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Simple Summary:
CDK9, in combination with Cyclin T1, is one of the major regulators of RNA Polymerase II mediated productive transcription of critical genes in any cell. The activity of CDK9 is significantly up-regulated in a wide variety of cancer entities, to aid in the overexpression of genes responsible for the regulation of functions, which are beneficial to the cancer cells, like proliferation, survival, cell cycle regulation, DNA damage repair and metastasis. Enhanced CDK9 activity, therefore, leads to poorer prognosis in many cancer types, offering the rationale to target it using small-molecule inhibitors. Several, increasingly specific inhibitors, have been developed, some of which are presently in clinical trials. Other approaches being tested involve combining inhibitors against CDK9 activity with those against CDK9’s upstream regulators like BRD4, SEC and HSP90; or downstream effectors like cMYC and MCL-1. The inhibition of CDK9’s activity holds the potential to be a highly effective anti-cancer therapeutic.
Abstract:
Cyclin Dependent Kinase 9 (CDK9) is one of the most important transcription regulatory members of the CDK family. In conjunction with its main cyclin partner—Cyclin T1, it forms the Positive Transcription Elongation Factor b (P-TEFb) whose primary function in eukaryotic cells is to mediate the positive transcription elongation of nascent mRNA strands, by phosphorylating the S2 residues of the YSPTSPS tandem repeats at the C-terminus domain (CTD) of RNA Polymerase II (RNAP II). To aid in this process, P-TEFb also simultaneously phosphorylates and inactivates a number of negative transcription regulators like 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole (DRB) Sensitivity-Inducing Factor (DSIF) and Negative Elongation Factor (NELF). Significantly enhanced activity of CDK9 is observed in multiple cancer types, which is universally associated with significantly shortened Overall Survival (OS) of the patients. In these cancer types, CDK9 regulates a plethora of cellular functions including proliferation, survival, cell cycle regulation, DNA damage repair and metastasis. Due to the extremely critical role of CDK9 in cancer cells, inhibiting its functions has been the subject of intense research, resulting the development of multiple, increasingly specific small-molecule inhibitors, some of which are presently in clinical trials. The search for newer generation CDK9 inhibitors with higher specificity and lower potential toxicities and suitable combination therapies continues. In fact, the Phase I clinical trials of the latest, highly specific CDK9 inhibitor BAY1251152, against different solid tumors have shown good anti-tumor and on-target activities and pharmacokinetics, combined with manageable safety profile while the phase I and II clinical trials of another inhibitor AT-7519 have been undertaken or are undergoing. To enhance the effectiveness and target diversity and reduce potential drug-resistance, the future of CDK9 inhibition would likely involve combining CDK9 inhibitors with inhibitors like those against BRD4, SEC, MYC, MCL-1 and HSP90.
The spindle assembly checkpoint (SAC) acts as a molecular safeguard in ensuring faithful chromosome transmission during mitosis, which is regulated by a complex interplay between phosphatases and kinases including PLK1. Adenomatous polyposis coli (APC) germline mutations cause aneuploidy and are responsible for familial adenomatous polyposis (FAP). Here we study the role of PLK1 in colon cancer cells with chromosomal instability promoted by APC truncation (APC-ΔC). The expression of APC-ΔC in colon cells reduces the accumulation of mitotic cells upon PLK1 inhibition, accelerates mitotic exit and increases the survival of cells with enhanced chromosomal abnormalities. The inhibition of PLK1 in mitotic, APC-∆C-expressing cells reduces the kinetochore levels of Aurora B and hampers the recruitment of SAC component suggesting a compromised mitotic checkpoint. Furthermore, Plk1 inhibition (RNAi, pharmacological compounds) promotes the development of adenomatous polyps in two independent ApcMin/+ mouse models. High PLK1 expression increases the survival of colon cancer patients expressing a truncated APC significantly.
Ligand stimulation of CD95 induces activation of Plk3 followed by phosphorylation of caspase-8
(2016)
Upon interaction of the CD95 receptor with its ligand, sequential association of the adaptor molecule FADD (MORT1), pro-forms of caspases-8/10, and the caspase-8/10 regulator c-FLIP leads to the formation of a death-inducing signaling complex. Here, we identify polo-like kinase (Plk) 3 as a new interaction partner of the death receptor CD95. The enzymatic activity of Plk3 increases following interaction of the CD95 receptor with its ligand. Knockout (KO) or knockdown of caspase-8, CD95 or FADD prevents activation of Plk3 upon CD95 stimulation, suggesting a requirement of a functional DISC for Plk3 activation. Furthermore, we identify caspase-8 as a new substrate for Plk3. Phosphorylation occurs on T273 and results in stimulation of caspase-8 proapoptotic function. Stimulation of CD95 in cells expressing a non-phosphorylatable caspase-8-T273A mutant in a rescue experiment or in Plk3-KO cells generated by CRISPR/Cas9 reduces the processing of caspase-8 prominently. Low T273 phosphorylation correlates significantly with low Plk3 expression in a cohort of 95 anal tumor patients. Our data suggest a novel mechanism of kinase activation within the Plk family and propose a new model for the stimulation of the extrinsic death pathway in tumors with high Plk3 expression.
Polo-like kinases (PLKs) belong to a five-membered family of highly conserved serine/threonine kinases (PLK1-5) that play differentiated and essential roles as key mitotic kinases and cell cycle regulators and with this in proliferation and cellular growth. Besides, evidence is accumulating for complex and vital non-mitotic functions of PLKs. Dysregulation of PLKs is widely associated with tumorigenesis and by this, PLKs have gained increasing significance as attractive targets in cancer with diagnostic, prognostic and therapeutic potential. PLK1 has proved to have strong clinical relevance as it was found to be over-expressed in different cancer types and linked to poor patient prognosis. Targeting the diverse functions of PLKs (tumor suppressor, oncogenic) are currently at the center of numerous investigations in particular with the inhibition of PLK1 and PLK4, respectively in multiple cancer trials. Functions of PLKs and the effects of their inhibition have been extensively studied in cancer cell culture models but information is rare on how these drugs affect benign tissues and organs. As a step further towards clinical application as cancer targets, mouse models therefore play a central role. Modelling PLK function in animal models, e.g., by gene disruption or by treatment with small molecule PLK inhibitors offers promising possibilities to unveil the biological significance of PLKs in cancer maintenance and progression and give important information on PLKs’ applicability as cancer targets. In this review we aim at summarizing the approaches of modelling PLK function in mice so far with a special glimpse on the significance of PLKs in ovarian cancer and of orthotopic cancer models used in this fatal malignancy.
Introduction: We examined if a combination of proliferation markers and estrogen receptor (ER) activity could predict early versus late relapses in ER-positive breast cancer and inform the choice and length of adjuvant endocrine therapy.
Methods: Baseline affymetrix gene-expression profiles from ER-positive patients who received no systemic therapy (n = 559), adjuvant tamoxifen for 5 years (cohort-1: n = 683, cohort-2: n = 282) and from 58 patients treated with neoadjuvant letrozole for 3 months (gene-expression available at baseline, 14 and 90 days) were analyzed. A proliferation score based on the expression of mitotic kinases (MKS) and an ER-related score (ERS) adopted from Oncotype DX® were calculated. The same analysis was performed using the Genomic Grade Index as proliferation marker and the luminal gene score from the PAM50 classifier as measure of estrogen-related genes. Median values were used to define low and high marker groups and four combinations were created. Relapses were grouped into time cohorts of 0-2.5, 0-5, 5-10 years.
Results: In the overall 10 years period, the proportional hazards assumption was violated for several biomarker groups indicating time-dependent effects. In tamoxifen-treated patients Low-MKS/Low-ERS cancers had continuously increasing risk of relapse that was higher after 5 years than Low-MKS/High-ERS cancers [0 to 10 year, HR 3.36; p = 0.013]. High-MKS/High-ERS cancers had low risk of early relapse [0-2.5 years HR 0.13; p = 0.0006], but high risk of late relapse which was higher than in the High-MKS/Low-ERS group [after 5 years HR 3.86; p = 0.007]. The High-MKS/Low-ERS subset had most of the early relapses [0 to 2.5 years, HR 6.53; p < 0.0001] especially in node negative tumors and showed minimal response to neoadjuvant letrozole. These findings were qualitatively confirmed in a smaller independent cohort of tamoxifen-treated patients. Using different biomarkers provided similar results.
Conclusions: Early relapses are highest in highly proliferative/low-ERS cancers, in particular in node negative tumors. Relapses occurring after 5 years of adjuvant tamoxifen are highest among the highly-proliferative/high-ERS tumors although their risk of recurrence is modest in the first 5 years on tamoxifen. These tumors could be the best candidates for extended endocrine therapy.
Heterogenous subtypes of breast cancer need to be analyzed separately. Pooling of datasets can provide reasonable sample sizes but dataset bias is an important concern. We assembled a combined dataset of 579 Affymetrix microarrays from triple negative breast cancer (TNBC) in Gene Expression Omnibus (GEO) series GSE31519. We developed a method for selecting comparable datasets and to control for the amount of dataset bias of individual probesets.
The taxanes are effective microtubule-stabilizing chemotherapy drugs that inhibit mitosis, induce apoptosis, and produce regression in a fraction of cancers that arise at many sites including the ovary. Novel therapeutic targets that augment taxane effects are needed to improve clinical chemotherapy response in CCNE1-amplified high grade serous ovarian cancer (HGSOC) cells. In this study, we conducted an siRNA-based kinome screen to identify modulators of mitotic progression in CCNE1-amplified HGSOC cells that may influence clinical paclitaxel response. PLK1 is overexpressed in many types of cancer, which correlates with poor prognosis. Here, we identified a novel synthetic lethal interaction of the clinical PLK1 inhibitor BI6727 and the microtubule-targeting drug paclitaxel in HGSOC cell lines with CCNE1-amplification and elucidated the underlying molecular mechanisms of this synergism. BI6727 synergistically induces apoptosis together with paclitaxel in different cell lines including a patient-derived primary ovarian cancer culture. Moreover, the inhibition of PLK1 reduced the paclitaxel-induced neurotoxicity in a neurite outgrowth assay. Mechanistically, the combinatorial treatment with BI6727/paclitaxel triggers mitotic arrest, which initiates mitochondrial apoptosis by inactivation of anti-apoptotic BCL-2 family proteins, followed by significant loss of the mitochondrial membrane potential and activation of caspase-dependent effector pathways. This conclusion is supported by data showing that BI6727/paclitaxel-co-treatment stabilizes FBW7, a component of SCF-type ubiquitin ligases that bind and regulate key modulators of cell division and growth including MCL-1 and Cyclin E. This identification of a novel synthetic lethality of PLK1 inhibitors and a microtubule-stabilizing drug has important implications for developing PLK1 inhibitor-based combination treatments in CCNE1-amplified HGSOC cells.
Paclitaxel is a frontline drug for the treatment of epithelial ovarian cancer (EOC). However, following paclitaxel-platinum based chemotherapy, tumor recurrence occurs in most ovarian cancer patients. Chromosomal instability (CIN) is a hallmark of cancer and represents genetic variation fueling tumor adaptation to cytotoxic effects of anticancer drugs. In this study, our Kaplan-Meier analysis including 263 ovarian cancer patients (stages I/II) revealed that high Polo-like kinase (PLK) 1 expression correlates with bad prognosis. To evaluate the role of PLK1 as potential cancer target within a combinatorial trial, we induced strong mitotic arrest in ovarian cancer cell lines by synergistically co-targeting microtubules (paclitaxel) and PLK1 (BI6727) followed by pharmaceutical inhibition of the Anaphase-Promoting Complex (APC/C) using proTAME. In short- and long-term experiments, this triple treatment strongly activated apoptosis in cell lines and primary ovarian cells derived from cancer patients. Mechanistically, BI6727/paclitaxel/proTAME stabilize Cyclin B1 and trigger mitotic arrest, which initiates mitochondrial apoptosis by inactivation of antiapoptotic BCL-2 family proteins, followed by activation of caspase-dependent effector pathways. This triple treatment prevented endoreduplication and reduced CIN, two mechanisms that are associated with aggressive tumors and the acquisition of drug resistance. This “two-punch strategy” (strong mitotic arrest followed by blocking mitotic exit) has important implications for developing paclitaxel-based combinatorial treatments in ovarian cancer.
Myomas, also known as fibroids, are a specific characteristic of the human species. No other primates develop fibroids. At a cellular level, myomas are benign hyperplastic lesions of uterine smooth muscle cells. There are interesting theoretical concepts that link the development of myomas in humans with the highly specific process of childbirth from an upright position and the resulting need for greatly increased “expulsive” forces during labor. Myomas might be the price our species pays for our bipedal and highly intelligent existence. Myomas affect, with some variability, all ethnic groups and approximately 50% of all women during their lifetime. While some remain asymptomatic, myomas can cause significant and sometimes life-threatening uterine bleeding, pain, infertility, and, in extreme cases, ureteral obstruction and death. Traditionally, over 50% of all hysterectomies were performed for fibroids, leading to a significant healthcare burden. In this article, we review the developments of the past 20 years with regard to multiple new treatment strategies that have evolved during this time.
Background: Reports of head and neck ultrasound examinations are frequently written by hand as free texts. Naturally, quality and structure of free text reports is variable, depending on the examiner’s individual level of experience. Aim of the present study was to compare the quality of free text reports (FTR) and structured reports (SR) of head and neck ultrasound examinations.
Methods: Both standard FTRs and SRs of head and neck ultrasound examinations of 43 patients were acquired by nine independent examiners with comparable levels of experience. A template for structured reporting of head and neck ultrasound examinations was created using a web-based approach. FTRs and SRs were evaluated with regard to overall quality, completeness, required time to completion, and readability by four independent raters with different specializations (Paired Wilcoxon test, 95% CI) and inter-rater reliability was assessed (Fleiss’ kappa). A questionnaire was used to compare FTRs vs. SRs with respect to user satisfaction (Mann-Whitney U test, 95% CI).
Results: By comparison, completeness scores of SRs were significantly higher than FTRs’ completeness scores (94.4% vs. 45.6%, p < 0.001), and pathologies were described in more detail (91.1% vs. 54.5%, p < 0.001). Readability was significantly higher in all SRs when compared to FTRs (100% vs. 47.1%, p < 0.001). The mean time to complete a report, however, was significantly higher in SRs (176.5 vs. 107.3 s, p < 0.001). SRs achieved significantly higher user satisfaction ratings (VAS 8.87 vs. 1.41, p < 0.001) and a very high inter-rater reliability (Fleiss’ kappa 0.92).
Conclusions: As compared to FTRs, SRs of head and neck ultrasound examinations are more comprehensive and easier to understand. On the balance, the additional time needed for completing a SR is negligible. Also, SRs yield high inter-rater reliability and may be used for high-quality scientific data analyses.