Olaparib combined with CDK12-IN-3 to promote genomic instability and cell death in ovarian cancer.

Large-scale phase III clinical trials of Olaparib have revealed benefits for ovarian cancer patients with BRCA gene mutations or homologous recombination deficiency (HRD). However, fewer than 50% of ovarian cancer patients have both BRCA mutations and HRD. Therefore, improving the effect of Olaparib in HR-proficient patients is of great clinical value. Here, a combination strategy comprising Olaparib and CDK12-IN-3 effectively inhibited the growth of HR-proficient ovarian cancer in cell line, patient-derived organoid (PDO), and mouse xenograft models. Furthermore, the combination strategy induced severe DNA double-strand break (DSB) formation, increased NHEJ activity in the G2 phase, and reduced HR activity in cancer cells. Mechanistically, the combination treatment impaired Ku80 poly(ADP-ribosyl)ation (PARylation) and phosphorylation, resulting in PARP1-Ku80 complex dissociation. After dissociation, Ku80 occupancy at DSBs and the resulting Ku80-primed NHEJ activity were increased. Owing to Ku80-mediated DNA end protection, MRE11 and Rad51 foci formation was inhibited after the combination treatment, suggesting that this treatment suppressed HR activity. Intriguingly, the combination strategy expedited cGAS nuclear relocalization, further suppressing HR and, conversely, increasing genomic instability. Moreover, the inhibitory effect on cell survival persisted after drug withdrawal. These findings provide a rationale for the clinical application of CDK12-IN-3 in combination with Olaparib.

MED12 and CDK8/19 Modulate Androgen Receptor Activity and Enzalutamide Response in Prostate Cancer.

Prostate cancer progression is driven by androgen receptor (AR) activity, which is a target for therapeutic approaches. Enzalutamide is an AR inhibitor that prolongs the survival of patients with advanced prostate cancer. However, resistance mechanisms arise and impair its efficacy. One of these mechanisms is the expression of AR-V7, a constitutively active AR splice variant. The Mediator complex is a multisubunit protein that modulates gene expression on a genome-wide scale. MED12 and cyclin-dependent kinase (CDK)8, or its paralog CDK19, are components of the kinase module that regulates the proliferation of prostate cancer cells. In this study, we investigated how MED12 and CDK8/19 influence cancer-driven processes in prostate cancer cell lines, focusing on AR activity and the enzalutamide response. We inhibited MED12 expression and CDK8/19 activity in LNCaP (AR+, enzalutamide-sensitive), 22Rv1 (AR-V7+, enzalutamide-resistant), and PC3 (AR-, enzalutamide-insensitive) cells. Both MED12 and CDK8/19 inhibition reduced cell proliferation in all cell lines, and MED12 inhibition reduced proliferation in the respective 3D spheroids. MED12 knockdown significantly inhibited c-Myc protein expression and signaling pathways. In 22Rv1 cells, it consistently inhibited the AR response, prostate-specific antigen (PSA) secretion, AR target genes, and AR-V7 expression. Combined with enzalutamide, MED12 inhibition additively decreased the AR activity in both LNCaP and 22Rv1 cells. CDK8/19 inhibition significantly decreased PSA secretion in LNCaP and 22Rv1 cells and, when combined with enzalutamide, additively reduced proliferation in 22Rv1 cells. Our study revealed that MED12 and CDK8/19 regulate AR activity and that their inhibition may modulate response to enzalutamide in prostate cancer.

Structural basis of Cdk7 activation by dual T-loop phosphorylation.

Cyclin-dependent kinase 7 (Cdk7) is required in cell-cycle and transcriptional regulation owing to its function as both a CDK-activating kinase (CAK) and part of transcription factor TFIIH. Cdk7 forms active complexes by associating with Cyclin H and Mat1, and is regulated by two phosphorylations in the activation segment (T loop): the canonical activating modification at T170 and another at S164. Here we report the crystal structure of the human Cdk7/Cyclin H/Mat1 complex containing both T-loop phosphorylations. Whereas pT170 coordinates basic residues conserved in other CDKs, pS164 nucleates an arginine network unique to the ternary Cdk7 complex, involving all three subunits. We identify differential dependencies of kinase activity and substrate recognition on the individual phosphorylations. CAK function is unaffected by T-loop phosphorylation, whereas activity towards non-CDK substrates is increased several-fold by T170 phosphorylation. Moreover, dual T-loop phosphorylation stimulates multisite phosphorylation of the RNA polymerase II (RNAPII) carboxy-terminal domain (CTD) and SPT5 carboxy-terminal repeat (CTR) region. In human cells, Cdk7 activation is a two-step process wherein S164 phosphorylation precedes, and may prime, T170 phosphorylation. Thus, dual T-loop phosphorylation can regulate Cdk7 through multiple mechanisms, with pS164 supporting tripartite complex formation and possibly influencing processivity, while pT170 enhances activity towards key transcriptional substrates.

Activation of the CDK7 Gene, Coding for the Catalytic Subunit of the Cyclin-Dependent Kinase (CDK)-Activating Kinase (CAK) and General Transcription Factor II H, by the Trans-Activator Protein Tax of Human T-Cell Leukemia Virus Type-1.

Human T-cell leukemia virus type-1 (HTLV-1) is the etiological agent of adult T-cell leukemia (ATL). The trans-activator protein Tax of HTLV-1 plays crucial roles in leukemogenesis by promoting proliferation of virus-infected cells through activation of growth-promoting genes. However, critical target genes are yet to be elucidated. We show here that Tax activates the gene coding for cyclin-dependent kinase 7 (CDK7), the essential component of both CDK-activating kinase (CAK) and general transcription factor TFIIH. CAK and TFIIH play essential roles in cell cycle progression and transcription by activating CDKs and facilitating transcriptional initiation, respectively. Tax induced CDK7 gene expression not only in human T-cell lines but also in normal peripheral blood lymphocytes (PHA-PBLs) along with increased protein expression. Tax stimulated phosphorylation of CDK2 and RNA polymerase II at sites reported to be mediated by CDK7. Tax activated the CDK7 promoter through the NF-κB pathway, which mainly mediates cell growth promotion by Tax. Knockdown of CDK7 expression reduced Tax-mediated induction of target gene expression and cell cycle progression. These results suggest that the CDK7 gene is a crucial target of Tax-mediated trans-activation to promote cell proliferation by activating CDKs and transcription.

Overexpression of Orysa;KRP4 drastically reduces grain filling in rice.

MAIN CONCLUSION: Excess of KRP4 in the developing kernels in rice causes poor filling of the grains possibly through inhibition of CDKA;2 and CDKB;1 activity mediated by its interaction with CDKF;3. The potential yield of the rice varieties producing compact and heavy panicles bearing numerous spikelets is compromised because a high percentage of spikelets remain poorly filled, reportedly because of a high expression of KRPs that causes suppression of endosperm cell proliferation. To test the stated negative relationship between KRP expression and grain filling, Orysa;KRP4 was overexpressed under the control of seed-specific glutelin promoter in IR-64 rice variety that shows good grain filling. The transgenic lines showed more than 15-fold increase in expression of KRP4 in the spikelets concomitant with nearly 50% reduction in grain filling compared with the wild type without producing any significant changes on the other yield-related parameters like panicle length and the spikelets numbers that were respectively 30.23 ± 0.89 cm and 229.25 ± 33.72 per panicle in the wild type, suggesting a highly organ-targeted effect of the genetic transformation. Yeast two-hybrid test revealed CDKF;3 as the interacting partner of KRP4, and CDKF;3 was found to interact with CDKA;2, CDKB;1 and CDKD;1. Significant decrease in grain filling in the transgenic lines compared with the wild type due to overexpression of KRP4 could be because of suppression of the activity of CDKB;1 and CDKA;2 by inhibition of their phosphorylation directly by CDKF;3, or mediated through inhibition of phosphorylation of CDKD;1 by CDKF;3. The study thus indicated that suppression of expression of KRP(s) by genetic manipulation of their promoters could be an important way of improving the yield of the rice varieties bearing compact and heavy panicles.

CDK12 controls transcription at damaged genes and prevents MYC-induced transcription-replication conflicts.

The identification of genes involved in replicative stress is key to understanding cancer evolution and to identify therapeutic targets. Here, we show that CDK12 prevents transcription-replication conflicts (TRCs) and the activation of cytotoxic replicative stress upon deregulation of the MYC oncogene. CDK12 was recruited at damaged genes by PARP-dependent DDR-signaling and elongation-competent RNAPII, to repress transcription. Either loss or chemical inhibition of CDK12 led to DDR-resistant transcription of damaged genes. Loss of CDK12 exacerbated TRCs in MYC-overexpressing cells and led to the accumulation of double-strand DNA breaks, occurring between co-directional early-replicating regions and transcribed genes. Overall, our data demonstrate that CDK12 protects genome integrity by repressing transcription of damaged genes, which is required for proper resolution of DSBs at oncogene-induced TRCs. This provides a rationale that explains both how CDK12 deficiency can promote tandem duplications of early-replicated regions during tumor evolution, and how CDK12 targeting can exacerbate replicative-stress in tumors.

Dual-site molecular glues for enhancing protein-protein interactions of the CDK12-DDB1 complex.

Protein-protein interactions (PPIs) stabilization with molecular glues plays a crucial role in drug discovery, albeit with significant challenges. In this study, we propose a dual-site approach, targeting the PPI region and its dynamic surroundings. We conduct molecular dynamics simulations to identify critical sites on the PPI that stabilize the cyclin-dependent kinase 12 - DNA damage-binding protein 1 (CDK12-DDB1) complex, resulting in further cyclin K degradation. This exploration leads to the creation of LL-K12-18, a dual-site molecular glue, which enhances the glue properties to augment degradation kinetics and efficiency. Notably, LL-K12-18 demonstrates strong inhibition of gene transcription and anti-proliferative effects in tumor cells, showing significant potency improvements in MDA-MB-231 (88-fold) and MDA-MB-468 cells (307-fold) when compared to its precursor compound SR-4835. These findings underscore the potential of dual-site approaches in disrupting CDK12 function and offer a structural insight-based framework for the design of cyclin K molecular glues.

Modeling the START transition in the budding yeast cell cycle.

Budding yeast, Saccharomyces cerevisiae, is widely used as a model organism to study the genetics underlying eukaryotic cellular processes and growth critical to cancer development, such as cell division and cell cycle progression. The budding yeast cell cycle is also one of the best-studied dynamical systems owing to its thoroughly resolved genetics. However, the dynamics underlying the crucial cell cycle decision point called the START transition, at which the cell commits to a new round of DNA replication and cell division, are under-studied. The START machinery involves a central cyclin-dependent kinase; cyclins responsible for starting the transition, bud formation, and initiating DNA synthesis; and their transcriptional regulators. However, evidence has shown that the mechanism is more complicated than a simple irreversible transition switch. Activating a key transcription regulator SBF requires the phosphorylation of its inhibitor, Whi5, or an SBF/MBF monomeric component, Swi6, but not necessarily both. Also, the timing and mechanism of the inhibitor Whi5's nuclear export, while important, are not critical for the timing and execution of START. Therefore, there is a need for a consolidated model for the budding yeast START transition, reconciling regulatory and spatial dynamics. We built a detailed mathematical model (START-BYCC) for the START transition in the budding yeast cell cycle based on established molecular interactions and experimental phenotypes. START-BYCC recapitulates the underlying dynamics and correctly emulates key phenotypic traits of ~150 known START mutants, including regulation of size control, localization of inhibitor/transcription factor complexes, and the nutritional effects on size control. Such a detailed mechanistic understanding of the underlying dynamics gets us closer towards deconvoluting the aberrant cellular development in cancer.

Understanding the Cytomegalovirus Cyclin-Dependent Kinase Ortholog pUL97 as a Multifaceted Regulator and an Antiviral Drug Target.

Herpesviral protein kinases, such as the therapy-relevant pUL97 of human cytomegalovirus (HCMV), are important for viral replication efficiency as well as pathogenesis, and represent key antiviral drug targets. HCMV pUL97 is a viral cyclin-dependent kinase (CDK) ortholog, as it shares functional and structural properties with human CDKs. Recently, the formation of vCDK/pUL97-cyclin complexes and the phosphorylation of a variety of viral and cellular substrate proteins has been demonstrated. Genetic mapping and structural modeling approaches helped to define two pUL97 interfaces, IF1 and IF2, responsible for cyclin binding. In particular, the regulatory importance of interactions between vCDK/pUL97 and host cyclins as well as CDKs has been highlighted, both as determinants of virus replication and as a novel drug-targeting option. This aspect was substantiated by the finding that virus replication was impaired upon cyclin type H knock-down, and that such host-directed interference also affected viruses resistant to existing therapies. Beyond the formation of binary interactive complexes, a ternary pUL97-cyclin H-CDK7 complex has also been described, and in light of this, an experimental trans-stimulation of CDK7 activity by pUL97 appeared crucial for virus-host coregulation. In accordance with this understanding, several novel antiviral targeting options have emerged. These include kinase inhibitors directed to pUL97, to host CDKs, and to the pUL97-cyclin H interactive complexes. Importantly, a statistically significant drug synergy has recently been reported for antiviral treatment schemes using combinations of pharmacologically relevant CDK7 and vCDK/pUL97 inhibitors, including maribavir. Combined, such findings provide increased options for anti-HCMV control. This review focuses on regulatory interactions of vCDK/pUL97 with the host cyclin-CDK apparatus, and it addresses the functional relevance of these key effector complexes for viral replication and pathogenesis. On this basis, novel strategies of antiviral drug targeting are defined.

Anti-Tumor and Chemosensitizing Effects of the CDK Inhibitor Dinaciclib on Cholangiocarcinoma In Vitro and In Vivo.

BACKGROUND/AIM: Cholangiocarcinoma (CCA) is a highly aggressive disease. Most of CCA patients are diagnosed in an advanced stage of the disease, when it is unresectable and there is chemoresistance, resulting in poor prognosis. However, effective therapeutic regimens and molecular targets for CCA remain poor. Cyclin-dependent kinases (CDKs) are key regulatory enzymes in cell cycle progression. Aberrant CDK activation is a hallmark of cancer. Dinaciclib is a small molecule inhibitor of multiple CDKs, currently under clinical evaluation for treating advanced malignancies. The efficacy of anti-tumor activity of dinaciclib against chemotherapy resistant CCA cells was examined in vitro and in vivo. MATERIALS AND METHODS: In this study, the effect of dinaciclib on growth and cell cycle in CCA cell lines were determined using the MTT assay and cell cycle analysis. The anti-tumor activity of dinaciclib was investigated in CCA-inoculated mice. In addition, the chemosensitizing effect of dinaciclib was investigated in gemcitabine-treated CCA cell lines. RESULTS: Dinaciclib significantly suppressed cell proliferation, induced G1/S phase cell cycle arrest and apoptosis of CCA cell lines. It significantly suppressed the growth of CCA cells in xenograft mouse models. We also found that dinaciclib significantly inhibited the growth of gemcitabine-resistant CCA cell lines (KKU-213A-GemR and KKU-100-GemR). Furthermore, dinaciclib significantly enhanced the anti-tumor activity of gemcitabine in CCA cell lines. CONCLUSION: Dinaciclib has the potential to be an effective therapeutic agent to control tumor cell growth of both parental and gemcitabine-resistant CCA cells.

Elucidating the Selective Mechanism of Drugs Targeting Cyclin-Dependent Kinases with Integrated MetaD-US Simulation.

Cyclin-dependent kinases (CDKs), including CDK12 and CDK13, play crucial roles in regulating the cell cycle and RNA polymerase II activity, making them vital targets for cancer therapies. SR4835 is a selective inhibitor of CDK12/13, showing significant potential for treating triple-negative breast cancer. To elucidate the selective mechanism of SR4835 among three CDKs (CDK13/12/9), we developed an innovative enhanced sampling method, integrated well-tempered metadynamics-umbrella sampling (IMUS). IMUS synergistically combines the comprehensive pathway exploration capability of well-tempered metadynamics (WT-MetaD) with the precise free energy calculation capability of umbrella sampling, enabling the efficient and accurate characterization of drug-target interactions. The accurate calculation of binding free energy and the detailed analysis of the kinetic mechanism of the drug-target interaction using IMUS successfully elucidate the drug selectivity mechanism targeting the three CDKs, showing that the selectivity is primarily arising from differences in the stability of H-bonds within the Hinge region of the kinases and the interaction patterns during the protein-ligand recognition process. These findings also underscore the utility of IMUS in efficiently and accurately capturing drug-target interaction processes with clear mechanisms.

The role of intrinsic protein disorder in regulation of cyclin-dependent kinases.

While the structure/function paradigm for folded domains was established decades ago, our understanding of how intrinsically disordered regions (IDRs) contribute to biological function is still evolving. IDRs exist as conformational ensembles that can range from highly compact to highly extended depending on their sequence composition. IDR sequences are less conserved than those of folded domains, but often display short, conserved segments termed short linear motifs (SLiMs), that often mediate protein-protein interactions and are often regulated by posttranslational modifications, giving rise to complex functionality when multiple, differently regulated SLiMs are combined. This combinatorial functionality was associated with signaling and regulation soon after IDRs were first recognized as functional elements within proteins. Here, we discuss roles for disorder in proteins that regulate cyclin-dependent kinases, the master timekeepers of the eukaryotic cell cycle. We illustrate the importance of intrinsic flexibility in the transmission of regulatory signals by these entirely disordered proteins.

Role of the CDKL1-SOX11 signaling axis in acute kidney injury.

The biology of the cyclin-dependent kinase-like (CDKL) kinase family remains enigmatic. Contrary to their nomenclature, CDKLs do not rely on cyclins for activation and are not involved in cell cycle regulation. Instead, they share structural similarities with mitogen-activated protein kinases and glycogen synthase kinase-3, although their specific functions and associated signaling pathways are still unknown. Previous studies have shown that the activation of CDKL5 kinase contributes to the development of acute kidney injury (AKI) by suppressing the protective SOX9-dependent transcriptional program in tubular epithelial cells. In the current study, we measured the functional activity of all five CDKL kinases and discovered that, in addition to CDKL5, CDKL1 is also activated in tubular epithelial cells during AKI. To explore the role of CDKL1, we generated a germline knockout mouse that exhibited no abnormalities under normal conditions. Notably, when these mice were challenged with bilateral ischemia-reperfusion and rhabdomyolysis, they were found to be protected from AKI. Further mechanistic investigations revealed that CDKL1 phosphorylates and destabilizes SOX11, contributing to tubular dysfunction. In summary, this study has unveiled a previously unknown CDKL1-SOX11 axis that drives tubular dysfunction during AKI.NEW & NOTEWORTHY Identifying and targeting pathogenic protein kinases holds potential for drug discovery in treating acute kidney injury. Our study, using novel germline knockout mice, revealed that Cdkl1 kinase deficiency does not affect mouse viability but provides protection against acute kidney injury. This underscores the importance of Cdkl1 kinase in kidney injury and supports the development of targeted small-molecule inhibitors as potential therapeutics.

Antitumoral activity of a CDK12 inhibitor in colorectal cancer through a liposomal formulation.

Colorectal cancer (CRC) is the third most common cancer worldwide. Recent experiments suggest that CDK12 can be a good therapeutic target in CRC, and therefore, novel inhibitors targeting this protein are currently in preclinical development. Lipid-based formulations of chemical entities have demonstrated the ability to enhance activity while improving the safety profile. In the present work, we explore the antitumor activity of a new CDK12 inhibitor (CDK12-IN-E9, CDK12i) and its lipid-based formulation (LP-CDK12i) in CRC models, to increase efficacy. SW620, SW480 and HCT116 CRC cell lines were used to evaluate the inhibitor and the liposomal formulation using MTT proliferation assay, 3D invasion cultures, flow cytometry, Western blotting and immunofluorescence experiments. Free-cholesterol liposomal formulations of CDK12i (LP-CDK12i) were obtained by solvent injection method and fully characterized by size, shape, polydispersity, encapsulation efficiency, and release profile and stability assessments. LP-CDK12i induced a higher antiproliferative effect compared with CDK12i as a free agent. The IC50 value was lower across all cell lines tested, leading to a reduction in cell proliferation and the formation of 3D structures. Evaluation of apoptosis revealed an increase in cell death, while biochemical studies demonstrated modifications of apoptosis and DNA damage components. In conclusion, we confirm the role of targeting CDK12 for the treatment of CRC and describe, for the first time, a liposomal formulation of a CDK12i with higher antiproliferative activity compared with the free compound.

Estimation of CDK inhibitors by RP-HPLC: application for pharmacokinetic interactions studies with PPIs.

Background: The study investigated pharmacokinetic interactions between palbociclib and ribociclib with proton pump inhibitors (PPIs) using the reverse-phase high-performance liquid chromatography (RP-HPLC) method.Methods: Developed RP-HPLC method quantified palbociclib and ribociclib in biological matrices. In vitro metabolic stability assays and in vivo studies in rats evaluated effect of omeprazole and esomeprazole on pharmacokinetics of palbociclib and ribociclib.Results: The RP-HPLC method was sensitive, accurate and linear. Esomeprazole and omeprazole decreased metabolic clearance of palbociclib and ribociclib by several folds. In vivo, esomeprazole elevated Cmax of palbociclib and ribociclib by 90.1% and 86.4%, whereas omeprazole reduced it by 32.0% and 16.8%, respectively.Conclusion: The RP-HPLC method was used to analyze in vitro and in vivo samples. Long-term treatment with PPIs affects pharmacokinetics of palbociclib and ribociclib, necessitating optimal chemotherapy regimen.

[Box: see text].

CDK8/19 inhibitor enhances arginase-1 expression in macrophages via STAT6 and p38 MAPK activation.

Macrophages polarize into alternatively activated M2 macrophages through interleukin (IL)-4, and they express high levels of arginase-1, which promotes anti-inflammatory responses. Several studies have confirmed the anti-inflammatory effects of cyclin-dependent kinase (CDK) 8/19 inhibition, and hence, numerous CDK8/19 inhibitors, such as BRD6989, have been developed. However, the effects of CDK8/19 inhibitors on arginase-1 expression in macrophages have not yet been elucidated. This study investigated the effects of CDK8/19 inhibitor on arginase-1 expression in IL-4-activated macrophages. The results showed that BRD6989 increased arginase-1 expression transcriptionally in murine peritoneal macrophages and the murine macrophage cell line RAW264.7 in an IL-4-dependent manner. In addition, the results indicated that BRD6989 enhances signal transducer and activator of transcription (STAT) 6 phosphorylation. Meanwhile, BRD6989 exhibited the capability to activate p38 mitogen-activated protein kinase (MAPK） even in the absence of IL-4 stimulation. Moreover, we observed that a p38 MAPK inhibitor suppressed the BRD6989-induced increase in arginase-1 expression. Besides, BRD6989 increased the surface expression of CD206, an M2 macrophage marker. Thus, this study demonstrated for the first time that CDK8/19 inhibition increases arginase-1 expression, suggesting that this mechanism involves the activation of STAT6 and p38 MAPK. This finding implies that CDK8/19 inhibition may facilitate the production of anti-inflammatory M2 macrophages.

Synthetic Approaches and Clinical Application of Representative Small-Molecule Inhibitors of Cyclin-Dependent Kinase for Cancer Therapy.

The regulation of the cancer cell cycle heavily relies on cyclin-dependent kinases (CDKs). Targeting CDKs has been identified as a promising approach for effective cancer therapy. In recent years, there has been significant attention paid towards developing small-molecule CDK inhibitors in the field of drug discovery. Notably, five such inhibitors have already received regulatory approval for the treatment of different cancers, including breast tumors, lung malignancies, and hematological malignancies. This review provides an overview of the synthetic routes used to produce 17 representative small-molecule CDK inhibitors that have obtained regulatory approval or are currently being evaluated through clinical trials. It also discusses their clinical applications for treating CDK-related diseases and explores the challenges and limitations associated with their use in a clinical setting, which will stimulate the further development of novel CDK inhibitors. By integrating therapeutic applications, synthetic methodologies, and mechanisms of action observed in various clinical trials involving these CDK inhibitors, this review facilitates a comprehensive understanding of the versatile roles and therapeutic potential offered by interventions targeting CDKs.

Cimicifugin, a broad-spectrum inhibitor of Theileria annulata and Plasmodium falciparum CDK7.

Cyclin-dependent kinase 7 is an attractive therapeutic target for the treatment of cancers, and a previous report suggested that Plasmodium falciparum CDK7 is a potential drug target for developing new anti-malarial drugs. In this study, we aimed to characterize and evaluate the drug target potential of Theileria annulata CDK7. Theileria annulata is responsible for tropical theileriosis, which induces a phenotype similar to cancerous cells like immortalization, hyperproliferation, and dissemination. Virtual screening of the MyriaScreen II library predicted 14 compounds with high binding energies to the ATP-binding pocket of TaCDK7. Three compounds (cimicifugin, ST092793, and ST026925) of these 14 compounds were non-cytotoxic to the uninfected bovine cells (BoMac cells). Cimicifugin treatment led to the activation of the extrinsic apoptosis pathway and induced autophagy in T. annulata-infected cells. Furthermore, cimicifugin also inhibited the growth of P. falciparum, indicating that it has both anti-theilerial and anti-malarial activities and that TaCDK7 and PfCDK7 are promising drug targets.

CDKL3 is a promising biomarker for diagnosis and prognosis prediction in patients with hepatocellular carcinoma.

Cyclin-dependent kinase-like 3 (CDKL3) has been identified as an oncogene in certain types of tumors. Nonetheless, its function in hepatocellular carcinoma (HCC) is poorly understood. In this study, we conducted a comprehensive analysis of CDKL3 based on data from the HCC cohort of The Cancer Genome Atlas (TCGA). Our analysis included gene expression, diagnosis, prognosis, functional enrichment, tumor microenvironment and metabolic characteristics, tumor burden, mRNA expression-based stemness, alternative splicing, and prediction of therapy response. Additionally, we performed a cell counting kit-8 assay, TdT-mediated dUTP nick-end Labeling staining, migration assay, wound healing assay, colony formation assay, and nude mouse experiments to confirm the functional relevance of CDKL3 in HCC. Our findings showed that CDKL3 was significantly upregulated in HCC patients compared to controls. Various bioinformatic analyses suggested that CDKL3 could serve as a potential marker for HCC diagnosis and prognosis. Furthermore, CDKL3 was found to be involved in various mechanisms linked to the development of HCC, including copy number variation, tumor burden, genomic heterogeneity, cancer stemness, and alternative splicing of CDKL3. Notably, CDKL3 was also closely correlated with tumor immune cell infiltration and the expression of immune checkpoint markers. Additionally, CDKL3 was shown to independently function as a risk predictor for overall survival in HCC patients by multivariate Cox regression analysis. Furthermore, the knockdown of CDKL3 significantly inhibited cell proliferation in vitro and in vivo, indicating its role as an oncogene in HCC. Taken together, our findings suggest that CDKL3 shows promise as a biomarker for the detection and treatment outcome prediction of HCC patients.