Two different methods of presenting quantitative gene expression exist: absolute and relative quantification. Absolute quantification calculates the copy number of the gene usually by relating the PCR signal to a standard curve. Relative gene expression presents the data of the gene of interest relative to some calibrator or internal control gene. A widely used method to present relative gene expression is the comparative C(T) method also referred to as the 2 (-DeltaDeltaC(T)) method. This protocol provides an overview of the comparative C(T) method for quantitative gene expression studies. Also presented here are various examples to present quantitative gene expression data using this method.

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Analyzing real-time PCR data by the comparative C(T) method.

Background A more accurate means of prognostication in breast cancer will improve the selection of patients for adjuvant systemic therapy. Methods Using microarray analysis to evaluate our previously established 70-gene prognosis profile, we classified a series of 295 consecutive patients with primary breast carcinomas as having a gene-expression signature associated with either a poor prognosis or a good prognosis. All patients had stage I or II breast cancer and were younger than 53 years old; 151 had lymph-node–negative disease, and 144 had lymph-node–positive disease. We evaluated the predictive power of the prognosis profile using univariable and multivariable statistical analyses. Results Among the 295 patients, 180 had a poor-prognosis signature and 115 had a good-prognosis signature, and the mean (±SE) overall 10-year survival rates were 54.6±4.4 percent and 94.5±2.6 percent, respectively. At 10 years, the probability of remaining free of distant metastases was 50.6±4.5 percent in the group with a...

/pdf/a-gene-expression-signature-as-a-predictor-of-survival-in-1f6irz2ok6.pdf

A Gene-Expression Signature as a Predictor of Survival in Breast Cancer

In recent years, members of the protein kinase family have been discovered at an accelerated pace. Most were first described, not through the traditional biochemical approach of protein purification and enzyme assay, but as putative protein kinase amino acid sequences deduced from the nucleotide sequences of molecularly cloned genes or complementary DNAs. Phylogenetic mapping of the conserved protein kinase catalytic domains can serve as a useful first step in the functional characterization of these newly identified family members.

http://science.sciencemag.org/content/sci/241/4861/42.full.pdf

The protein kinase family: conserved features and deduced phylogeny of the catalytic domains.

Whether and how human tumours are genetically unstable has been debated for decades. There is now evidence that most cancers may indeed be genetically unstable, but that the instability exists at two distinct levels. In a small subset of tumours, the instability is observed at the nucleotide level and results in base substitutions or deletions or insertions of a few nucleotides. In most other cancers, the instability is observed at the chromosome level, resulting in losses and gains of whole chromosomes or large portions thereof. Recognition and comparison of these instabilities are leading to new insights into tumour pathogenesis.

Genetic instabilities in human cancers

Over the past decade, the Nomenclature Committee on Cell Death (NCCD) has formulated guidelines for the definition and interpretation of cell death from morphological, biochemical, and functional perspectives. Since the field continues to expand and novel mechanisms that orchestrate multiple cell death pathways are unveiled, we propose an updated classification of cell death subroutines focusing on mechanistic and essential (as opposed to correlative and dispensable) aspects of the process. As we provide molecularly oriented definitions of terms including intrinsic apoptosis, extrinsic apoptosis, mitochondrial permeability transition (MPT)-driven necrosis, necroptosis, ferroptosis, pyroptosis, parthanatos, entotic cell death, NETotic cell death, lysosome-dependent cell death, autophagy-dependent cell death, immunogenic cell death, cellular senescence, and mitotic catastrophe, we discuss the utility of neologisms that refer to highly specialized instances of these processes. The mission of the NCCD is to provide a widely accepted nomenclature on cell death in support of the continued development of the field.

https://biblio.ugent.be/publication/8555786/file/8555788.pdf

Molecular mechanisms of cell death: recommendations of the Nomenclature Committee on Cell Death 2018.

Exceptional advances in molecular biology and genetic research have expedited cancer drug development tremendously. The declared paradigm is the development of 'personalized and tailored drugs' that precisely target the specific molecular defects of a cancer patient. It is therefore appropriate to revisit the intellectual foundations of the development of such agents, as many have shown great clinical success. One hundred years ago, Paul Ehrlich, the founder of chemotherapy, received the Nobel Prize for Physiology or Medicine. His postulate of creating 'magic bullets' for use in the fight against human diseases inspired generations of scientists to devise powerful molecular cancer therapeutics.

Paul Ehrlich's magic bullet concept: 100 years of progress.

Eph Receptors and Ligands Comprise Two Major Specificity Subclasses and Are Reciprocally Compartmentalized during Embryogenesis

Human polo-like kinase 1 (PLK1) is essential during mitosis and in the maintenance of genomic stability. PLK1 is overexpressed in human tumours and has prognostic potential in cancer, indicating its involvement in carcinogenesis and its potential as a therapeutic target. The use of different PLK1 inhibitors has increased our knowledge of mitotic regulation and allowed us to assess their ability to suppress tumour growth in vivo. We address the structural features of the kinase domain and the unique polo-box domain of PLK1 that are most suited for drug development and discuss our current understanding of the therapeutic potential of PLK1.

Targeting polo-like kinase 1 for cancer therapy

Polo-like kinase 1 (PLK1), which helps orchestrate key events in the cell cycle, has been preclinically validated as an anticancer target, whereas it seems that the closely related kinases PLK2 and PLK3 might act as tumour suppressors. Strebhardt reviews recent insights into the biology of PLKs, with emphasis on their role in malignant transformation, and examines progress in the development of small-molecule PLK1 inhibitors as anticancer drugs. The polo-like kinase 1 (PLK1) acts in concert with cyclin-dependent kinase 1–cyclin B1 and Aurora kinases to orchestrate a wide range of critical cell cycle events. Because PLK1 has been preclinically validated as a cancer target, small-molecule inhibitors of PLK1 have become attractive candidates for anticancer drug development. Although the roles of the closely related PLK2, PLK3 and PLK4 in cancer are less well understood, there is evidence showing that PLK2 and PLK3 act as tumour suppressors through their functions in the p53 signalling network, which guards the cell against various stress signals. In this article, recent insights into the biology of PLKs will be reviewed, with an emphasis on their role in malignant transformation, and progress in the development of small-molecule PLK1 inhibitors will be examined.

Multifaceted polo-like kinases: drug targets and antitargets for cancer therapy

Polo-like kinases (Plks) play pivotal roles in the regulation of cell cycle progression. Plk1, the best characterized family member among mammalian Plks, strongly promotes the progression of cells through mitosis. Furthermore, Plk1 is found to be overexpressed in a variety of human tumors and its expression correlates with cellular proliferation and prognosis of tumor patients. Although all Plks share two conserved elements, the N-terminal Ser/Thr kinase domain and a highly homologues C-terminal region termed the polo-box motif, their functions diverge considerably. While Plk1 is inhibited by different checkpoint pathways, Plk2 and Plk3 are activated by the spindle checkpoint or the DNA damage checkpoint. Thus, Plk2 and Plk3 seem to inhibit oncogenic transformation. Deregulation of Plk1 activity contributes to genetic instability, which in turn leads to oncogenic transformation. In contrast, Plk2 and Plk3 are involved in checkpoint-mediated cell cycle arrest to ensure genetic stability, thereby inhibiting the accumulation of genetic defects. In this review, we shall discuss the roles of Plks in oncogenesis and Plk1 as a target for therapeutic intervention against cancer.

Klaus Strebhardt

Papers

Paul Ehrlich's magic bullet concept: 100 years of progress.

Eph Receptors and Ligands Comprise Two Major Specificity Subclasses and Are Reciprocally Compartmentalized during Embryogenesis

Targeting polo-like kinase 1 for cancer therapy

Multifaceted polo-like kinases: drug targets and antitargets for cancer therapy

Polo-like kinases and oncogenesis