Study on the Biological Function of Abemecilib in Inhibiting the Proliferation, Invasion and Migration of Small Cell Lung Cancer with High c-Myc Expression / 中国肺癌杂志

BACKGROUND@#Small cell lung cancer (SCLC) with high c-Myc expression is prone to relapse and metastasis, leading to extremely low survival rate. Cyclin-dependent kinases 4 and 6 (CDK4/6) inhibitor Abemaciclib plays a key role in the treatment of tumors, but the effects and mechanisms on SCLC remain unclear. This study was to analyze the effect and molecular mechanism of Abemaciclib in inhibiting proliferation, migration and invasion of SCLC with high c-Myc expression, with a view to expanding a new direction for reducing the recurrence and metastasis.@*METHODS@#Proteins interacting with CDK4/6 were predicted using the STRING database. The expressions of CDK4/6 and c-Myc in 31 cases of SCLC cancer tissues and paired adjacent normal tissues were analyzed by immunohistochemistry. The effects of Abemaciclib on the proliferation, invasion and migration of SCLC were detected by CCK-8, colony formation assay, Transwell and migration assay. Western blot was used to detect the expressions of CDK4/6 and related transcription factors. Flow cytometry was used to analyze the effects of Abemaciclib on the cell cycle and checkpoint of SCLC.@*RESULTS@#The expression of CDK4/6 was associated with c-Myc by STRING protein interaction network. c-Myc can directly modalize achaete-scute complex homolog 1 (ASCL1), neuronal differentiation 1 (NEUROD1) and Yes-associated protein 1 (YAP1). Moreover, CDK4 and c-Myc regulate the expression of programmed cell death ligand 1 (PD-L1). Immunohistochemistry showed that the expressions of CDK4/6 and c-Myc in cancer tissues were higher than those in adjacent tissues(P<0.0001). CCK-8, colony formation assay, Transwell and migration assay verified that Abemaciclib could effectively inhibit the proliferation, invasion and migration of SBC-2 and H446OE(P<0.0001). Western blot analysis further showed that Abemaciclib not only inhibited CDK4 (P<0.05) and CDK6 (P<0.05), but also affected c-Myc (P<0.05), ASCL1 (P<0.05), NEUROD1 (P<0.05) and YAP1 (P<0.05), which are related to SCLC invasion and metastasis. Flow cytometry showed that Abemaciclib not only inhibited the cell cycle progression of SCLC cells (P<0.0001), but also significantly increased PD-L1 expression on SBC-2 (P<0.01) and H446OE (P<0.001).@*CONCLUSIONS@#Abemaciclib significantly inhibits the proliferation, invasion, migration and cell cycle progression of SCLC by inhibiting the expressions of CDK4/6, c-Myc, ASCL1, YAP1 and NEUROD1. Abemaciclib can also increase the expression of PD-L1 in SCLC.

Recent Progress of Palbociclib in Treatment of HR+/HER2- Breast Cancer / 肿瘤防治研究

As the first cyclin-dependent kinases 4 and 6 inhibitors, palbociclib significantly improved the survival of the patients with the hormone receptor-positive and human epidermal growth factor receptor-2 negative breast cancer. Palbociclib is a crucial landmark in the development history of antineoplastic drugs. This article reviews the mechanism of palbociclib, and summarizes the clinical trials, side effects, and the application of palbociclib.

Recent progress of targeted small molecular CDK9 degraders based on PROTAC technology / 药学学报

CDKs proteins are a kind of cell cycle protein-dependent kinases, which serve as important roles in controlling cell division and transcriptional stages. Among them, CDK9, as a key regulator responsible for the transcriptional elongation of cells, drives the development of various malignant cells and is considered as an important target in the field of anti-tumor drug development. However, the CDK family proteins feature high conservativeness and similarity in structure, leading to the poor selectivity and severe side effects for traditional small-molecular CDK9 inhibitors, which has limited their clinical applications. In view of this, there is an urgent need to investigate CDK9 targets through a novel strategy. The PROTAC is an emerging drug discovery strategy that the degrader could specifically recognize the target protein through indirect linkage with ubiquitin ligases and ultimately eliminate the target protein through the ubiquitination degradation system. This paper provides a brief overview of the structure and function of CDK9 protein, its relationship with the poor prognosis of clinical diseases, as well as the currently reported small molecular inhibitors. The latest research progress on the targeted degradation of CDK9 protein based on PROTAC technology is highlighted. Finally, the development prospects of this target protein in this novel technology field are summarized and prospected, aiming to provide a reference for the development of antitumor drugs in this direction.

Peroxidase from foxtail millet bran exerts anti-colorectal cancer activity via targeting cell-surface GRP78 to inactivate STAT3 pathway

Molecular targeted therapy has become an emerging promising strategy in cancer treatment, and screening the agents targeting at cancer cell specific targets is very desirable for cancer treatment. Our previous study firstly found that a secretory peroxidase of class III derived from foxtail millet bran (FMBP) exhibited excellent targeting anti-colorectal cancer (CRC) activity in vivo and in vitro, whereas its underlying target remains unclear. The highlight of present study focuses on the finding that cell surface glucose-regulated protein 78 (csGRP78) abnormally located on CRC is positively correlated with the anti-CRC effects of FMBP, indicating it serves as a potential target of FMBP against CRC. Further, we demonstrated that the combination of FMBP with the nucleotide binding domain (NBD) of csGRP78 interfered with the downstream activation of signal transducer and activator of transcription 3 (STAT3) in CRC cells, thus promoting the intracellular accumulation of reactive oxygen species (ROS) and cell grown inhibition. These phenomena were further confirmed in nude mice tumor model. Collectively, our study highlights csGRP78 acts as an underlying target of FMBP against CRC, uncovering the clinical potential of FMBP as a targeted agent for CRC in the future.

Selective inhibition of CDK4/6: A safe and effective strategy for developing anticancer drugs

The sustained cell proliferation resulting from dysregulation of the cell cycle and activation of cyclin-dependent kinases (CDKs) is a hallmark of cancer. The inhibition of CDKs is a highly promising and attractive strategy for the development of anticancer drugs. In particular, third-generation CDK inhibitors can selectively inhibit CDK4/6 and regulate the cell cycle by suppressing the G1 to S phase transition, exhibiting a perfect balance between anticancer efficacy and general toxicity. To date, three selective CDK4/6 inhibitors have received approval from the U.S. Food and Drug Administration (FDA), and 15 CDK4/6 inhibitors are in clinical trials for the treatment of cancers. In this perspective, we discuss the crucial roles of CDK4/6 in regulating the cell cycle and cancer cells, analyze the rationale for selectively inhibiting CDK4/6 for cancer treatment, review the latest advances in highly selective CDK4/6 inhibitors with different chemical scaffolds, explain the mechanisms associated with CDK4/6 inhibitor resistance and describe solutions to overcome this issue, and briefly introduce proteolysis targeting chimera (PROTAC), a new and revolutionary technique used to degrade CDK4/6.

Regulation of Plasmodium falciparum cell cycle involving Cyclins and Cyclin Dependent Kinases

@#Protein kinases (PKs) are regulators of protein phosphorylation in many infectious diseases, including malaria. However, the cellular functions of majority of PKs in Plasmodium falciparum remain unknown. The mechanisms involved in P. falciparum cell cycle progress are not fully understood. The activation of cyclin-dependent kinases (CDKs), which constitute a PK family that includes crucial regulators of cell cycle progression in eukaryotes, is strictly being coordinated by the interaction with specific cyclins at well-defined points within the cell cycle. These cyclin/CDK complexes are very well characterised in humans, but little is known in P. falciparum. This review expand our understanding of the characteristic of CDKs and cyclins in P. falciparum, and paves the way for further investigations on the precise molecular role of these crucial regulatory proteins in mosquito and human. This represents a valuable step towards the elucidation of cell cycle control mechanisms in malaria parasites.

Application of CDK4 / 6 inhibitors in the treatment of hormone receptor-positive breast cancer / 国际肿瘤学杂志

With roles of blocking cell proliferation,cyclin-dependent protein kinase (CDK)4 / 6 inhi-bitors are effective and safe complementary therapies for hormone receptor-positive breast cancer. CDK4 / 6 inhibitors combined with endocrine therapy significantly improve the prognosis of patients with advanced or metastatic breast cancer,especially those with endocrine resistance. Indications for CDK4 / 6 inhibitors are also expected to broaden into early-stage breast cancer. However,mechanisms of resistance of CDK4 / 6 inhibitors remain elusive.

Abnormal Mitochondria in a Non-human Primate Model of MPTP-induced Parkinson's Disease: Drp1 and CDK5/p25 Signaling

Mitochondria continuously fuse and divide to maintain homeostasis. An impairment in the balance between the fusion and fission processes can trigger mitochondrial dysfunction. Accumulating evidence suggests that mitochondrial dysfunction is related to neurodegenerative diseases such as Parkinson's disease (PD), with excessive mitochondrial fission in dopaminergic neurons being one of the pathological mechanisms of PD. Here, we investigated the balance between mitochondrial fusion and fission in the substantia nigra of a non-human primate model of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD. We found that MPTP induced shorter and abnormally distributed mitochondria. This phenomenon was accompanied by the activation of dynamin-related protein 1 (Drp1), a mitochondrial fission protein, through increased phosphorylation at S616. Thereafter, we assessed for activation of the components of the cyclin-dependent kinase 5 (CDK5) and extracellular signal-regulated kinase (ERK) signaling cascades, which are known regulators of Drp1(S616) phosphorylation. MPTP induced an increase in p25 and p35, which are required for CDK5 activation. Together, these findings suggest that the phosphorylation of Drp1(S616) by CDK5 is involved in mitochondrial fission in the substantia nigra of a non-human primate model of MPTP-induced PD.

Application of CDK4/6 inhibitors in the treatment of hormone receptor-positive breast cancer / 国际肿瘤学杂志

With roles of blocking cell proliferation, cyclin-dependent protein kinase (CDK) 4/6 inhi-bitors are effective and safe complementary therapies for hormone receptor-positive breast cancer. CDK4/6 inhibitors combined with endocrine therapy significantly improve the prognosis of patients with advanced or metastatic breast cancer, especially those with endocrine resistance. Indications for CDK4/6 inhibitors are also expected to broaden into early-stage breast cancer. However, mechanisms of resistance of CDK4/6 inhibitors remain elusive.

MNAT1 expression in non-small cell lung cancer and its biological cellular impact / 中华病理学杂志

Objective@#To investigate the expression and significance of MNAT1 in non-small cell lung cancer (NSCLC) and to explore the biological impact of MNAT1 expression in lung cancer cells at the cellular level and related signaling pathway.@*Methods@#Forty-eight cases of NSCLC tissues and paired normal tissues was collected at Nanfang Hospital, Southern Medical University from 2015 to 2017. The expression level of MNAT1 was detected by immunohistochemistry, and the relationship between MNAT1 and clinicopathological features was analyzed. The expression of MNAT1 was detected in lung cancer cells, MNAT1 level was analyzed after knocking down in A549 and H322 cells by siRNA; Plasmid vector of overexpressing MNAT1 was constructed, followed by transfecting H1299 cells and observing proliferation and migration at the cellular level. Flow cytometry was used to analyze the effect of the expression of MNAT1 on cell cycle, and Western blot was used to explore the possible molecular mechanism of MNAT1 on cell proliferation and cell cycle.@*Results@#Immunohistochemistry showed that the expression score of MNAT1 was (4.07±3.55) in normal lung tissue and (7.33±4.09) in NSCLC tissue (P<0.01), and correlated with lymph node metastasis. At the cellular level, MNAT1 promoted cell proliferation(P<0.05), migration(P<0.05) and cell cycle progression(P<0.01).@*Conclusions@#MNAT1 may be involved in the development of non-small cell lung cancer.MNAT1 affects cell cycle and proliferation through the Akt/p21 pathway.

Application of selective CDK4/6 inhibitors in solid cancers therapy / 国际肿瘤学杂志

Cyclin-dependent kinase (CDK) inhibitors have been approved for the treatment of ER +/HER2-advanced breast cancer,and preclinical study and clinical trials are under way for multiple solid cancers.CDK4/6 inhibitors have shown good efficacy for human solid cancers,and it's future development direction are biomarkers,drug resistance and combined therapy in the future.

A study on mechanism underlying the inhibition of berberine on proliferation of human melanoma A375 cells / 中华皮肤科杂志

Objective To evaluate the effect of berberine on the expression of cell cycle-related miRNA and its target gene in human melanoma A375 cells.Methods In vitro cultured A375 cells were classified into several groups to be treated with berberine at different concentrations of 0 (blank control group),20,40,60,80,and 100 μmol/L,respectively,for 48 hours.qRT-PCR was performed to determine the mRNA expression of miRNA-582-5p and its target gene cyclin-dependent kinase 1 (CDK1),miRNA-188-5p and its target gene CDK2,Cyclin D1 and Cyclin A.Western blot analysis was conducted to measure the protein expression of cell cycle-related proteins CDK1,CDK2,Cyclin D1 and Cyclin A.Results qRT-PCR showed that compared with the blank control group,the 20 and 40 μmol/L berberine groups had a similar expression of miRNA-582-5p (both P ＞ 0.05),but the 60 and 80 μmol/L berberine groups had a significantly up-regulated expression of miRNA-582-5p (both P ＜ 0.05).Compared with the blank control group,all the 5 berberine groups had a significantly increased expression of miRNA-188-5p (F =22.600,P =0.002).However,the mRNA expression of CDK2,CDK1,Cyclin A and Cyclin D1 gradually decreased along with the increase of berberine concentrations (F =51.976,248.510,626.671 and 312.740,respectively,all P ＜ 0.001).Western blot analysis revealed that berberine decreased the protein expression of CDK1,CDK2,Cyclin D1 and Cyclin A (F =138.124,110.966,278.772 and 140.167,all P ＜ 0.001).Conclusion Berberine can decrease the expression of cell cycle-related proteins CDK1,CDK2,Cyclin D1 and Cyclin A,likely by decreasing their mRNA expression and increasing the expression of miRNA-582-5p and miRNA-188-5p,and then block the cell cycle progression of A375 cells and inhibit the growth of tumor.

Progress in the relevance of CDK12 with tumors / 肿瘤

Cancer represents a pathological characteristic of uncontrolled cell division. The cyclin-dependent kinases (CDKs) are a kinds of key factors regulating cell cycle progression and transcription process. Since the dysregulation of CDKs is a frequently occurring event driving tumorigenesis, CDKs have been tested extensively as the targets for cancer therapy. CDK12 is a transcription-associated kinase which participates in various cellular processes, including DNA damage response, cellular development and differentiation, as well as precursor mRNA splicing and processing. The mutations and amplification of CDK12 gene have been recently reported in different types of malignancies, such as loss-of-function mutations in high-grade serous ovarian carcinomas, which suggests that CDK12 is a tumor suppressor. On the contrary, CDK12 is overexpressed in other tumors, which suggests that CDK12 has some properties of oncogene. This paper reviews the structure and function of CDK12 gene, as well as the relationship between CDK12 gene and cancers.

The role of p21/CIP1/WAF1 (p21) in the negative regulation of the growth hormone/growth hormone receptor and epidermal growth factor/epidermal growth factor receptor pathways, in growth hormone transduction defect

PURPOSE: Growth hormone transduction defect (GHTD) is characterized by severe short stature, impaired STAT3 (signal transducer and activator of transcription-3) phosphorylation and overexpression of the cytokine inducible SH2 containing protein (CIS) and p21/CIP1/WAF1. To investigate the role of p21/CIP1/WAF1 in the negative regulation of the growth hormone (GH)/GH receptor and Epidermal Growth Factor (EGF)/EGF Receptor pathways in GHTD. METHODS: Fibroblast cultures were developed from gingival biopsies of 1 GHTD patient and 1 control. The protein expression and the cellular localization of p21/CIP1/WAF1 was studied by Western immunoblotting and immunofluorescence, respectively: at the basal state and after induction with 200-μg/L human GH (hGH) (GH200), either with or without siRNA CIS (siCIS); at the basal state and after inductions with 200-μg/L hGH (GH200), 1,000-μg/L hGH (GH1000) or 50-ng/mL EGF. RESULTS: After GH200/siCIS, the protein expression and nuclear localization of p21 were reduced in the patient. After successful induction of GH signaling (control, GH200; patient, GH1000), the protein expression and nuclear localization of p21 were reduced. After induction with EGF, p21 translocated to the cytoplasm in the control, whereas in the GHTD patient it remained located in the nucleus. CONCLUSIONS: In the GHTD fibroblasts, when CIS is reduced, either after siCIS or after a higher dose of hGH (GH1000), p21’s antiproliferative effect (nuclear localization) is also reduced and GH signaling is activated. There also appears to be a positive relationship between the 2 inhibitors of GH signaling, CIS and p21. Finally, in GHTD, p21 seems to participate in the regulation of both the GH and EGF/EGFR pathways, depending upon its cellular location.

Cyclin-dependent kinase 7 gene silencing or knockout represses proliferation of ovarian cancer cells and the mechanism research / 肿瘤

Objective: To investigate the role of cyclin-dependent kinase 7 (CDK7) in the proliferation of ovarian cancer cells. Methods: The ovarian cancer cell lines OVCA433, TOV-112D and IGROVl were transfected with specific siRNA to downregulate the expression of CDK7 gene, then the cell proliferation viability was detected by CCK-8 method. The CDK7 gene in ovarian cancer HEY, OVCA420, OVCA433 and IGROVl cells was knocked out by clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated endonuclease 9 (Cas9) system, then the clone formation ability was detected by plate clone formation assay. The ovarian cancer TOV-112D, IGROV1, OVCA433, OVCAR8, OV90, SKOV3 and COV413B cells were treated with CDK7-inhibitor THZl, then the cell proliferation viability and clone formation ability were detected by CCK-8 method and clone formation assay, respectively. Furthermore, the expression level of CDK7 protein and the phosphorylation level of RNA polymerase n (RNAPolE) in IGROV1, OVCA433, SKOV3 and COV41 3B cells treated with THZl were analyzed by Western blotting. Results: The proliferation and clone formation abilities of various ovarian cancer cells were significantly decreased after CDK7 gene was silenced or knocked out (all P < 0.05). THZ1 repressed the proliferation and clon e formation of ovarian cancer cells, an d downregulated the expression of CDK7 and the phosphorylation of RNAPoln (all P < 0.05). Conclusion: CDK7 may promote the proliferation of ovarian cancer cells by regulating the phosphorylation of RNAPoln.

The role of PD0332991 in clonogenicity of mantle cell lymphoma cells / 中国肿瘤临床

Objective:To investigate the function and mechanism of cyclin-dependent kinase 4/6 (CDK4/6) inhibitor PD0332991 on clo-nogenicity of mantle cell lymphoma (MCL) cells. Methods:The effect of PD0332991 on MCL cell cycle distribution was assessed by flow cytometry;Western blot was used to test expression level of Rb protein and phosphorylated Rb protein in MCL after treatment with PD0332991;colony forming assay was performed to test the role of PD0332991 and mitoxantrone and their combination on colo-ny forming activity in MCL. Results:Flow cytometry revealed that PD0332991 can increase G0/G1 phase MCL cells and significantly de-crease S phase cells, leading to G0/G1 cell arrest. Western blot confirmed that PD0332991 exerted no effect on Rb protein expression but suppressed levels of phosphorylated Rb protein. Colony forming assay showed that PD0332991 significantly suppressed colony for-mation and enhanced the effect of mitoxantrone on colony forming activity in MCL. Conclusion:This study revealed that CDK4/6 inhib-itor PD0332991 induced G0/G1 cell arrest and increased the effect of mitoxantrone on MCL clonogenicity by suppressing levels of phosphorylated Rb protein.

Multiple cutaneous melanomas associated with gastric and brain metastases

Abstract The occurrence of multiple primary melanomas in a single individual is rare. Most commonly, malignant melanocytic lesions subsequent to the initial diagnosis of melanoma are secondary cutaneous metastases. We report a patient with gastrointestinal bleeding from gastric metastasis of cutaneous melanoma. During clinical evaluation and staging, we discovered a brain metastasis associated with 3 synchronous primary cutaneous melanomas. We suggest the research on the mutation in the cyclin-dependent kinase inhibitor 2A (CDKN2A) (INK4a) in such cases. We also emphasize the importance of clinical examination and dermoscopy of the entire tegument, even after a malignant melanocytic lesion is identified.

Expression of CCRK in small cell lung cancer and its clinical significance / 中国医师杂志

Objective To investigate the pathogenesis of small cell lung cancer ( SCLC) and to ex-plore the expression of cell cycle related kinase ( CCRK) in SCLC and its clinical significance.Methods Reverse transcription polymerase chain reaction ( RT-PCR) and Western blot were performed to examine ex-pression of CCRK in SCLC and normal tissues.Results The expressions of gene [(0.51 ±0.11)IU/L] and protein [(0.61 ±0.13)IU/L] of CCRK in SCLC tissues were significantly higher than normal tissues [(0.30 ±0.08)IU/L, (0.34 ±0.09)IU/L] ( P 0.05 ) .Conclusions The expressions of gene and protein of CCRK in SCLC tissues were significantly higher than normal tissues. CCRK promoted the occurrence and progress of SCLC.Chem can restrain effectually the excessive expres-sion of CCRK.The expressions of gene and protein of CCRK in the different clinical curative effect group had significant difference.

Verification of Differential Expression Genes after CacyBP/SIP Nuclear Translocation in Colon Carcinoma Cell Line / 天津医药

Objective To verify the genes screened by the polymerase chain reaction (PCR) chip of cell cycle. Methods The colon cancer cells SW480 were randomized into two groups, the test group (with gastrin stimulation) and con-trol group (without gastrin stimulation). The method of Western blot was used to detect the expression of calcylin binding pro-tein/Siah-1 interacting protein (Cacybp/SIP) before and after gastrin stimulation. The differential expression genes, cyclin de-pendent kinase 8 (CDK8) and cyclin dependent kinase subunit (CKS2), were verified by using real-time quantitative PCR (qRT-PCR). Results It was found that before the stimulation, CacyBP/SIP was located and expressed in cytoplasm, and then in both cytoplasm and nucleus after gastrin stimulation. The qRT-PCR results of CDK8 and CKS2 genes were consis-tent with those of microarray detection. The expressions of CDK8 and CKS2 were up-regulated (P < 0.05). Conclusion The stimulation of human gastrin can lead to the nuclear translocation of CacyBP/SIP. The results of microarray are reliable, and the differentially expressed genes screened through gene chip deserve further study.

The importance of regulatory role of panaxoside Rg1 in Cdk5 on hippocampal neuron radioactive damage protection / 中国医师杂志

Objective To discuss the importance of regulatory role of panaxoside Rg1 in Cdk5 in the process of hippocampal neuron radioactive damage protection.Methods Radioactive brain damage in vivo 40 models were built,and divided into 4 groups,including 0 Gy group (short for blank group),pure panaxoside Rg1 preconditioning group (short for control group),30 Gy group (short for model group),and 30Gy + panaxoside Rg1 preconditioning group (short for traditional Chinese medicine group).Hippocampal neurons were separated and trained.Hippocampal neuron apoptosis condition was tested in every group by 4′,6-diamidino2-phenylindole (DAPI) staining method.The p35 and p25 protein expressions were tested with Western blot.Cdk5 was restrained by Cdk5 restrainer roscovitine.Hippocampal neuron damage after Cdk5 blocking-up was observed with changes of X ray in every group.Results Compared with blank group,no significant difference was found in nuclear shrinkage percentage,the number of neuron survival,and the protein expression levels of p35 and p25 in control group ; nuclear shrinkage percentage and the protein expression levels of p35 and p25 were significantly increased and the number of neuron survival was significantly decreased in the model group and traditional Chinese medicine group (P ＜ 0.05).Compared with model group,nuclear shrinkage percentage and the protein expression levels of p35 and p25 were significantly decreased and the number of neuron survival was significantly increased in the traditional Chinese medicine group (P ＜ 0.05).For the addition of dimethyl sulfoxide (DMSO) in every group,the nuclear shrinkage percentage was not significantly changed in control group compared with blank group,was significantly increased in model group and traditional Chinese medicine group compared with blank group (P ＜ 0.05),and was significantly decreased in traditional Chinese medicine group compared with model group (P ＜ 0.05).Conclusions Panaxoside Rg1 can reduce neuron apoptosis by controlling Cdk5,and plays a protective role in hippocampal neuron radioactive damage.