Background: To investigate deep vein thrombosis (DVT) in hospitalized patients with coronavirus disease 2019 (COVID-19), we performed a single institutional study to evaluate its prevalence, risk factors, prognosis, and potential thromboprophylaxis strategies in a large referral and treatment center. Methods: We studied a total of 143 patients with COVID-19 from January 29, 2020 to February 29, 2020. Demographic and clinical data, laboratory data, including ultrasound scans of the lower extremities, and outcome variables were obtained, and comparisons were made between groups with and without DVT. Results: Of the 143 patients hospitalized with COVID-19 (age 63±14 years, 74 [51.7%] men), 66 patients developed lower extremity DVT (46.1%: 23 [34.8%] with proximal DVT and 43 [65.2%] with distal DVT). Compared with patients who did not have DVT, patients with DVT were older and had a lower oxygenation index, a higher rate of cardiac injury, and worse prognosis, including an increased proportion of deaths (23 [34.8%] versus 9 [11.7%]; P =0.001) and a decreased proportion of patients discharged (32 [48.5%] versus 60 [77.9%]; P <0.001). Multivariant analysis showed an association only between CURB-65 (confusion status, urea, respiratory rate, and blood pressure) score 3 to 5 (odds ratio, 6.122; P =0.031), Padua prediction score ≥4 (odds ratio, 4.016; P =0.04), D-dimer >1.0 μg/mL (odds ratio, 5.818; P <0.014), and DVT in this cohort, respectively. The combination of a CURB-65 score 3 to 5, a Padua prediction score ≥4, and D-dimer >1.0 μg/mL has a sensitivity of 88.52% and a specificity of 61.43% for screening for DVT. In the subgroup of patients with a Padua prediction score ≥4 and whose ultrasound scans were performed >72 hours after admission, DVT was present in 18 (34.0%) patients in the subgroup receiving venous thromboembolism prophylaxis versus 35 (66.0%) patients in the nonprophylaxis group ( P =0.010). Conclusions: The prevalence of DVT is high and is associated with adverse outcomes in hospitalized patients with COVID-19. Prophylaxis for venous thromboembolism may be protective in patients with a Padua protection score ≥4 after admission. Our data seem to suggest that COVID-19 is probably an additional risk factor for DVT in hospitalized patients.

Our effort to identify novel drug-resistant genes in cyclophosphamide-resistant EMT6 mouse mammary tumors led us to the identification of SPF45. Simultaneously, other groups identified SPF45 as a component of the spliceosome that is involved in alternative splicing. We isolated the human homologue and examined the normal human tissue expression, tumor expression, and the phenotype caused by overexpression of human SPF45. Our analyses revealed that SPF45 is expressed in many, but not all, normal tissues tested with predominant expression in normal ductal epithelial cells of the breast, liver, pancreas, and prostate. Our analyses using tissue microarrays and sausages of tumors indicated that SPF45 is highly expressed in numerous carcinomas including bladder, breast, colon, lung, ovarian, pancreatic, and prostate. Interestingly, this study revealed that overexpression of SPF45 in HeLa, a cervical carcinoma cell line, resulted in drug resistance to doxorubicin and vincristine, two chemotherapeutic drugs commonly used in cancer. To our knowledge, this is the first study showing tumor overexpression of an alternate splicing factor resulting in drug resistance. The development of multidrug resistance is a major hurdle in the treatment of cancer. Although some genes that confer multidrug resistance are known, it is clear that many other genes remain to be identified. The characterization of novel mechanisms that lead to drug resistance would enable the development of a targeted new generation of anti-cancer drugs. To identify novel genes involved in drug resistance we performed suppressivesubtractive polymerase chain reaction analyses of the cyclophosphamide-resistant EMT6 mouse mammary tumor cells and the parental EMT6 tumors. This led to the identification of a gene that was highly homologous to a DNA damage response protein, DRT111 of Arabidopsis (unpublished observations). 1 While this work was in progress, Neubauer and colleagues, 1 identified a novel protein from the spliceosome. An analysis of all proteins present in the spliceosomal complex led to the identification of 25 previously identified proteins and 20 novel proteins. One of the novel proteins thus identified was named SPF45 (splicing factor 45 kd). This protein was identical to the one that was identified by our suppressive-subtractive polymerase chain reaction analysis. Neubauer and colleagues, 1 further showed that SPF45 was a nuclear protein that co-localizes with U1A snRNP in nuclear speckles, a known reservoir of splicing factors. [2][3][4] Recently Rappsilber and colleagues, 5 have described the identification of 311 proteins in the human spliceosomal complex of which 96 were novel proteins. Again, in this study, SPF45 was identified as a component of the spliceosome.Lallena and colleagues, 6 demonstrated that SPF45 regulates the alternate splicing of the Drosophila sexlethal gene (sxl). SXL protein is expressed only in female flies, but not in males. 7 Exon3 of Sxl plays a very important role in determining the expression of SXL because its inclus...

Transforming growth factor beta (TGF-beta) signaling pathways regulate a wide variety of cellular processes including cell proliferation, differentiation, extracellular matrix deposition, development, and apoptosis. TGF-beta type-I receptor (TbetaRI) is the major receptor that triggers several signaling events by activating downstream targets such as the Smad proteins. The intracellular kinase domain of TbetaRI is essential for its function. In this study, we have identified a short phospho-Smad peptide, pSmad3(-3), KVLTQMGSPSIRCSS(PO4)VS as a substrate of TbetaRI kinase for in vitro kinase assays. This peptide is uniquely phosphorylated by TbetaRI kinase at the C-terminal serine residue, the phosphorylation site of its parent Smad protein in vivo. Specificity analysis demonstrated that the peptide is phosphorylated by only TbetaRI and not TGF-beta type-II receptor kinase, indicating that the peptide is a physiologically relevant substrate suitable for kinetic analysis and screening of TbetaRI kinase inhibitors. Utilizing pSmad3(-3) as a substrate, we have shown that novel pyrazole compounds are potent inhibitors of TbetaRI kinase with K(i) value as low as 15 nM. Kinetic analysis revealed that these pyrazoles act through the ATP-binding site and are typical ATP competitive inhibitors with tight binding kinetics. More importantly, these compounds were shown to inhibit TGF-beta-induced Smad2 phosphorylation in vivo in NMuMg mammary epithelial cells with potency equivalent to the inhibitory activity in the in vitro kinase assay. Cellular selectivity analysis demonstrated that these pyrazoles are capable of inhibiting activin signaling but not bone morphogenic protein or platelet-derived growth factor signal transduction pathways. Further functional analysis revealed that pyrazoles are capable of blocking the TGF-beta-induced epithelial-mesenchymal transition in NMuMg cells, a process involved in the progression of cancer, fibrosis, and other human diseases. These pyrazoles provide a foundation for future development of potent and selective TbetaRI kinase inhibitors to treat human disease.

b Picorna-like viruses in the Picornavirales order are a large group of positive-strand RNA viruses that include numerous important pathogens for plants, insects, and humans. In these viruses, nonstructural protein 2C is one of the most conserved proteins and contains ATPase activity and putative RNA helicase activity. Here we expressed 2C protein of Ectropis obliqua picorna-like virus (EoV; genus Iflavirus, family Iflaviridae, order Picornavirales) in a eukaryotic expression system and determined that EoV 2C displays ATP-independent nucleic acid helix destabilizing and strand annealing acceleration activity in a concentration-dependent manner, indicating that this picornaviral 2C is more like an RNA chaperone than like the previously predicted RNA helicase. Our further characterization of EoV 2C revealed that divalent metal ions, such as Mg 2؉ and Zn 2؉ , inhibit 2C-mediated helix destabilization to different extents. Moreover, we determined that EoV 2C also contains ATPase activity like that of other picornaviral 2C proteins and further assessed the functional relevance between its RNA chaperone-like and ATPase activities using mutational analysis as well as their responses to Mg 2؉ . Our data show that, when one of the two 2C activities was dramatically inhibited or almost abolished, the other activity could remain intact, showing that the RNA chaperone-like and ATPase activities of EoV 2C can be functionally separated. This report reveals that a picorna-like virus 2C protein displays RNA helix destabilizing and strand annealing acceleration activity, which may be critical for picornaviral replication and pathogenesis, and should foster our understanding of picorna-like viruses and viral RNA chaperones.