Alain Pierré

Introduction to sol-gel processing

Preface. 1. General Introduction. 2. The Chemistry of Precursors Solutions. 3. Colloidal Particles and Sols. 4. Gelation. 5. Gels. 6. New Types of Sol-Gel Derived Materials. 7. Phase Transformations. 8. Sintering Sol-Gel Ceramics. 9. Applications of Sol-Gel Processing. Appendix.

Characterization of a Large Panel of Patient-Derived Tumor Xenografts Representing the Clinical Heterogeneity of Human Colorectal Cancer

Purpose: Patient-derived xenograft models are considered to represent the heterogeneity of human cancers and advanced preclinical models. Our consortium joins efforts to extensively develop and characterize a new collection of patient-derived colorectal cancer (CRC) models. Experimental Design: From the 85 unsupervised surgical colorectal samples collection, 54 tumors were successfully xenografted in immunodeficient mice and rats, representing 35 primary tumors, 5 peritoneal carcinoses and 14 metastases. Histologic and molecular characterization of patient tumors, first and late passages on mice includes the sequence of key genes involved in CRC (i.e., APC, KRAS, TP53), aCGH, and transcriptomic analysis. Results: This comprehensive characterization shows that our collection recapitulates the clinical situation about the histopathology and molecular diversity of CRC. Moreover, patient tumors and corresponding models are clustering together allowing comparison studies between clinical and preclinical data. Hence, we conducted pharmacologic monotherapy studies with standard of care for CRC (5-fluorouracil, oxaliplatin, irinotecan, and cetuximab). Through this extensive in vivo analysis, we have shown the loss of human stroma cells after engraftment, observed a metastatic phenotype in some models, and finally compared the molecular profile with the drug sensitivity of each tumor model. Through an experimental cetuximab phase II trial, we confirmed the key role of KRAS mutation in cetuximab resistance. Conclusions: This new collection could bring benefit to evaluate novel targeted therapeutic strategies and to better understand the basis for sensitivity or resistance of tumors from individual patients. Clin Cancer Res; 18(19); 5314–28. ©2012 AACR.

Novel protein scaffolds as emerging therapeutic proteins: from discovery to clinical proof-of-concept

Recent advances in combinatorial protein engineering have made it possible to develop immunoglobulin (Ig)-based and non-Ig protein scaffolds that can potentially substitute for most whole antibody-associated properties and currently translate into biologicals with drug-like properties. During the past 10 years, the most validated scaffolds have reached the clinical development phase and, recently, one of them [Kalbitor(®) (Dyax)] has made it to the market, making these alternative scaffold proteins viable drug candidates in a post-antibody landscape. Interestingly, several scaffolds include an immune-active component as part of their therapeutic mode of action, which yielded spectacular clinical efficacy in some hematological malignancies. Here, we review the most recent clinical advances and analyze their benefits for patients.

Encapsulation of lipases in aerogels

Abstract Pseudomonas cepacia (PC) lipase was encapsulated in silica and aluminosilicate gels made by different chemical processes. Some samples were dried by the supercritical method in CO 2 to obtain aerogels, while other ones made by the same wet chemical procedure were dried by evaporation to produce xerogels. The biocatalysts which were obtained were compared in the esterification reaction of 1-octanol by lauric acid. In all cases, the aerogels performed better than the corresponding xerogels. This biocatalysis behavior is discussed in relationship with the textural and structural characteristics of the gels, studied by nitrogen sorption, 29 Si NMR and 27 Al NMR. It is proposed that supercritical drying avoids a simultaneous compression of the gel network and of the enzyme, hence improving the biocatalytic activity.

TTK/hMPS1 is an attractive therapeutic target for triple-negative breast cancer.

Triple-negative breast cancer (TNBC) represents a subgroup of breast cancers (BC) associated with the most aggressive clinical behavior. No targeted therapy is currently available for the treatment of patients with TNBC. In order to discover potential therapeutic targets, we searched for protein kinases that are overexpressed in human TNBC biopsies and whose silencing in TNBC cell lines causes cell death. A cohort including human BC biopsies obtained at Institut Curie as well as normal tissues has been analyzed at a gene-expression level. The data revealed that the human protein kinase monopolar spindle 1 (hMPS1), also known as TTK and involved in mitotic checkpoint, is specifically overexpressed in TNBC, compared to the other BC subgroups and healthy tissues. We confirmed by immunohistochemistry and reverse phase protein array that TNBC expressed higher levels of TTK protein compared to the other BC subgroups. We then determined the biological effects of TTK depletion by RNA interference, through analyses of tumorigenic capacity and cell viability in different human TNBC cell lines. We found that RNAi-mediated depletion of TTK in various TNBC cell lines severely compromised their viability and their ability to form colonies in an anchorage-independent manner. Moreover, we observed that TTK silencing led to an increase in H2AX phosphorylation, activation of caspases 3/7, sub-G1 cell population accumulation and high annexin V staining, as well as to a decrease in G1 phase cell population and an increased aneuploidy. Altogether, these data indicate that TTK depletion in TNBC cells induces apoptosis. These results point out TTK as a protein kinase overexpressed in TNBC that may represent an attractive therapeutic target specifically for this poor prognosis associated subgroup of breast cancer.

Radiosensitization by Chir-124, a selective CHK1 inhibitor: effects of p53 and cell cycle checkpoints.

Checkpoint kinase-1 (CHK1) is a key regulator of the DNA damage-elicited G2-M checkpoints. The aim of the present study was to investigate the effects of a selective CHK1 inhibitor, Chir124, on cell survival and cell cycle progression following ionizing radiation (IR). Treatment with Chir-124 resulted in reduced clonogenic survival and abrogated the IR- induced G2-M arrest in a panel of isogenic HCT116 cell lines after IR. This radiosensitizing effect was relatively similar between p53-/- and p53-sufficient wild type (WT) HCT116 cells. However, the number of mitotic cells (as measured by assessing the phosphorylation of mitotic proteins) increased dramatically in p53-/- HCT116 cells after concomitant Chir-124 exposure, compared to IR alone, while no such effect was observed in p53-sufficient WT HCT116 cells. In p53-/- cells, Chir-124 treatment induced a marked accumulation of polyploid cells that were characterized by micronucleation or multinucleation. p21-/- HCT116 cells displayed a similar pattern of response as p53-/- cells. Chir-124 was able to radiosensitize HCT116 cells that lack checkpoint kinase-2 (CHK2) or that were deficient for the spindle checkpoint protein Mad2. Finally, Chir-124 could radiosensitize tetraploid cell lines, which were relatively resistant against DNA damaging agents. Altogether these results suggest that Chir-124-mediated radiosensitization is profoundly influenced by the p53 and cell cycle checkpoint system.

Synthesis and pharmacological evaluation of novel non-lactone analogues of camptothecin.

Ten novel camptothecin (CPT) derivatives devoid of the lactone function in the E-ring were synthesized and evaluated as anticancer agents. Several of these CPT analogues bearing a five-membered E-ring are potent inhibitors of the DNA relaxation and cleavage reactions catalyzed by topoisomerase I and exhibit promising cytotoxic activities with IC(50) values in the nM range. This is the first successful design of lactone-free CPT, providing thus a new avenue to the development of topoisomerase I targeted antitumor agents.

Alkylation of guanine in DNA by S23906-1, a novel potent antitumor compound derived from the plant alkaloid acronycine

The discovery of a new DNA-targeted antitumor agent is a challenging enterprise, and the elucidation of its mechanism of action is an essential first step in investigating the structural and biological consequences of DNA modification and to guide the rational design of analogues. Here, we have dissected the mode of action of the newly discovered antitumor agent S23906-1. Gel retardation experiments reveal that the diacetate compound S23906-1 and its monoacetate analogue S28687 form highly stable covalent adducts with DNA. The covalent adducts formed between S23906-1 and a 7-bp hairpin oligonucleotide duplex were identified by spectrometry. In contrast, the inactive compound S23907, lacking the two acetate groups of S23906-1, fails to yield covalent DNA adducts, indicating that the C1-C2 functionality is the DNA reactive moiety. DNase I footprinting and DNA alkylation experiments indicate that S23906-1 reacts primarily with guanine residues. A 30-mer oligonucleotide containing only G.C bp forms highly stable complexes with S23906-1 and S28687, whereas the equivalent A.T oligonucleotide is not a good substrate for these two drugs. The use of an oligonucleotide duplex containing inosines instead of guanosines identifies the guanine 2-amino group exposed in the minor groove of DNA as the potential reactive site. The reactivity of S23906-1 toward the guanine-N2 group was independently confirmed by fluorescence spectroscopy. Covalent DNA adducts were also identified in the genomic DNA of B16 melanoma cells exposed to S23906-1, and the specific accumulation of the drug in the nucleus of the cells was visualized by confocal microscopy. The elucidation of the mechanism of action of this highly potent anticancer agent opens a new field for future drug design.

S49076 is a Novel Kinase Inhibitor of MET, AXL and FGFR with Strong Preclinical Activity Alone and in Association with Bevacizumab

Aberrant activity of the receptor tyrosine kinases MET, AXL, and FGFR1/2/3 has been associated with tumor progression in a wide variety of human malignancies, notably in instances of primary or acquired resistance to existing or emerging anticancer therapies. This study describes the preclinical characterization of S49076, a novel, potent inhibitor of MET, AXL/MER, and FGFR1/2/3. S49076 potently blocked cellular phosphorylation of MET, AXL, and FGFRs and inhibited downstream signaling in vitro and in vivo. In cell models, S49076 inhibited the proliferation of MET- and FGFR2-dependent gastric cancer cells, blocked MET-driven migration of lung carcinoma cells, and inhibited colony formation of hepatocarcinoma cells expressing FGFR1/2 and AXL. In tumor xenograft models, a good pharmacokinetic/pharmacodynamic relationship for MET and FGFR2 inhibition following oral administration of S49076 was established and correlated well with impact on tumor growth. MET, AXL, and the FGFRs have all been implicated in resistance to VEGF/VEGFR inhibitors such as bevacizumab. Accordingly, combination of S49076 with bevacizumab in colon carcinoma xenograft models led to near total inhibition of tumor growth. Moreover, S49076 alone caused tumor growth arrest in bevacizumab-resistant tumors. On the basis of these preclinical studies showing a favorable and novel pharmacologic profile of S49076, a phase I study is currently underway in patients with advanced solid tumors. Mol Cancer Ther; 12(9); 1749–62. ©2013 AACR.

Syntheses and antiproliferative activities of rebeccamycin analogues bearing two 7-azaindole moieties

As a part of structure-activity relationship studies on rebeccamycin analogues, compounds containing two aza-indole moieties were synthesized bearing either a methyl group or a hydrogen atom on the imide nitrogen. The azaindole substructures were expected to enhance the cytotoxicity toward tumor cell lines through stronger hydrogen bonding with the target enzyme(s). The cytotoxicities of compounds 8, 10 and 19 against a panel of tumor cell lines were examined and compared with those of rebeccamycin, dechlorinated rebeccamycin 2 and N-methylated analogue A. Their effect on the L1210 cell cycle was also evaluated. Compound 19, having an imide NH function had the strongest cytotoxicity towards L1210 cells and induced the largest accumulation of cells in the G2+M phases of the cell cycle. In contrast to their non-aza analogues, which were cytotoxic for all the cell lines tested, diaza compounds 10 and 19 showed selectivity for some cell lines.