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Cyclin-dependent kinase 6 (EC 2.7.11.22) play significant roles in numerous biological processes and triggers cell cycle events. CDK6 controlled the transcriptional regulation. A dysregulated function of CDK6 is linked with the development of progression of multiple tumor types. Thus, it is considered as an effective drug target for cancer therapy. Based on the direct roles of CDK4/6 in tumor development, numerous inhibitors developed as promising anti-cancer agents. CDK4/6 inhibitors regulate the G1 to S transition by preventing Rb phosphorylation and E2F liberation, showing potent anti-cancer activity in several tumors, including HR+/HER2- breast cancer. CDK4/6 inhibitors such as abemaciclib, palbociclib, and ribociclib, control cell cycle, provoke cell senescence, and induces tumor cell disturbance in pre-clinical studies. Here, we discuss the roles of CDK6 in cancer along with the present status of CDK4/6 inhibitors in cancer therapy. We further discussed, how structural features of CDK4/6 could be implicated in the design and development of potential anti-cancer agents. In addition, the therapeutic potential and limitations of available CDK4/6 inhibitors are described in detail. Recent pre-clinical and clinical information for CDK4/6 inhibitors are highlighted. In addition, combination of CDK4/6 inhibitors with other drugs for the therapeutic management of cancer are discussed.

Background CDK4/6 inhibitors have been used to treat hormone receptor-positive HER2-negative advanced breast cancer. Their benefit in HER2-positive breast cancer has not been determined yet. In this study, we investigated the effects of HER2 on CDK4/6 activity by assessing the level of downstream phosphorylated retinoblastoma protein (pRb) in HER2-positive breast cancer (HER2 positivity is defined by immunohistochemical study or FISH, regardless of ER status) to determine if these cases may be responsive to CDK4/6 inhibitors. Materials and Methods One hundred and thirty cases of breast biopsies with invasive carcinoma were collected, including 77 cases of HER2+ (39 cases of ER +PR±HER2+ and 38 cases of ER-PR-HER2+) and 53 cases of HER2- (ER-PR-HER2-) breast cancer. Immunohistochemical study of pRb was performed and the pRb level was assessed by H-score (intensity x percentage of positive cells). Results The pRb H-score ranges from 3 to 270. The average H-scores for the ER-PR-HER2+, ER+PR±HER2+ and ER-PR-HER2- groups are 115.8 ± 75.8, 93.1 ± 68.6 and 63.1 ± 65.6, respectively. By comparison, HER2+ cases have significantly higher pRb levels than HER2- cases (P = .001). Among HER2+ cases, there was a trend of positive correlation between the HER2 gene copy number, and the pRb level although not statistically significant (r = 0.192, 95% CI, [-0.033, 0.399], P = .09). Conclusion In breast cancer, HER2 positivity leads to significantly higher levels of CDK4/6 activity as reflexed by pRb. Breast cancer that is positive for HER2 may respond to CDK4/6 inhibitors and pRb may potentially be used as a biomarker to predict the responsiveness.

Acute lymphoblastic leukemia (ALL) is a common malignancy in children. In this study, we aimed to explore putative mechanisms of microRNA-155-5p (miR-155-5p) involvement in childhood ALL (cALL) via interactions with casitas B-lineage lymphoma (CBL), interferon regulatory factor 4 (IRF4), and cyclin-dependent kinase 6 (CDK6). Bioinformatic analysis was performed initially to identify differentially expressed genes in cALL. The expression levels of miR-155-5p, CBL, IRF4, and CDK6 in peripheral blood lymphocytes from clinical ALL samples were determined using RT-qPCR and Western blot assays. A dual-luciferase reporter gene assay was used to ascertain a possible targeting relationship between miR-155-5p and CBL, CCK-8 assay and flow cytometry were used to measure cell activity and apoptosis of ALL cells. Co-IP was performed to investigate the interaction between CBL and IRF4 and the ubiquitination level of IRF4. Furthermore, in vivo validation was performed inducing xenograft tumor models with ALL cells in nude mice. As indicated by bioinformatic analysis, miR-155-5p and CDK6 were upregulated and CBL was downregulated in ALL. miR-155-5p was found to target CBL to inhibit CBL expression. miR-155-5p promoted the proliferation of ALL cells and inhibited their apoptosis by inhibiting the expression of CBL, which otherwise degraded IRF4 protein through ubiquitination, leading to inhibited CDK6 expression. Collectively, the results show that miR-155-5p can promote the development of cALL via the regulation on CBL-mediated IRF4/CDK6 axis. The authors explored the role of microRNA-155-5p (miR-155-5p) in childhood acute lymphoblastic leukemia (cALL). They found that miRNA-155-5p targets the E3 ubiquitin ligase CBL and inhibits its expression, reducing the ubiquitination and degradation of interferon regulatory factor 4, thus elevating expression of cyclin-dependent kinase 6. These findings suggest a basis for potential future therapeutic strategies for cALL.

Introduction Cyclin-dependent kinase (CDK)4/6 inhibitor is a first-line therapy for metastatic ER+/HER2-breast cancer. However, there are limited data on safety of combined radiotherapy (RT) and CDK4/6 inhibition. Methods We conducted a retrospective study of women with metastatic breast cancer who received palliative RT within 14 days of CDK4/6 inhibitor use. The primary endpoint was toxicity per Common Terminology Criteria for Adverse Events v5. Secondary endpoints were pain response and local control based on clinical assessment and imaging. Results Thirty patients underwent 36 RT courses with palbociclib (n = 34 courses, 94.4%) or abemaciclib (n = 2, 5.6%). RT was delivered before, concurrently or after CDK4/6 inhibitors in 7 (19.4%), 8 (22.2%), and 21 (58.3%) of cases with median 3.5 days from RT to closest CDK4/6 inhibitor administration. Median RT dose was 30Gy (range 8–40.05Gy). Treated sites included brain (n = 5, 11.6%), spine (n = 19, 44.2%), pelvis (n = 9, 20.9%), other bony sites (n = 6, 14.0%) and others (n = 4, 9.3%). No acute grade ≥3 non-hematologic toxicity occurred. No increased hematologic toxicity was attributable to RT with grade 3 hematologic toxicities rates 16.7%, 0%, and 6.7% before, during, and 2 weeks after RT completion. All but one patient (29/30) achieved symptom relief. Local control rates were 94.4%, 91.7% at 6 and 12 months. Conclusions The use of RT within 2 weeks of CDK4/6 inhibitors had low acceptable toxicity and high efficacy, suggesting that it is safe for palliation of metastatic breast cancer., Highlights • CDK4/6 inhibitors with endocrine therapy is a preferred first line therapy for HR+, HER2-metastatic breast cancer. • Limited data exists on safety of combined radiotherapy and CDK4/6 inhibition in patients with metastatic breast cancer. • This retrospective cohort study included 30 patients who underwent palliative RT with combined CDK4/6 inhibitor use. • RT within 2 weeks of CDK4/6 inhibitor had low toxicity and high efficacy, supporting its safety in metastatic breast cancer.

Background Regulatory B cells (Bregs) play an important role in maintaining immune homeostasis and have the potential to induce tolerance. Previous work has found that Breg cells are involved in heart transplantation tolerance. However, the effect of Breg on the transplantation tolerance and the underlying mechanisms remain to be clarified. Methods Using a within-species heart transplantation model, we aimed to investigate the role of CD19+CD5+CD1dhigh Bregs isolated from transplanted mice in preventing transplant rejection in vivo. We also explored the effects of CD40 and tumor necrosis factor receptor-associated factor 6 (TRAF6) ubiquitin ligase on Breg-mediated prolongation of survival in heart transplant (HT) mice, and the regulatory effects of downstream Cdk4 and Cdk6 proteins on dendritic cells (DCs), which clarified the function and molecular mechanism of Breg cells in HT mice. Results Our data suggest that adoptive transfer of the transplanted Bregs served as an effective tolerance-inducing mechanism in HT mice and was involved in the CD40-TRAF6 signaling pathway in DCs. Moreover, DCs collected from the Breg treated HT mice also prolonged the survival of HT mice. Furthermore, DC-specific knockout of TRAF6 diminished Breg-mediated prolongation of survival in HT mice. Interestingly, gut microbes from donors increased the survival of cardiac allografts both in both the absence and presence of Bregs but were not implicated in CD40-TRAF6 signaling. Conclusions These findings reveal a role of Breg cells in the induction of transplantation tolerance through the blockade of the CD40-TRAF6 signaling pathway, which might be used in the treatment of HT in the clinic.

Undoubtedly, the development of COVID-19 vaccines displays a critical step towards ending this devastating pandemic, considering their protective benefits in the general population. Yet, data regarding their efficacy and safety in cancer patients are limited. Herein we provide the initial analysis of immune responses after the first dose of vaccination in 21 breast cancer patients receiving cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors. The levels of neutralizing antibodies post vaccination were similar to the matched healthy controls, whereas no safety issues have been raised. Further exploration is needed to reduce the uncertainty of SARS-CoV-2 immunity among cancer patients under treatment.

Recurrent gene mutations and copy number alterations in cancer patients are presumably associated with resistance to targeted therapy. In the present study, we assessed the gene mutations and copy number alterations that recurrently occurred in cetuximab-treated patients with metastatic colorectal cancer (mCRC). Targeted next-generation sequencing was performed in the tumor samples obtained pre- and postcetuximab treatment to assess the variations that occurred during cetuximab treatment. Moreover, we identified the emergent gene mutations (CDK6, EPHA3, ERCC2, MYC, PCMTD1, PIK3CA, PRIM2, RICTOR, and ZNRF3) and copy number alterations (ARAF, BCL2, BRCA2, EGFR, MYC, and SMAD4) that were recurrently observed only in postprogression samples and not in pretreatment or posttreatment samples from patients revealing clinical response. Furthermore, to identify the feasible candidate variations implicated in treatment resistance, we examined the variants with clonal expansion during treatment and discovered PCBP1 as a variant associated with posttreatment progression. Various recurrent mutations were enriched in the TGF-beta signaling pathway. Collectively, we identified recurrent variations in mCRC samples exhibiting post-cetuximab progression. Additionally, future studies are required to evaluate the therapeutic potential of these variations.

Summary Background Cyclin-dependent kinase 4/6 inhibitors (CDKIs) are oral targeted agents approved for use in combination with endocrine therapy as first-line or second-line treatment of patients with hormone receptor-positive, HER2-negative, advanced or metastatic breast cancer. We previously reported the pooled analyses of progression-free survival in patients in specific clinicopathological subgroups, all of whom received consistent benefit from the addition of a CDKI to hormonal therapy. Here, we report the pooled overall survival results in patients treated with a CDKI and fulvestrant. Methods In this exploratory analysis, we pooled individual patient data from three phase 3 randomised trials of CDKI or placebo in combination with fulvestrant in patients with breast cancer submitted to the US Food and Drug Administration and approved before Aug 1, 2020, in support of marketing applications. All analysed patients were aged at least 18 years, had an Eastern Cooperative Oncology Group performance status of 0–1, had hormone receptor-positive, HER2-negative advanced or metastatic breast cancer, and received at least one dose of CDKI or placebo in combination with fulvestrant. The median overall survival was estimated using Kaplan-Meier methods, and hazard ratios (HRs) with corresponding 95% CIs were estimated using Cox regression models. Patients were analysed collectively, by number of previous lines of systemic endocrine therapy in any disease setting (first-line or endocrine naive vs second-line and later), and in various clinicopathological subgroups of interest. The estimated median overall survival was not reported by group when the pooled population included patients treated across lines of therapy because of potential patient heterogeneity. All results presented are considered exploratory and hypothesis generating. Findings Across the three pooled trials, 1960 patients were randomly assigned between Oct 7, 2013, and June 10, 2016 (12 patients were not treated and 1296 [66%] patients were randomly assigned to CDKI and 652 [33%] to placebo). In all treated patients (n=1948), the estimated HR for overall survival was 0·77 (95% CI 0·68–0·88), with a median follow-up of 43·7 months (IQR 37·8–47·7) and deaths in 935 (48%) of the 1948 patients. The difference in estimated median overall survival was 7·1 months, favouring CDKIs. In patients who received CDKIs or placebo in combination with fulvestrant as first-line systemic endocrine therapy (two trials; n=396), the estimated HR for overall survival was 0·74 (95% CI 0·52–1·07), with a median follow-up of 39·4 months (IQR 37·0–42·2). 123 (31%) of these patients died. The difference in estimated median overall survival could not be calculated because median overall survival was not estimable (95% CI 50·9–not estimable) in the CDKI group and was 45·7 months (95% CI 41·7–not estimable) in the placebo group. In patients who received CDKIs or placebo in combination with fulvestrant as second-line or later systemic endocrine therapy (three trials; n=1552), the estimated HR for overall survival was 0·77 (95% CI 0·67–0·89), with a median follow-up of 45·1 months (95% CI 39·2–48·5). 812 (52%) of these patients died. The difference in estimated median overall survival was 7·0 months, favouring CDKIs. Interpretation The addition of CDKIs to fulvestrant resulted in a consistent overall survival benefit in all pooled patients and within most clinicopathological subgroups of interest. These findings support the existing standard of care of CDKIs plus fulvestrant for the treatment of patients with hormone receptor-positive, HER2-negative, advanced breast cancer. Funding None.

Abnormal activity of the core cell-cycle machinery is seen in essentially all tumor types and represents a driving force of tumorigenesis. Recent studies revealed that cell-cycle proteins regulate a wide range of cellular functions, in addition to promoting cell division. With the clinical success of CDK4/6 inhibitors, it is becoming increasingly clear that targeting individual cell-cycle components may represent an effective anti-cancer strategy. Here, we discuss the potential of inhibiting different cell-cycle proteins for cancer therapy.

Summary Background Alpelisib, a PI3Kα-selective inhibitor and degrader, plus fulvestrant showed efficacy in hormone receptor-positive, HER2-negative, PIK3CA-mutated advanced breast cancer in SOLAR-1; limited data are available in the post-cyclin-dependent kinase 4/6 inhibitor setting. BYLieve aimed to assess alpelisib plus endocrine therapy in this setting in three cohorts defined by immediate previous treatment; here, we report results from cohort A. Methods This ongoing, phase 2, multicentre, open-label, non-comparative study enrolled patients with hormone receptor-positive, HER2-negative, advanced breast cancer with tumour PIK3CA mutation, following progression on or after previous therapy, including CDK4/6 inhibitors, from 114 study locations (cancer centres, medical centres, university hospitals, and hospitals) in 18 countries worldwide. Participants aged 18 years or older with an Eastern Cooperative Oncology Group performance status of 2 or less, with no more than two previous anticancer treatments and no more than one previous chemotherapy regimen, were enrolled in three cohorts. In cohort A, patients must have had progression on or after a CDK4/6 inhibitor plus an aromatase inhibitor as the immediate previous treatment. Patients received oral alpelisib 300 mg/day (continuously) plus fulvestrant 500 mg intramuscularly on day 1 of each 28-day cycle and on day 15 of cycle 1. The primary endpoint was the proportion of patients alive without disease progression at 6 months per local assessment using Response Evaluation Criteria in Solid Tumors, version 1.1, in patients with a centrally confirmed PIK3CA mutation. This trial is registered with ClinicalTrials.gov , NCT03056755 . Findings Between Aug 14, 2017, and Dec 17, 2019 (data cutoff), 127 patients with at least 6 months' follow-up were enrolled into cohort A. 121 patients had a centrally confirmed PIK3CA mutation. At data cutoff, median follow-up was 11·7 months (IQR 8·5–15·9). 61 (50·4%; 95% CI 41·2–59·6) of 121 patients were alive without disease progression at 6 months. The most frequent grade 3 or worse adverse events were hyperglycaemia (36 [28%] of 127 patients), rash (12 [9%]), and rash maculopapular (12 [9%]). Serious adverse events occurred in 33 (26%) of 127 patients. No treatment-related deaths were reported. Interpretation BYLieve showed activity of alpelisib plus fulvestrant with manageable toxicity in patients with PIK3CA-mutated, hormone receptor-positive, HER2-negative advanced breast cancer, after progression on a CDK4/6 inhibitor plus an aromatase inhibitor. Funding Novartis Pharmaceuticals.

Radiotherapy combined with chemotherapy is a common modality in abdominal cancer treatment. However, intestinal syndrome induced by radiation is a main factor leading to the poor prognosis of radiotherapy. In this work, we found that miR-378a-3p was markedly up-regulated in the small intestines of mice after total abdominal irradiation. Knocking-down (or overexpression) of miR-378a-3p increased (or decreased) the radiosensitivity of the small intestine cells HIEC and FHs-74-Int. Comet assay and γ-H2AX staining demonstrated that miR-378a-3p exerted its radioprotective function by reducing the accumulation of DNA damage in the cells and tissues of the small intestines. Mechanistically, miR-378a-3p could interact with the 3' UTR of CDK6 through complementary sequences and thus inhibited CDK6 expression in the small intestine cells. Rescue experiments suggested that the repression of miR-378a-3p overexpression on cell radiosensitivity and DNA damage accumulation was abrogated by the forced expression of CDK6. In summary, our results revealed for the first time that miR-378a-3p regulated the radiosensitivity and DNA damage response of small intestines by targeting CDK6. MiR-378a-3p may serve as a promising biomarker and radioprotective target in abdominal cancer.

Background Osteosarcoma (OS) is one of the most commonly diagnosed malignant tumors that mainly affects children and adolescents. The underlying molecular mechanisms that are responsible for the initiation and development of OS are still not clear. Increasing evidence suggested the tumor suppressor role of microRNA-524-5p in a variety of cancers via targeting key pathways involved in tumorigenesis. The aim of this study was to characterize the function of miR-524-5p in OS. Methods A total 50 paired OS tissues and adjacent normal tissues were collected from OS patients. The expression of miR-524-5p in OS tissues and cells was detected by RT-qPCR. The CCK-8 assay, flow cytometry and transwell assay were applied to determine the proliferation and invasion abilities of OS cells. The targets of miR-524-5p were predicted using the miRDB dataset and confirmed by luciferase reporter assay and western blot analysis. Results The expression of miR-524-5p was decreased in OS tissues and cell lines. OS patients with lymph node metastasis harbored relative lower level of miR-524-5p. Overexpression of miR-524-5p in OS cells significantly suppressed the proliferation, drove cell cycle arrest and apoptosis. The mechanism investigation revealed that miR-524-5p bound the 3′-untranslated region (UTR) of Cyclin Dependent Kinase 6 (CDK6) and repressed the expression of CDK6 in OS cells. Overexpressed CDK6 was found in OS tissues, which was inversely correlated with that of miR-524-5p. Moreover, forced expression of CDK6 significantly reversed the anti-cancer effects of miR-524-5p on the proliferation, apoptosis and cell cycle arrest of OS cells. Conclusions Our results identified the tumor-suppressive role of miR-524-5p in OS via targeting CDK6, which may lead to the identification of novel therapeutic target for the treatment of OS.

Pseudogenes have been reported to exert oncogenic or tumor-suppressive functions in cancer. However, the expression, role, and mechanism of pseudogene-derived RNAs in breast cancer remain unclear. The RNA levels and prognostic values of pseudogenes in breast cancer were determined. The levels of RP11-480I12.5 in cell lines and clinical samples were validated by quantitative real-time PCR. In vitro effects of RP11-480I12.5 on cell growth were measured by cell counting kit-8 (CCK-8) assay, colony formation assay, cell counting assay, and flow cytometry analysis. Xenograft model was established to detect its in vivo effect. The potential mechanism of RP11-480I12.5 was also studied by a combination of bioinformatic analysis and experimental confirmation. Finally, the possible functional parental genes of RP11-480I12.5 in breast cancer were explored. After a series of bioinformatic analyses, RP11-480I12.5 was selected as the most potential pseudogene in breast cancer. RP11-480I12.5 expression was significantly upregulated in breast cancer cell lines and clinical breast cancer tissues. Knockdown of RP11-480I12.5 markedly suppressed cell proliferation and colony formation, induced cell apoptosis of breast cancer in vitro, and inhibited tumor growth in vivo. Four transcripts of RP11-480I12.5 (001/002/003/004) were identified. Only overexpression of RP11-480I12.5-004 significantly enhanced cell growth of breast cancer both in vitro and in vivo. RP11-480I12.5-004 mainly located in cytoplasm and increased AKT3 and CDK6 mRNA expression, at least in part, by competitively binding to miR-29c-3p. Six parental genes of RP11-480I12.5 were found, among which TUBA1B and TUBA1C were statistically linked to RP11-480I12.5 expression, possessed prognostic values, and were upregulated in breast cancer. Our findings suggested that pseudogene-derived long non-coding RNA (lncRNA) RP11-480I12.5-004 promoted growth and tumorigenesis of breast cancer via increasing AKT3 and CDK6 expression by competitively binding to miR-29c-3p., Graphical Abstract, By combination of in silico analysis and experimental validation, Lou et al. elucidated that pseudogene-derived lncRNA RP11-480I12.5-004 promoted growth and tumorigenesis of breast cancer via increasing AKT3 and CDK6 expression by competitively binding to miR-29c-3p.

Background/aims Studies have found that LncRNA CYTOR is an important regulator of cancer. However, the function of lncRNA CYTOR in pancreatic cancer (PC) is unclear. This study amid to explore the regulation of lncRNA CYTOR in PC. Methods The expression of CYTOR and miR-205-5p in PC was detected by RT-qPCR. CCK-8 assay, colony formation assay and scratch test were conducted to detect the effects of CYTOR and miR-205-5p on proliferation and migration of PC cells. Target gene prediction and screening and luciferase reporter assays were used to verify downstream target genes of CYTOR and miR-205-5p. The expression of Cyclin-dependent protein kinase 6 (CDK6) was detected by Western blotting. The tumor growth in mice was detected by in vivo experiments in nude mice. Results The expression of LncRNA CYTOR was significantly elevated in PC. Knockdown of CYTOR significantly inhibited cell proliferation and migration of PC cells. In vivo animal studies showed that CYTOR promoted tumor growth. MiR-205-5p was a direct target of CYTOR, and the expression levels of miR-205-5p were significantly reduced in PC cell lines. Furthermore, co-transfection of shCYTOR with miR-205-5p inhibitor partially abolished the effect of shCYTOR on cell proliferation and migration. In addition, CYTOR was negatively correlated with the expression of miR-205-5p. CDK6 was a direct target of miR-205-5p, and miR-205-5p mimic and sh CYTOR significantly reduced the expression levels of CDK6. Conclusion CYTOR can promote PC progression by modulating the miR-205-5p/CDK6 axis, which may be a potential therapeutic target for PC.

Summary The development and approval of cyclin-dependent kinase (CDK) 4 and 6 inhibitors for hormone receptor-positive and human epidermal growth factor receptor 2 (HER2)-negative metastatic breast cancer represents a major milestone in cancer therapeutics. Three different oral CDK4/6 inhibitors, palbociclib, ribociclib, and abemaciclib, have significantly improved progression-free survival by a number of months when combined with endocrine therapy. More recently, improvement in overall survival has been reported with ribociclib and abemaciclib. The toxicity profile of all three drugs is well described and generally easily manageable with dose reductions when indicated. More myelotoxicity is observed with palbociclib and ribociclib, but more gastrointestinal toxicity is observed with abemaciclib. Emerging data is shedding light on the resistance mechanisms associated with CDK4/6 inhibitors, including cell cycle alterations and activation of upstream tyrosine kinase receptors. A number of clinical trials are exploring several important questions regarding treatment sequencing, combinatorial strategies, and the use of CDK4/6 inhibitors in the adjuvant and neoadjuvant settings, thereby further expanding and refining the clinical application of CDK4/6 inhibitors for patients with breast cancer.

Glioblastoma is the most common malignant primary brain tumor among adults and one of the most lethal cancers. It is characterized by the deregulation of signaling pathways involving proliferation, growth, survival, and other factors. MicroRNAs (miRNAs) play a role in the regulation of genes by affecting the 3′ untranslated region (UTR) of mRNA and affect many cell functions. The present study showed that miR-129 decreased the expression of retinoblastoma and p53 signaling pathways’ genes, including CDK4, CDK6, and MDM2. The real-time PCR data indicated that expression of CDK4 in U251 and U87 cell lines declined by 69.8% and 47% (p

BRAF is a serine-threonine kinase that harbors activating mutations in ∼7% of human malignancies and ∼60% of melanomas. Despite initial clinical responses to BRAF inhibitors (BRAFi), patients frequently develop drug resistance. To identify candidate therapeutic targets for BRAFi-resistant melanoma, we conducted CRISPR screens in melanoma cells harboring an activating BRAF mutation that had also acquired resistance to BRAFi. The screens identified pathways and genes critical for BRAFi resistance in melanoma cells. To investigate the mechanisms and pathways enabling resistance to BRAFi in melanomas, we integrated expression data, ATAC-seq, and CRISPR screen results. We identified the JUN family of transcription factors and the ETS family transcription factor ETV5 as key regulators of CDK6 that enabled resistance to BRAFi in melanoma cells. Our findings reveal genes whose loss of function conferred resistance to a selective BRAF inhibitor, providing new insight into signaling pathways that contribute to acquired resistance in melanoma.

Cyclin-dependent kinase 4/6 inhibitors (CDK4/6i) combined with endocrine therapy (ET) are now a backbone of treatment for hormone receptor-positive/human epidermal growth factor receptor 2 (HER2)-negative advanced breast cancer. CDK4/6i plus ET is more effective than ET alone in this setting; however, the risk of grade 3-4 adverse events also increases. Approved agents in this class have similar efficacies, but important differences due to their structural and pharmacological properties. We review biomarkers and discuss determinants to inform a rational approach to therapy choice when selecting the most appropriate ET and CDK4/6i partners. We also identify subgroups that may benefit from specific ET-CDK4/6i combinations and discuss strategies to overcome resistance. This personalized approach aims to minimize treatment-related toxicities that may affect patient QoL and compliance, and ultimately therapy efficacy.

Selective CDK4/6 inhibitors, such as palbociclib, ribociclib, and abemaciclib, have been approved in combination with hormone therapy for the treatment of patients with HR+, HER2-negative advanced or metastatic breast cancer (mBC). Despite their promising activity, approximately 10 % of patients have de novo resistance, while the rest of them will develop acquired resistance after 24-28 months when used as first-line therapy and after a shorter period when used as second-line therapy. Various mechanisms of resistance to CDK4/6 inhibitors have been described, including cell cycle-related mechanisms, such as RB loss, p16 amplification, CDK6 or CDK4 amplification, and cyclin E-CDK2 amplification. Other bypass mechanisms involve the activation of FGFR or PI3K/AKT/mTOR pathways. Identifying the different mechanisms by which resistance to CDK4/6 inhibitors occurs may help to design new treatment strategies to improve patient outcomes. This review presents the currently available knowledge on the mechanisms of resistance to CDK4/6 inhibitors, explores possible treatment strategies that could overcome this therapeutic problem, and summarizes relevant recent clinical trials.

Lung adenocarcinoma (LUAD) is associated with poor prognosis. Identifying novel cancer targets and helpful therapeutic strategies remains a serious clinical challenge. This study detected differentially expressed genes in The Cancer Genome Atlas (TCGA) LUAD data collection. We also identified a predictive DNA biomarker, G protein-coupled receptor 37 (GPR37), which was verified as a prognostic biomarker with a critical role in tumor progression. In human LUAD specimens and microarray analyses, we determined that GPR37 was significantly upregulated and associated with a poor prognosis. GPR37 downregulation markedly inhibited the proliferation and migration of LUAD both in vitro and in vivo. Mechanistically, GPR37 could bind to CDK6, thereby facilitating tumor progression in LUAD by inducing cell cycle arrest at the G1 phase. GPR37 also facilitates tumorigenesis in xenograft tumors in vivo. High-throughput screening for GPR37-targeted drugs was performed using the Natural Products Library, which revealed the potential of Hypocrellin B to inhibit GPR37 and cell growth in LUAD. We demonstrated that Hypocrellin B suppressed LUAD cell proliferation and migration both in vitro and in vivo via GPR37 inhibition. Collectively, our findings reveal the role of GPR37 in LUAD progression and migration and the potential of GPR37 as a target for the treatment of LUAD. Thus, the specific inhibition of GPR37 by the natural product Hypocrellin B may possess the potential for the treatment of LUAD.

Chemotherapy remains the main treatment option for patients with several tumor types. In this issue of Cancer Cell, Salvador-Barbero et al. demonstrate that treatment with CDK4/6 inhibitors after application of taxanes (or other chemotherapeutic compounds) strongly potentiates the anti-tumor effect due to repression of DNA repair machinery.

Targeting CDK4/6 has been identified as an effective therapeutics for treatment of cancer. We herein reported the discovery of a series of 6-(2-(methylamino)ethyl)-5,6,7,8-tetrahydro-1,6-naphthyridin-2-amine derivatives as CDK4/6 inhibitors against cancer. Compound 3c, which displayed high potency and selectivity on CDK4/6 (IC50 = 0.710/1.10 nM) over a variety of other kinases, possessed desirable antiproliferative activities, excellent metabolic properties, and favorable pharmacokinetic characters. In MCF-7, Colo-205, and A549 xenograft models, compound 3c exhibited significant tumor growth inhibitions with low toxicities, which could be a promising drug candidate for further development.

Summary Background Most head and neck squamous-cell carcinomas (HNSCCs) are driven by p16INK4A inactivation and cyclin D1 overexpression that results in hyperactivation of cyclin-dependent kinase 4 and 6 (CDK4/6), rather than by the human papillomavirus (HPV). Deregulated cyclin D1 expression also causes resistance to EGFR inhibitors. We previously reported that palbociclib (a selective CDK4/6 inhibitor) given with cetuximab (an EGFR inhibitor) was safe. The aim of this study was to establish the proportion of patients achieving an objective response with palbociclib and cetuximab in recurrent or metastatic HNSCC. Methods We did a multicentre, multigroup, phase 2 trial to evaluate the activity of palbociclib and cetuximab in platinum-resistant (group 1) and cetuximab-resistant (group 2) HPV-unrelated HNSCC. The study was done across eight university sites in the USA. Eligibility required measurable disease (according to Response Evaluation Criteria in Solid Tumors, version 1·1 [RECIST 1·1]), Eastern Cooperative Oncology Group (ECOG) performance status of 0–2, age of 18 years or older, and disease progression on platinum but cetuximab-naive (group 1) or disease progression on cetuximab (group 2). All patients received palbociclib orally (125 mg/day, on days 1–21) and intravenous cetuximab (400 mg/m2 on cycle one, day 1, then 250 mg/m2 once per week) in 28-day cycles. The primary endpoint was objective response (complete responses and partial responses per RECIST 1·1). Analyses were done per protocol. This trial was registered with ClinicalTrials.gov, NCT02101034, and is ongoing, but both groups are closed to accrual. Findings Between Oct 19, 2015, and Nov 7, 2018, 62 patients were enrolled onto the trial: 30 patients were enrolled in group 1 and 32 in group 2. Median follow-up was 5·4 months (IQR 4·4–12·1) for group 1 and 5·5 months (4·3–8·3) for group 2. In group 1, of 28 evaluable patients, an objective response was achieved by 11 (39%; 95% CI 22–59). In group 2, of 27 evaluable patients, an objective response was achieved by five (19%; 6–38) in group 2. The most common grade 3–4 palbociclib-related adverse event was neutropenia (in 21 [34%] of 62 patients). No treatment-related deaths occurred. Interpretation In patients with platinum-resistant or cetuximab-resistant HPV-unrelated HNSCC, palbociclib and cetuximab results in promising activity outcomes. Further studies of CDK4/6 inhibitors are warranted in HPV-unrelated HNSCC. Funding Pfizer.

The GAIT (gamma-interferon-activated inhibitor of translation) complex or miR-297-RISC (RNA-induced silencing complex), together with hnRNP L or hnRNP L-bearing complex, operates an RNA switch in myeloid cells that regulates stress-dependent expression of vascular endothelial growth factor-A (VEGFA). Here, we have shown that hnRNP L directs multiple hypoxia-inducible RNA switches simultaneously and regulates expression of these oncogenic genes in addition to VEGFA. Bioinformatic and polysome profiling-microarray screens have identified DNM1L (Dynamin 1-like) and PHF21A (PHD finger protein 21A) mRNAs as regulated at the translational level by GAIT-dependent, hnRNP L-directed RNA switches. We have also uncovered CDK6 (Cyclin dependent kinase 6), MKLN1 (Muskelin 1) and EIF5 (Eukaryotic initiation factor 5) as novel miR-297-dependent, hnRNP L-directed RNA switch transcripts. Src Kinase is required for the phosphorylation of hnRNP L and activation of the RNA switch pathway. Knockdown of hnRNP L sensitizes the human U937 monocytic cells under hypoxia stress but not in normoxia via inducing cell apoptosis partially due to the reduced translation of hnRNP L target mRNAs. Collectively, our findings suggest that commonly controlled genes by the hnRNP L-directed RNA switches form a translational regulon that promotes hypoxia resistance and cell survival.

The freshwater red-eared slider (Trachemys scripta elegans) is found not only in freshwater but also in coastal saline habitats. Hyperosmotic salinity can induce cell damage. p53, regarded as the guardian of the genome, is very important and versatile in response to the change of environment. In this study, the role of p53 in T. s. elegans under environmental salinity change will be explored. The results indicated that amino acid sequence of p53 showed high similarity to p53 of other species. In addition, the expression of p53 showed differences in various tissues under normal condition. Under salinity stress, the mRNA levels of p53 in the liver increased significantly at 48 h with 15‰ group (15 practical salinity units-exposed group). In the heart, p53 mRNA levels increased at 6 h in 5‰ (5 practical salinity units) and 15‰ groups. Furthermore, the changes of p21 mRNA expression levels in liver and heart were similar to p53, while cyclin D1, cyclin-dependent kinase4 (CDK4) and cyclin-dependent kinase6 (CDK6) showed opposite changes to p53. Moreover, Bax and caspase 3 mRNA expression levels were similar to p53, respectively, while Bcl-2 showed opposite changes. The positive cells of apoptosis were found in the liver of 15‰ at 48 h and 30 d of chronic stress. Taken together, these results indicated that the T. s. elegans may protect itself by regulating cell cycle progression and apoptosis of damaged cells under salinity stress, which played an important role for T. s. elegans in salinity adaptation.

Objective: The aim of this study was to investigate the mechanism of acupotomology (Apo) in the prevention of articular cartilage destruction via the promotion of chondrocyte proliferation and chondrocyte expression of cell cycle regulators, CyclinD1, CDK4 and CDK6 in a rabbit knee osteoarthritis (KOA) model. Methods: Twenty-eight rabbits were randomly divided into a control group, an OA (osteoarthritis) model group, an Apo (acupotomology) group and EA (electro-acupuncture) group (n = 7). Improved Videman's method was used to induce a rabbit model of KOA over 6 weeks. One week later, acupotomy and electro-acupuncture therapy was applied to animals in the respective groups and treatment lasted 4 weeks. Following these treatments, quantitative real-time PCR, immunohistochemical staining and western blotting were performed to assess the mRNA and protein levels of cell cycle regulators CyclinD1 (Cell cycle protein D1), CDK4 (Cyclin-dependent kinase 4) and CDK6 (Cyclin-dependent kinase 6). Ethology measures and knee morphology were also compared among groups. Results: The Lequesne MG index score of morphology was increased (P .05). Compared with the EA group, the morphology score was decreased in Apo group (P

Background Human urothelial carcinoma associated 1 (UCA1) has been recognized as an oncogenic lncRNA in various cancers, except infantile hemangioma (IH). This study attempts to explore the functional role of lncRNA UCA1 in IH. Methods qRT-PCR was carried out to detect the expression of lncRNA UCA1 in human IH tissues. Two hemangioma cell lines (EOMA and HemECs) were transfected with shRNAs specific for lncRNA UCA1, or a plasmid for expression lncRNA UCA1. The expression of miR-200c in cell was suppressed or overexpressed by miRNA-mediated transfection. CCK-8 assay, flow cytometry, Transwell assay, and Western blot were performed to detect cell survival, migration and invasion. Results LncRNA UCA1 was up-regulated in proliferating-phase hemangioma samples, as compared to involuting-phase. Silence of lncRNA UCA1 significantly reduced EOMA cells viability, migration and invasion, and induced apoptosis. These observations were coupled with the down-regulations of CyclinD1, CDK6 and CDK4, the cleavage of caspase-3 and caspase-9, as well as the decreased expression levels of MMP-9 and Vimentin. miR-200c was highly expressed in lncRNA UCA1 silenced-cells. Besides, the anti-tumor effects of lncRNA UCA1 silence towards EOMA cells were reversed by miR-200c suppression. Same effects of lncRNA UCA1 and miR-200c on HemECs cells were observed. Furthermore, silence of lncRNA UCA1 repressed mTOR, AMPK and Wnt/β-catenin signaling via a miR-200c-dependent fashion. Conclusion This study evidences that silence of lncRNA UCA1 inhibits hemangioma cells growth, migration and invasion possibly via its regulation on miR-200c expression and the activation of mTOR, AMPK and Wnt/β-catenin signaling pathways.

Development of selective kinase inhibitors that target the ATP binding site continues to be a challenge largely due to similar binding pockets. Palbociclib is a cyclin-dependent kinase inhibitor that targets the ATP binding site of CDK4 and CDK6 with similar potency. The enzymatic function associated with the kinase can be effectively probed using kinase inhibitors however the kinase-independent functions cannot. Herein, we report a palbociclib based PROTAC that selectively degrades CDK6 while sparing the homolog CDK4. We used competition studies to characterize the binding and mechanism of CDK6 degradation.

Adrenocortical cancer (ACC) is an aggressive malignancy with no available effective treatments; therefore, exploring the molecular mechanisms involved in the initiation and progression of ACC is quite important. Here, we found that the long noncoding RNA urothelial carcinoma-associated 1 (UCA1) was highly expressed in ACC tissues and closely associated with the TNM stage and metastasis of ACC patients. Overexpression of UCA1 significantly promoted the proliferation and suppressed the apoptosis of ACC cells. Mechanism study showed that UCA1 acted as sponge of miR-298 and decreased the expression abundance of miR-298 in ACC cells. Further investigation identified that miR-298 bound the 3'-UTR of the cyclin-dependent kinase 6 (CDK6) and inhibited the expression of CDK6. Consistently, ectopic expressed UCA1 suppressed miR-298 and up-regulated the expression of CDK6, which promoted the cell cycle progression of ACC cells. Taken together, our results identified the potential oncogenic function of UCA1 in ACC by regulating the miR-298-CDK6 axis.

Cisplatin is a widely used chemotherapeutic drug that often causes acute kidney injury (AKI) in cancer patients. The contribution of miRNAs to the cisplatin-induced renal tubular epithelial cell injury remains largely unknown. Here we performed an integrative network analysis of miRNA and mRNA expression profiles to shed light into the underlying mechanism of cisplatin-induced renal tubular epithelial cell injury. Microarray analysis identified 47 differentially expressed miRNAs, among them 26 were upregulated and 21 were downregulated. Moreover, integrating dysregulated miRNAs target prediction and altered mRNA expression enabled us to identify 1181 putative target genes for further bioinformatics analysis. Gene ontology (GO) analysis revealed that the putative target genes were involved in apoptosis process and regulation of transcription. Pathway analysis indicated that the top upregulated pathways included MAPK and p53 signaling pathway, while the top downregulated pathways were PI3K-Akt and Wnt signaling pathway. Further network analysis showed that MAPK signaling pathway and apoptosis with the highest degree were identified as core pathways, hsa-miR-9-3p and hsa-miR-371b-5p as the most critical miRNAs, and CASK, ASH1L, CDK6 etc. as hub target genes. In addition, the expression level change of selected five microRNAs (hsa-miR-4299, hsa-miR-297, hsa-miR-3135b, hsa-miR-9-3p, and hsa-miR-371b-5p) and two mRNAs( CASK and CDK6) were validated in cisplatin-induced HK-2 cells. Furthermore, a similar trend of expression level change was observed in NRK-52E cells by cisplatin treatment. Overall, our results provide the molecular basis and potential targets for the treatment of cisplatin-induced renal tubular cell injury.

Bisphenol S (BPS), exhibiting estrogenic activity, has been reported to promote cell proliferation in MCF-7 breast cancer cells; however, the underlying mechanism remains unclear. In this study, BPS (1–100 μM) significantly promoted cell proliferation in ERα positive MCF-7 cells, but not in ERα negative MDA-MB-231 or SK-BR-3 cells, confirming the important role of ERα in BPS-induced cell proliferation. Results of the flow cytometry analysis indicated that 10 μM BPS promoted MCF-7 proliferation by accelerating G1 to S phase transition of the cell cycle. BPS increased cyclin D1 expression and phospho-retinoblastoma (p-Rb) levels, resulting in the release of E2F transcription factors and the increased expression of downstream cyclin E2 and cyclin A2 genes to promote the cell cycle progression. Moreover, BPS-induced Rb phosphorylation and cell cycle progression were prevented by the ERα inhibitor ICI 182,780 and methylpiperidino pyrazole, as well as cyclin-dependent kinase 4 (CDK4) and CDK6 inhibitor PD 0332991, indicating that the underlying mechanisms involve ERα-dependent pathways but also mediated by cyclin D-CDK4/6. Overall, our result showed, for the first time, that BPS promoted cell cycle progression and cell proliferation through the ERα-cyclin D-CDK4/6-pRb pathway in MCF-7 breast cancer cells. This study provides a novel insight regarding the potential role of cyclin D-CDK4/6-pRb pathway in mediating the proliferative effects of BPS in breast cancer cells.

Nerve growth factor receptor (NGFR), a member of kinase protein, is emerging as an important target for Glioblastoma (GBM) treatment. Overexpression of NGFR is observed in many metastatic cancers including GBM, promoting tumor migration and invasion. Hydrazones have been reported to effectively interact with receptor tyrosine kinases (RTKs). We report herein the synthesis of 23 arylhydrazones of active methylene compounds (AHAMCs) compounds and their anti-proliferative activity against GBM cell lines, LN229 and U87. Compound R234, 2-(2-(2,4-dioxopentan-3-ylidene)hydrazineyl)benzonitrile, was identified as the most active anti-neoplastic compound, with the IC50 value ranging 87 μM - 107 μM. Molecular docking simulations of the synthesized compounds into the active site of tyrosine receptor kinase A (TrkA), demonstrated a strong binding affinity with R234 and concurs well with the obtained biological results. R234 was found to be a negative regulator of PI3K/Akt/mTOR pathway and an enhancer of p53 expression. In addition, R234 treated GBM cells exhibited the downregulation of cyclins, cyclin-dependent kinases and other key molecules involved in cell cycle such as CCNE, E2F, CCND, CDK6, indicating that R234 induces cell cycle arrest at G1/S. R234 also exerted its apoptotic effects independent of caspase3/7 activity, in both cell lines. In U87 cells, R234 induced oxidative effects whereas LN229 cells annulled oxidative stress. The study thus concludes that R234, being a negative modulator of RTKs and cell cycle inhibitor, may represent a novel class of anti-GBM drugs.

The MYCN oncoprotein induces the proliferation of neuroblastoma cells through regulating gene transcription. The E2F transcription factor 5 (E2F5) plays an oncogenic role in several types of cancer, however, its role in neuroblastoma cells remains poorly characterized. We report here that MYCN positively regulates E2F5 expression in neuroblastoma cells. Analyses of chromatin immunoprecipitation (ChIP) and reporter gene assay demonstrate that MYCN directly binds to a Myc E-Box DNA binding motif within the promoter of E2F5 gene, whereby inducing its transcription. In addition, E2F5 knockdown inhibits the proliferation of MYCN-amplified neuroblastoma cells. Furthermore, E2F5 knockdown inhibits cell cycle progression, which could be attributed to its regulation in the expression of related cyclin-dependent kinases (CDKs), including CDK2 and CDK6. These results suggest that MYCN-regulated E2F5 upregulation is an important mechanism through which MYCN induces the proliferation of neuroblastoma cells. In conclusion, this study identifies E2F5 as a pro-proliferative factor in MYCN-amplified neuroblastoma cells, and also implicates it as a potential target in neuroblastoma treatment.

Gliomas are the most common primary brain malignant tumors in humans. Glioblastoma multiforme(GBM) is the most malignant intracranial tumor with a relatively poor prognosis. There promote us to find effective anti-cancer therapies to reduce cancer mortality. By using bioinformatic analysis, we found SSFA2 as a gene with elevated expression in the glioma tissues. We detected the expression of SSFA2 in glioma tissues and in the glioma cell lines, as well as in normal brain tissues. SSFA2 expression was higher in glioma tissues, especially in glioblastoma multiforme than normal brain tissues. Subsequently, we found that down-regulate SSFA2 in glioma cell lines can regulate the cell cycle to reduce the proliferation ability and induce the early apoptosis rate in shSSFA2 cells relative to control cells. Moreover, we found that down-regulate SSFA2 in glioma cell line U87(shSSFA2-U87) inhibited the growth effectiveness compared to the control cell line U87. These result reveals us that SSFA2 may act as oncogene to promote the progression of glioma. For further research specific mechanisms of SSFA2 in gliomas, we used the gene chip to detect the downstream gene in U87. We found that 30 genes also may be as target gene of SSFA2, and we testify the protein expression by western-blot. The result reveal that IL1A, IL1B and CDK6 as target gene of SSFA2 to regulate the progression of glioma. These finding suggest that SSFA2 could be a new therapeutic target for gliomas.

Three Cyclin Dependent Kinase 4/6 (CDK) inhibitors have been approved by the United Stated Food and Drug Administration for front line treatment of advanced hormone receptor positive breast cancer based on improvements in progression free survival against endocrine monotherapy. Two clinical trials have so far reported results on overall survival but both are negative. CDK inhibitors are usually tolerated well but they do add to inconvenience and cost - for example, grade III-IV neutropenia occur at a frequency of over 60% requiring frequent blood work at least during the initial months of treatment. These drugs cost over $ 13,500 for a 4-week cycle in the United States, and are responsible for billions of dollars annually in drug cost alone. Importantly, many women with metastatic breast cancer do well for a long time with endocrine therapy alone and CDK inhibitors do not have a predictive marker. Selective use of these agents in later lines may improve substantially the convenience and cost without compromise in overall outcome. However, with results demonstrating impressive improvements in PFS published in major medical journals coupled with patients' natural desire for “best available” options, the trend among oncologists is to prescribe these drugs as the default front-line treatment. In this commentary I caution readers against over interpretation of results from the CDK inhibitor trials, describe adverse consequences of routine front-line use, and explain why selective use in later line may yield a higher value.

Illuminating the correlations between non-coding RNAs and protein-coding genes are of great interest to understand more about the molecular mechanisms that drive malignant transformation and understanding such correlations will enable development of more specific and efficient targeted therapeutics. Accordingly, in this comprehensive meta-analysis study, we tried to determine correlations between long non-coding RNA (lncRNA), microRNA (miRNA) and messenger RNA (mRNA) molecules that are involved in the pathogenesis of colorectal cancer (CRC). For the present study, current colorectal cancer studies published until 20 August 2017 and associated with the lncRNA-miRNA-mRNA interactions was included. The current literature search was done online in Pubmed, Embase and Web of Science databases. These databases have been screened with three keywords; “lncRNA”, “miRNA” and “colorectal cancer”. As a result, colorectal neoplasia differentially expressed (CRNDE) was determined to be consistently up-regulated in CRC tissues and inversely associated with miR-181a-5p. Also, CRNDE expression was significantly associated with lymph node metastasis (p = 0.032). Additionally, a significant association was determined between CRC and survival time in the OS analysis of FER1L4, TUSC7, UCC and lincRNA-ROR. More importantly, there was a negative correlation between lncRNA-miRNA expressions (p = 0.001). Particularly, CRNDE/miR-181a-5p, FER1L4/miR-106a-5p, TUSC7/miR-211, UCC/miR-143 and lincRNA-ROR/miR-145 was negatively correlated. In addition, there was a significant positive correlation between GAPLINC/CD44 and TUSC7/CDK6 in CRC tissues (p = 0.000). Overall analysis showed that lncRNAs and mRNAs, which are targets of the same miRNA, have positive interactions. In addition, miRNAs were shown to have negative correlations with target lncRNA/mRNA.

MicroRNA-107 (miR-107) plays a regulatory role in obesity and insulin resistance, but the mechanisms of its function in adipocytes have not been elucidated in detail. Here we show that overexpression of miR-107 in pre- and mature human Simpson-Golabi-Behmel syndrome (SGBS) adipocytes attenuates differentiation and lipid accumulation. Our results suggest that miR-107 controls adipocyte differentiation via CDK6 and Notch signaling. CDK6 is a validated target of miR-107 and was downregulated upon miR-107 overexpression. Notch3, a signaling receptor involved in adipocyte differentiation, has been shown to decrease upon CDK6 depletion; Here Notch3 and its target Hes1 were downregulated by miR-107 overexpression. In mature adipocytes, miR-107 induces a triglyceride storage defect by impairing glucose uptake and triglyceride synthesis. To conclude, our data suggests that miR-107 has distinct functional roles in preadipocytes and mature adipocytes; Its elevated expression at these different stages of adipocytes may on one hand dampen adipogenesis, and on the other, promote ectopic fatty acid accumulation and reduced glucose tolerance.

Tripartite motif family-like 2 (TRIML2), a member of the TRIM proteins family, is closely related to Alzheimer's disease, however, no studies of TRIML2 have been published in the cancer research literature. In the current study, we investigated the expression level of TRIML2 and its molecular mechanisms in human oral squamous cell carcinoma (OSCC); reverse transcriptase-quantitative polymerase chain reaction, immunoblot analysis, and immunohistochemistry showed that TRIML2 is up-regulated significantly in OSCCs in vitro and in vivo. TRIML2 knockdown OSCC cells showed decreased cellular proliferation by cell-cycle arrest at G1 phase that resulted from down-regulation of CDK4, CDK6, and cyclin D1 and up-regulation of p21Cip1 and p27Kip1. Surprisingly, resveratrol, a polyphenol, led to not only down-regulation of TRIML2 but also cell-cycle arrest at G1 phase similar to TRIML2 knockdown experiments. Taken together, we concluded that TRIML2 might play a significant role in tumoral growth and that resveratrol may be a new drug for treating OSCC by interfering with TRIML2 function.

Dermatopontin (DPT), a noncollagenous extracellular matrix component, has been illustrated to regulate cellular proliferation and invasiveness in several types of neoplasms. Nevertheless, the biological functions of DPT in cell proliferation, especially papillary thyroid cancer (PTC) cell proliferation, remain unknown, as do the mechanisms underlying its effects. In this study, we detected low DPT expression in PTC, which was related to higher T classifications. Ectopic DPT expression impeded cell proliferation both in vitro and in vivo. Furthermore, we illustrated that DPT down-regulated MYC, which in turn targeted CDK4, CDK6 and p21, through the ERK pathway. These results suggest that DPT regulates CDK4, CDK6 and p21, through MEK-ERK-MYC signaling to repress PTC proliferation.

Synthetic transformations of steroids for drug discovery and improvement of drug effectiveness have been an important part of modern medicinal chemistry and pharmaceutical sciences. Pentacyclic triterpenoids, being represented in the nature by various structures and biogenetically related to steroids, can largely expand the spectrum of biologically active steroidal agents via synthesis of the so-called “triterpenoid-steroid” hybrids. In the presented work, the nitrile anion cyclizations of 3,4-secolupane and 3,4-seco-oleanane nitriles and follow-up synthetic transformations of the cyclized products with formation of the gemm-dimethyl-free A ring “triterpenoid-steroid” hybrids were studied. Furthermore, the resulting cyclic compounds were modified at C3, C4, and/or C5 positions of ring A, as well as at C20, C28, and C30 positions of the isopropylidene moiety in the case of lupane triterpenoids. The cytotoxic effect of the synthesized compounds against seven cancer cell lines HEp-2, HCT 116, MS, RD TE32, A549, MCF7, and PC3 was evaluated. The in silico identification of potential anticancer protein targets with regard to the compounds, which were active at micromolar concentrations against tested cell lines, was carried out. The molecular docking studies showed that compound 19, which demonstrated most pronounced cytotoxicity (IC50 0.64–3.17 μM) against all tested cell lines, fits well the active sites of CDK6 and HER2/neu.

Background: Increasing evidence confirmed that circRNAs play a variety of roles in the biological processes of cancer, including tumor cell proliferation, differentiation, and apoptosis. In this study, we identified differentially expressed circRNAs and explored its regulatory role in the development of bladder cancer. Methods: circRNA microarray and qRT-PCR were used to identify the dysregulation of circLAMA3 in bladder cancer. The precise role of circLAMA3 in cell proliferation was identified by bioinformatics analysis, RNA-fluorescence in situ hybridization, and function loss and gain experiment. The potential mechanism of circLAMA3 in bladder cancer by assessed via Western blotting, luciferase reporter gene, and TRAP experiment. Findings: Functional experiments demonstrated that circLAMA3 significantly inhibited the proliferation, migration, and invasion of bladder cancer cell and also inhibited bladder cancer growth in vivo. Mechanically, CircLAMA3 directly binds MYCN mRNA and promotes the degradation of MYCN mRNA to reduce MYCN protein expression in bladder cancer cells. Decreased MYCN protein inhibits the promoter activity and expression level of CDK6. Ultimately, circLAMA3 affects DNA replication by downregulating CDK6, resulting in G0/G1 phase arrest and inhibition of bladder cancer proliferation. Interpretation: We report a new regulatory mechanism that circRNA might directly bind mRNA and participate in the regulation of this mRNA fate. Moreover, CircLAMA3 can significantly affect the progress of bladder cancer and is expected to be potential biomarkers and therapeutic targets for bladder cancer diagnosis and treatment. Funding: This work was supported by the National Natural Science Foundation of China(NSFC81903356). Declaration of Interest: None to declare. Ethical Approval: All of the organizational uses were approved by Ethics Committee of Wenzhou Medical University and the patients provided informed consent.

Application of adipose-derived mesenchymal stromal cells (AMSCs)-derived extracellular vesicles (EVs) in skin wound healing has been documented. In this study, we investigated the therapeutic potential of AMSCs-derived EVs in skin wound healing through delivery of microRNA-10b (miR-10b). HaCaT cells were treated with H

Circular RNAs (circRNAs) are increasingly implicated in the development of oral squamous cell carcinoma (OSCC). Here, we explored the precise role of circRNA oxysterol binding protein like 10 (circ_OSBPL10, circ_0008549) in OSCC pathogenesis.Ribonuclease (RNase) R assay was performed to assess the stability of circ_OSBPL10. The levels of circ_OSBPL10, microRNA (miR)-299-3p and cyclin-dependent kinase 6 (CDK6) mRNA were gauged by qRT-PCR. CDK6 protein level was measured by western blot. Cell proliferation was detected by MTT and colony formation assays. Cell cycle distribution and apoptosis were measured by flow cytometry. Cell migration and invasion were evaluated using transwell assay. Dual-luciferase reporter assay was used to identify the relationship between miR-299-3p and circ_OSBPL10 or CDK6. Animal studies were performed to evaluate the role of circ_OSBPL10 in tumor growth in vivo.Circ_OSBPL10 was up-regulated in OSCC tissues and cells. Silencing of circ_OSBPL10 hindered cell proliferation, cell cycle progression, colony formation, migration, invasion, and promoted apoptosis in vitro and diminished tumor growth in vivo. Mechanistically, circ_OSBPL10 directly targeted miR-299-3p, and circ_OSBPL10 silencing affected cell functional properties in vitro by up-regulating miR-299-3p. CDK6 was a direct and functional target of miR-299-3p. The circ_OSBPL10/miR-299-3p axis regulated cell functional properties in vitro via CDK6. Moreover, circ_OSBPL10 acted as a competing endogenous RNA (ceRNA) to regulate CDK6 expression through miR-299-3p.Our present findings first demonstrate that circ_OSBPL10 can regulate the functional behaviors of OSCC cells at least partially by miR-299-3p/CDK6 axis, highlighting circ_OSBPL10 as a promising therapeutic target for OSCC.

Objectives: Ipatasertib (IPAT) is an orally administered selective protein kinase B (AKT) inhibitor that has demonstrated clinical activity in triple-negative breast and metastatic prostate cancer. Given that the AKT/mTOR pathway is altered in the vast majority of endometrioid endometrial cancers (ECs), it is logical that IPAT may also have efficacy in EC. Thus, we aimed to evaluate the anti-proliferative effects of IPAT in human endometrioid cell lines as well as in the LKB1fl/flp53fl/fl genetically engineered mouse model of endometrioid EC. Methods: The human endometrioid EC cell lines ECC-1 (PTEN mutant) and HEC-1A (PTEN wild type, PIK3CA mutant, KRAS mutant) were exposed to varying concentrations of IPAT (Genentech). Cell proliferation was assessed by MTT and colony assays. Cell cycle progression was measured by Cellometer. Apoptosis was assessed by cleaved caspase-3 assay. Cellular stress was assessed using tetra-methylrhodamine ester (TMRE) and DCFDA assays. Western immunoblotting determined effects of IPAT on BCL-2, MCL-1, Bip, CDK4, CDK6, cyclin D1, pAKT, pS6, PERK, calnexin, and PDI in both cell lines.LKB1fl/flp53fl/fl mice were used to evaluate the effect of IPAT on endometrial tumor growth. The mice were treated with placebo or IPAT (15mg/kg daily intraperitoneally for 4 weeks) starting 8 weeks after tumor induction via AdCre injection. Results: IPAT inhibited cellular proliferation in a dose dependent fashion in both cell lines after 72 hours of treatment. Median IC50 was 0.18µM in ECC-1 and 3.5µM in HEC-1A. IPAT induced the activity of cleaved caspase 3 by 15.6-fold in ECC-1 and 1.41-fold in HEC-1A (p Conclusions: IPAT significantly inhibited cell proliferation and induced apoptosis and cellular stress via inhibition of the AKT/mTOR pathway in human endometrioid EC cell lines. Furthermore, IPAT reduced tumor growth in a transgenic mouse model of endometrioid EC. These results suggest that IPAT is worthy of further exploration in clinical trials for EC.

Objectives: Ipatasertib (IPAT) is an orally administered selective protein kinase B (AKT) inhibitor that has demonstrated clinical activity in triple-negative breast and metastatic prostate cancer. Given that the AKT/mTOR pathway is altered in the vast majority of endometrioid endometrial cancers (ECs), it is logical that IPAT may also have efficacy in EC. Thus, we aimed to evaluate the anti-proliferative effects of IPAT in human endometrioid cell lines as well as in the LKB1fl/flp53fl/fl genetically engineered mouse model of endometrioid EC. Methods: The human endometrioid EC cell lines ECC-1 (PTEN mutant) and HEC-1A (PTEN wild type, PIK3CA mutant, KRAS mutant) were exposed to varying concentrations of IPAT (Genentech). Cell proliferation was assessed by MTT and colony assays. Cell cycle progression was measured by Cellometer. Apoptosis was assessed by cleaved caspase-3 assay. Cellular stress was assessed using tetramethylrhodamine ester (TMRE) and DCFDA assays. Western immunoblotting determined effects of IPAT on BCL-2, MCL-1, Bip, CDK4, CDK6, cyclin D1, pAKT, pS6, PERK, calnexin, and PDI in both cell lines. LKB1fl/flp53fl/fl mice were used to evaluate the effect of IPAT on endometrial tumor growth. The mice were treated with placebo or IPAT (15mg/kg daily intraperitoneally for 4 weeks) starting 8 weeks after tumor induction via AdCre injection. Results: IPAT inhibited cellular proliferation in a dose dependent fashion in both cell lines after 72 hours of treatment. Median IC50 was 0.18µM in ECC-1 and 3.5µM in HEC-1A. IPAT induced the activity of cleaved caspase 3 by 15.6-fold in ECC-1 and 1.41-fold in HEC-1A (p Conclusions: IPAT significantly inhibited cell proliferation and induced apoptosis and cellular stress via inhibition of the AKT/mTOR pathway in human endometrioid EC cell lines. Furthermore, IPAT reduced tumor growth in a transgenic mouse model of endometrioid EC. These results suggest that IPAT is worthy of further exploration in clinical trials for EC.

Everyone carries a set of genetic variants that contribute to regulation of the levels of blood cells, with unknown clinical impact. One of them, rs445 within the cell-cycle checkpoint gene CDK6, reduces the levels of myeloid cell types including granulocytes. We treated CD3+ T cells and whole blood with palbociclib in 41 individuals, who were stratified by genotype for analyses. In T cells we assessed cell cycle and apoptosis, whereas in whole blood, apoptosis in activated (CD11b+), unactivated (CD11b-) granulocytes, cytotoxic (CD8 + CD4-), and helper (CD8-CD4+) T cells. We find that rs445 modulates the immune response of CD8+ T cells. It also increases the level of apoptotic CD11b + activated granulocytes after palbociclib treatment, which, in synergy with neutropenia, may affect drug related adverse events. These results suggest that the effect of palbociclib treatment may depend on underlying genetically encoded individual immune response as well as the direct response to the drug.

CDK4/6 were attractive chemotherapeutic targets for the treatment of malignant tumors, CDK4/6 selective inhibitors have made outstanding contributions in the treatment of breast cancer. However, these inhibitors share a single skeleton of N-(pyridin-2-yl) pyrimidin-2-amine which cannot overcome the side effects in clinical application. In our previous study, an N'- acetylpyrrolidine-1-carbohydrazide was hit as the initial fragment by analyzing the active site characteristics of CDK6. Two series of N-(pyridin-3-yl) proline were obtained by fragment growth method. The QSAR study was carried out according to the in vitro activities data against CDK4/6, and two compounds 7c and 7p with potent inhibitory activities were found to interact with CDK4 in different binding conformation. They showed potential inhibition of cell proliferation against the breast cancer cell, and 7c exhibited promised anti-breast cancer effect in vivo.

Circular RNAs (circRNAs) have been reported to play critical roles in the malignant progression of diverse human cancers, including multiple myeloma (MM). This study aimed to explore the functional role and underlying mechanism of circ_SEC61A1 in MM progression.Quantitative real-time polymerase chain reaction (qRT-PCR) assay was performed to detect the expression levels of circ_SEC61A1, microRNA (miR)-660-5p and cyclin-dependent kinase 6 (CDK6) mRNA. The localization of circ_SEC61A1 in MM cells was tested by the subcellular fractionation location assay. Actinomycin D assay was conducted to determine the characteristics of circ_SEC61A1. Cell proliferation was evaluated by colony formation assay, 5-ethynyl-2'-deoxyuridine (EdU) incorporation assay and 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Western blot assay was exploited to examine the expression of proteins. Cell migration and invasion were tested via transwell assay, and cell apoptosis was measured by flow cytometry analysis. Dual-luciferase reporter assay was utilized to confirm the interaction between miR-660-5p and circ_SEC61A1 or CDK6.Circ_SEC61A1 level was increased in MM tissues and cells. Circ_SEC61A1 was a stable circRNA and mainly located in cytoplasm. Circ_SEC61A1 silence restrained the proliferation, metastasis and expedited the apoptosis in MM cells. CDK6 was identified as the target of miR-660-5p, and circ_SEC61A1 sponged miR-660-5p to positively regulate CDK6 expression. The inhibitory impacts of circ_SEC61A1 knockdown on the progression of MM cells were mitigated by miR-660-5p inhibition. MiR-660-5p overexpression blocked the malignant phenotypes of MM cells by targeting CDK6.Our study manifested that circ_SEC61A1 could accelerate MM progression at least partially through modulating miR-660-5p/CDK6 axis.