial cell transforming sequence 2 (ECT2). Decreased phosphorylation of ECT2 at the residue T359, critical for its active conformational change, was validated by western blot. In addition, the ECT2 T359-containing phosphopeptide was detected in both basal and luminal patient-derived breast cancers breast cancer phosphoproteomics data on the Clinical Proteomic Tumor Analysis Consortium (CPTAC) Assay portal. Together, for the first time, this implicates ECT2 phosphorylation in breast cancer, which has been proposed as a therapeutic target in lung cancer. In conclusion, this data suggests that low expression of DLC1 is associated with poor prognosis. Targeting ECT2 phosphopeptides could provide a promising mechanism for controlling poor prognosis seen in DLC1low ER+ breast cancer.
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    Vasculogenic skin reprogramming requires TET-mediated gene demethylation in fibroblasts for rescuing impaired perfusion in diabetes
    (Springer Nature, 2024-11-27) Mohanty, Sujit K.; Singh, Kanhaiya; Kumar, Manishekhar; Verma, Sumit S.; Srivastava, Rajneesh; Gnyawali, Surya C.; Palakurti, Ravichand; Sahi, Ajay K.; El Masry, Mohamed S.; Banerjee, Pradipta; Kacar, Sedat; Rustagi, Yashika; Verma, Priyanka; Ghatak, Subhadip; Hernandez, Edward; Rubin, J. Peter; Khanna, Savita; Roy, Sashwati; Yoder, Mervin C.; Sen, Chandan K.; Surgery, School of Medicine
    Tissue nanotransfection (TNT) topically delivers Etv2, Foxc2, and Fli1 (EFF) plasmids increasing vasculogenic fibroblasts (VF) and promoting vascularization in ischemic murine skin. Human dermal fibroblasts respond to EFF nanoelectroporation with elevated expression of endothelial genes in vitro, which is linked to increased ten-eleven translocase 1/2/3 (TET) expression. Single cell RNA sequencing dependent validation of VF induction reveals a TET-dependent transcript signature. TNTEFF also induces TET expression in vivo, and fibroblast-specific EFF overexpression leads to VF-transition, with TET-activation correlating with higher 5-hydroxymethylcytosine (5-hmC) levels in VF. VF emergence requires TET-dependent demethylation of endothelial genes in vivo, enhancing VF abundance and restoring perfusion in diabetic ischemic limbs. TNTEFF improves perfusion and wound closure in diabetic mice, while increasing VF in cultured human skin explants. Suppressed in diabetes, TET1/2/3 play a critical role in TNT-mediated VF formation which supports de novo blood vessel development to rescue diabetic ischemic tissue.