Search based algorithms for test sequence generation in functional testing

Information and Software Technology (DOI: 10.1016/j.infsof.2014.07.014)The generation of dynamic test sequences from a formal specification, complementing traditional testing methods in order to find errors in the source code. Objective In this paper we extend one specific combinatorial test approach, the Classification Tree Method (CTM), with transition information to generate test sequences. Although we use CTM, this extension is also possible for any combinatorial testing method. Method The generation of minimal test sequences that fulfill the demanded coverage criteria is an NP-hard problem. Therefore, search-based approaches are required to find such (near) optimal test sequences. Results The experimental analysis compares the search-based technique with a greedy algorithm on a set of 12 hierarchical concurrent models of programs extracted from the literature. Our proposed search-based approaches (GTSG and ACOts) are able to generate test sequences by finding the shortest valid path to achieve full class (state) and transition coverage. Conclusion The extended classification tree is useful for generating of test sequences. Moreover, the experimental analysis reveals that our search-based approaches are better than the greedy deterministic approach, especially in the most complex instances. All presented algorithms are actually integrated into a professional tool for functional testing.Spanish Ministry of Economy and Competitiveness and FEDER under contract TIN2011-28194 and fellowship BES-2012-055967. Project 8.06/5.47.4142 in collaboration with the VSB-Tech. Univ. of Ostrava, Universidad de Málaga, Andalucía Tech. and EU Grant ICT-257574 (FITTEST project)

Inhibition of human papilloma virus E2 DNA binding protein by covalently linked polyamides

Polyamides are a class of heterocyclic small molecules with the potential of controlling gene expression by binding to the minor groove of DNA in a sequence-specific manner. To evaluate the feasibility of this class of compounds as antiviral therapeutics, molecules were designed to essential sequence elements occurring numerous times in the HPV genome. This sequence element is bound by a virus-encoded transcription and replication factor E2, which binds to a 12 bp recognition site as a homodimeric protein. Here, we take advantage of polyamide:DNA and E2:DNA co-crystal structural information and advances in polyamide synthetic chemistry to design tandem hairpin polyamides that are capable of displacing the major groove-binding E2 homodimer from its DNA binding site. The binding of tandem hairpin polyamides and the E2 DNA binding protein to the DNA site is mutually exclusive even though the two ligands occupy opposite faces of the DNA double helix. We show with circular permutation studies that the tandem hairpin polyamide prevents the intrinsic bending of the E2 DNA site important for binding of the protein. Taken together, these results illustrate the feasibility of inhibiting the binding of homodimeric, major groove-binding transcription factors by altering the local DNA geometry using minor groove-binding tandem hairpin polyamides

Inhibition of the Sec61 translocon overcomes cytokine-induced glucocorticoid resistance in T-cell acute lymphoblastic leukaemia.

Glucocorticoid (GC) resistance is a poor prognostic factor in T-cell acute lymphoblastic leukaemia (T-ALL). Interleukin-7 (IL-7) mediates GC resistance via GC-induced upregulation of IL-7 receptor (IL-7R) expression, leading to increased pro-survival signalling. IL-7R reaches the cell surface via the secretory pathway, so we hypothesized that inhibiting the translocation of IL-7R into the secretory pathway would overcome GC resistance. Sec61 is an endoplasmic reticulum (ER) channel that is required for insertion of polypeptides into the ER. Here, we demonstrate that KZR-445, a novel inhibitor of Sec61, potently attenuates the dexamethasone (DEX)-induced increase in cell surface IL-7R and overcomes IL-7-induced DEX resistance

Synthesis, in Vitro Covalent Binding Evaluation, and Metabolism of ¹⁴C‑Labeled Inhibitors of 11β-HSD1

In this letter, we reported the design and synthesis of three potent, selective, and orally bioavailable 11β-HSD1 inhibitors labeled with ¹⁴C: AMG 456 (<b>1</b>), AM-6949 (<b>2</b>), and AM-7715 (<b>3</b>). We evaluated the covalent protein binding of the labeled inhibitors in human liver microsomes in vitro and assessed their potential bioactivation risk in humans. We then studied the in vitro mechanism of <b>2</b> in human hepatocytes and the formation of reactive intermediates. Our study results suggest that <b>1</b> and <b>3</b> have low potential for metabolic bioactivation in humans, while <b>2</b> has relatively high risk

Signal peptide mimicry primes Sec61 for client-selective inhibition.

Preventing the biogenesis of disease-relevant proteins is an attractive therapeutic strategy, but attempts to target essential protein biogenesis factors have been hampered by excessive toxicity. Here we describe KZR-8445, a cyclic depsipeptide that targets the Sec61 translocon and selectively disrupts secretory and membrane protein biogenesis in a signal peptide-dependent manner. KZR-8445 potently inhibits the secretion of pro-inflammatory cytokines in primary immune cells and is highly efficacious in a mouse model of rheumatoid arthritis. A cryogenic electron microscopy structure reveals that KZR-8445 occupies the fully opened Se61 lateral gate and blocks access to the lumenal plug domain. KZR-8445 binding stabilizes the lateral gate helices in a manner that traps select signal peptides in the Sec61 channel and prevents their movement into the lipid bilayer. Our results establish a framework for the structure-guided discovery of novel therapeutics that selectively modulate Sec61-mediated protein biogenesis