Longitudinal Monitoring of Intra-Tumoural Heterogeneity Using Optical Barcoding of Patient-Derived Colorectal Tumour Models.

Geno- and phenotypic heterogeneity amongst cancer cell subpopulations are established drivers of treatment resistance and tumour recurrence. However, due to the technical difficulty associated with studying such intra-tumoural heterogeneity, this phenomenon is seldom interrogated in conventional cell culture models. Here, we employ a fluorescent lineage technique termed "optical barcoding" (OBC) to perform simultaneous longitudinal tracking of spatio-temporal fate in 64 patient-derived colorectal cancer subclones.

CD44v6 Defines a New Population of Circulating Tumor Cells Not Expressing EpCAM.

Circulating tumor cells (CTCs) are promising diagnostic and prognostic tools for clinical use. In several cancers, including colorectal and breast, the CTC load has been associated with a therapeutic response as well as progression-free and overall survival. However, counting and isolating CTCs remains sub-optimal because they are currently largely identified by epithelial markers such as EpCAM.

CSK-homologous kinase (CHK/MATK) is a potential colorectal cancer tumour suppressor gene epigenetically silenced by promoter methylation.

Hyperactivation of SRC-family protein kinases (SFKs) contributes to the initiation and progression of human colorectal cancer (CRC). Since oncogenic mutations of SFK genes are rare in human CRC, we investigated if SFK hyperactivation is linked to dysregulation of their upstream inhibitors, C-terminal SRC kinase (CSK) and its homolog CSK-homologous kinase (CHK/MATK). We demonstrate that expression of CHK/MATK but not CSK was significantly downregulated in CRC cell lines and primary tumours compared to normal colonic tissue.

Laminin 521 enhances self-renewal via STAT3 activation and promotes tumor progression in colorectal cancer.

Remodeling of basement membrane proteins contributes to tumor progression towards the metastatic stage. One of these proteins, laminin 521 (LN521), sustains embryonic and induced pluripotent stem cell self-renewal, but its putative role in cancer is poorly described. In the present study we found that LN521 promotes colorectal cancer (CRC) cell self-renewal and invasion.

Compromized DNA repair as a basis for identification of cancer radiotherapy patients with extreme radiosensitivity.

A small percentage of cancer radiotherapy patients develop abnormally severe side effects as a consequence of intrinsic radiosensitivity. We analysed the γ-H2AX response to ex-vivo irradiation of peripheral blood lymphocytes (PBL) and plucked eyebrow hair follicles from 16 patients who developed severe late radiation toxicity following radiotherapy, and 12 matched control patients. Longer retention of the γ-H2AX signal and lower colocalization efficiency of repair factors in over-responding patients confirmed that DNA repair in these individuals was compromised.

Radiotherapy for Non-Small Cell Lung Cancer Induces DNA Damage Response in Both Irradiated and Out-of-field Normal Tissues.

Purpose: To study the response of irradiated and out-of-field normal tissues during localized curative intent radiotherapy.

Experimental Design: Sixteen patients with non-small cell lung carcinoma (NSCLC) received 60 Gy in 30 fractions of definitive thoracic radiotherapy with or without concurrent chemotherapy. Peripheral blood lymphocytes (PBL) and eyebrow hairs were sampled prior, during, and after radiotherapy.

Evaluation of Severe Combined Immunodeficiency and Combined Immunodeficiency Pediatric Patients on the Basis of Cellular Radiosensitivity.

Pediatric patients with severe or nonsevere combined immunodeficiency have increased susceptibility to severe, life-threatening infections and, without hematopoietic stem cell transplantation, may fail to thrive. A subset of these patients have the radiosensitive (RS) phenotype, which may necessitate conditioning before hematopoietic stem cell transplantation, and this conditioning includes radiomimetic drugs, which may significantly affect treatment response. To provide statistical criteria for classifying cellular response to ionizing radiation as the measure of functional RS screening, we analyzed the repair capacity and survival of ex vivo irradiated primary skin fibroblasts from five dysmorphic and/or developmentally delayed pediatric patients with severe combined immunodeficiency and combined immunodeficiency.

A pattern of early radiation-induced inflammatory cytokine expression is associated with lung toxicity in patients with non-small cell lung cancer.

Purpose: Lung inflammation leading to pulmonary toxicity after radiotherapy (RT) can occur in patients with non-small cell lung cancer (NSCLC). We investigated the kinetics of RT induced plasma inflammatory cytokines in these patients in order to identify clinical predictors of toxicity.

Experimental Design: In 12 NSCLC patients, RT to 60 Gy (30 fractions over 6 weeks) was delivered; 6 received concurrent chemoradiation (chemoRT) and 6 received RT alone.

Mobilization of viable tumor cells into the circulation during radiation therapy.

Purpose: To determine whether radiation therapy (RT) could mobilize viable tumor cells into the circulation of non-small cell lung cancer (NSCLC) patients.

Methods And Materials: We enumerated circulating tumor cells (CTCs) by fluorescence microscopy of blood samples immunostained with conventional CTC markers. We measured their DNA damage levels using γ-H2AX, a biomarker for radiation-induced DNA double-strand breaks, either by fluorescence-activated cell sorting or by immunofluorescence microscopy.

Crystal structures of the LsrR proteins complexed with phospho-AI-2 and two signal-interrupting analogues reveal distinct mechanisms for ligand recognition.

Quorum sensing (QS) is a cell-to-cell communication system responsible for a variety of bacterial phenotypes including virulence and biofilm formation. QS is mediated by small molecules, autoinducers (AIs), including AI-2 that is secreted by both Gram-positive and -negative microbes. LsrR is a key transcriptional regulator that governs the varied downstream processes by perceiving AI-2 signal, but its activation via autoinducer-binding remains poorly understood.

AI-2 analogs and antibiotics: a synergistic approach to reduce bacterial biofilms.

Quorum sensing (QS), the process of autoinducer-mediated cell-cell signaling among bacteria, facilitates biofilm formation, virulence, and many other multicellular phenotypes. QS inhibitors are being investigated as antimicrobials because of their potential to reduce symptoms of infectious disease while slowing the emergence of resistant strains. Autoinducer-2 (AI-2) analogs have been shown to inhibit genotypic QS responses among many bacteria.

Altering the communication networks of multispecies microbial systems using a diverse toolbox of AI-2 analogues.

There have been intensive efforts to find small molecule antagonists for bacterial quorum sensing (QS) mediated by the "universal" QS autoinducer, AI-2. Previous work has shown that linear and branched acyl analogues of AI-2 can selectively modulate AI-2 signaling in bacteria. Additionally, LsrK-dependent phosphorylated analogues have been implicated as the active inhibitory form against AI-2 signaling.

Tumour targeting of Auger emitters using DNA ligands conjugated to octreotate.

Purpose: The objective of the study was to conjugate the DNA binding ligand para-[(125)I]-iodoHoechst to octreotate, and to explore the tumour targeting potential of this conjugate in the octreotate-somatostatin receptor system.

Methods: We synthesized a Hoechst analogue containing a tri-butylstannyl group in the para position of phenyl ring, conjugated it to the N-terminal amino group of octreotate and prepared (125)I-labelled conjugate by iododestannylation. We used the somatostatin receptor (SSTR2) over-expressing cell line A427-7 derived from its parent A427 human non-small cell lung carcinoma cell line to investigate SSTR2 affinity and receptor-mediated internalisation of the conjugate, and the mouse A427-7 tumour xenograft model for in vivo biodistribution studies of the radiolabelled conjugate.

Paradigm shift in discovering next-generation anti-infective agents: targeting quorum sensing, c-di-GMP signaling and biofilm formation in bacteria with small molecules.

Small molecules that can attenuate bacterial toxin production or biofilm formation have the potential to solve the bacteria resistance problem. Although several molecules, which inhibit bacterial cell-to-cell communication (quorum sensing), biofilm formation and toxin production, have been discovered, there is a paucity of US FDA-approved drugs that target these processes. Here, we review the current understanding of quorum sensing in important pathogens such as Pseudomonas aeruginosa, Escherichia coli and Staphylococcus aureus and provide examples of experimental molecules that can inhibit both known and unknown targets in bacterial virulence factor production and biofilm formation.

Synthetic analogs tailor native AI-2 signaling across bacterial species.

The widespread use of antibiotics and the emergence of resistant strains call for new approaches to treat bacterial infection. Bacterial cell-cell communication or "quorum sensing" (QS) is mediated by "signatures" of small molecules that represent targets for "quenching" communication and avoiding virulent phenotypes. Only a handful of small molecules that antagonize the action of the "universal" autoinducer, AI-2, have been reported.

Biological screening of a diverse set of AI-2 analogues in Vibrio harveyi suggests that receptors which are involved in synergistic agonism of AI-2 and analogues are promiscuous.

C1-alkyl AI-2 analogues do not induce bioluminescence in V. harveyi on their own but enhance the bioluminescence induced by AI-2 in a synergistic fashion. A new facile synthesis of AI-2 facilitates the synthesis of a diverse set of AI-2 analogues and biological screening suggests that receptors that are involved in the synergistic bioluminescence production in V.