Investigation of the interaction between non-small cell lung cancer cells and immortalised normal bronchial epithelial cells

Non-small cell lung cancer consists of a diverse range of molecular and pathological features. This may be due in part to the critical interaction between normal and lung cancer cells. Consequently resulting in ‘normal’ cells acting in a malignant fashion. This project aims to identify pathways responsible for this altered ‘normal’ behaviour.

CDCA3 regulates the cell cycle and modulates cisplatin sensitivity in non-small cell lung cancer

Cisplatin-based regimens are currently the most effective chemotherapy for non-small cell lung cancer (NSCLC). Cisplatin forms DNA crosslinks to stall DNA replication and induce apoptosis. However, intrinsic and acquired chemoresistance is a major therapeutic problem. We have identified ‘cell division cycle associated protein 3’ (CDCA3) as a novel protein that may prove useful in delaying or preventing cisplatin resistance in NSCLC. CDCA3 functions as part of an ubiquitin ligase complex to degrade the endogenous cell cycle inhibitors. While a role for CDCA3 in disease is emerging with elevated expression noted in oral squamous cell carcinoma, little else is known about CDCA3 or whether this protein may prove useful clinically.

Inflammatory meditated mechanisms of cisplatin resistance in non-small cell lung cancer

The majority of non-small cell lung cancer (NSCLC) patients present with advanced stage disease, where chemotherapy is usually the most common treatment option. While somewhat effective, patients treated with cisplatin-based chemotherapy will eventually develop resistance. Multiple pathways have been implicated in chemo-resistance, however the critical underlying mechanisms have yet to be elucidated. The aim of this project is to determine the role of inflammatory mediators in cisplatin resistance.

Non-Gaussian Cosmological Perturbations from Hybrid Inflation and Preheating

This thesis consists of four research papers and an introduction providing some background. The structure in the universe is generally considered to originate from quantum fluctuations in the very early universe. The standard lore of cosmology states that the primordial perturbations are almost scale-invariant, adiabatic, and Gaussian. A snapshot of the structure from the time when the universe became transparent can be seen in the cosmic microwave background (CMB). For a long time mainly the power spectrum of the CMB temperature fluctuations has been used to obtain observational constraints, especially on deviations from scale-invariance and pure adiabacity. Non-Gaussian perturbations provide a novel and very promising way to test theoretical predictions. They probe beyond the power spectrum, or two point correlator, since non-Gaussianity involves higher order statistics. The thesis concentrates on the non-Gaussian perturbations arising in several situations involving two scalar fields, namely, hybrid inflation and various forms of preheating. First we go through some basic concepts -- such as the cosmological inflation, reheating and preheating, and the role of scalar fields during inflation -- which are necessary for the understanding of the research papers. We also review the standard linear cosmological perturbation theory. The second order perturbation theory formalism for two scalar fields is developed. We explain what is meant by non-Gaussian perturbations, and discuss some difficulties in parametrisation and observation. In particular, we concentrate on the nonlinearity parameter. The prospects of observing non-Gaussianity are briefly discussed. We apply the formalism and calculate the evolution of the second order curvature perturbation during hybrid inflation. We estimate the amount of non-Gaussianity in the model and find that there is a possibility for an observational effect. The non-Gaussianity arising in preheating is also studied. We find that the level produced by the simplest model of instant preheating is insignificant, whereas standard preheating with parametric resonance as well as tachyonic preheating are prone to easily saturate and even exceed the observational limits. We also mention other approaches to the study of primordial non-Gaussianities, which differ from the perturbation theory method chosen in the thesis work.

Psychoacoustic sampling as a reliable, non-invasive method to monitor orthopteran species diversity in tropical forests

We evaluated trained listener-based acoustic sampling as a reliable and non-invasive method for rapid assessment of ensiferan species diversity in tropical evergreen forests. This was done by evaluating the reliability of identification of species and numbers of calling individuals using psychoacoustic experiments in the laboratory and by comparing psychoacoustic sampling in the field with ambient noise recordings made at the same time. The reliability of correct species identification by the trained listener was 100% for 16 out of 20 species tested in the laboratory. The reliability of identifying the numbers of individuals correctly was 100% for 13 out of 20 species. The human listener performed slightly better than the instrument in detecting low frequency and broadband calls in the field, whereas the recorder detected high frequency calls with greater probability. To address the problem of pseudoreplication during spot sampling in the field, we monitored the movement of calling individuals using focal animal sampling. The average distance moved by calling individuals for 17 out of 20 species was less than 1.5 m in half an hour. We suggest that trained listener-based sampling is preferable for crickets and low frequency katydids, whereas broadband recorders are preferable for katydid species with high frequency calls for accurate estimation of ensiferan species richness and relative abundance in an area.

On the maximal rate of non-square STBCs from complex orthogonal designs

A Linear Processing Complex Orthogonal Design (LPCOD) is a p x n matrix epsilon, (p >= n) in k complex indeterminates x(1), x(2),..., x(k) such that (i) the entries of epsilon are complex linear combinations of 0, +/- x(i), i = 1,..., k and their conjugates, (ii) epsilon(H)epsilon = D, where epsilon(H) is the Hermitian (conjugate transpose) of epsilon and D is a diagonal matrix with the (i, i)-th diagonal element of the form l(1)((i))vertical bar x(1)vertical bar(2) + l(2)((i))vertical bar x(2)vertical bar(2)+...+ l(k)((i))vertical bar x(k)vertical bar(2) where l(j)((i)), i = 1, 2,..., n, j = 1, 2,...,k are strictly positive real numbers and the condition l(1)((i)) = l(2)((i)) = ... = l(k)((i)), called the equal-weights condition, holds for all values of i. For square designs it is known. that whenever a LPCOD exists without the equal-weights condition satisfied then there exists another LPCOD with identical parameters with l(1)((i)) = l(2)((i)) = ... = l(k)((i)) = 1. This implies that the maximum possible rate for square LPCODs without the equal-weights condition is the same as that or square LPCODs with equal-weights condition. In this paper, this result is extended to a subclass of non-square LPCODs. It is shown that, a set of sufficient conditions is identified such that whenever a non-square (p > n) LPCOD satisfies these sufficient conditions and do not satisfy the equal-weights condition, then there exists another LPCOD with the same parameters n, k and p in the same complex indeterminates with l(1)((i)) = l(2)((i)) = ... = l(k)((i)) = 1.

Afatinib versus gefitinib as first-line treatment of patients with EGFR mutation-positive non-small-cell lung cancer (LUX-Lung 7): a phase 2B, open-label, randomised controlled trial

Background The irreversible ErbB family blocker afatinib and the reversible EGFR tyrosine kinase inhibitor gefitinib are approved for first-line treatment of EGFR mutation-positive non-small-cell lung cancer (NSCLC). We aimed to compare the efficacy and safety of afatinib and gefitinib in this setting. Methods This multicentre, international, open-label, exploratory, randomised controlled phase 2B trial (LUX-Lung 7) was done at 64 centres in 13 countries. Treatment-naive patients with stage IIIB or IV NSCLC and a common EGFR mutation (exon 19 deletion or Leu858Arg) were randomly assigned (1:1) to receive afatinib (40 mg per day) or gefitinib (250 mg per day) until disease progression, or beyond if deemed beneficial by the investigator. Randomisation, stratified by EGFR mutation type and status of brain metastases, was done centrally using a validated number generating system implemented via an interactive voice or web-based response system with a block size of four. Clinicians and patients were not masked to treatment allocation; independent review of tumour response was done in a blinded manner. Coprimary endpoints were progression-free survival by independent central review, time-to-treatment failure, and overall survival. Efficacy analyses were done in the intention-to-treat population and safety analyses were done in patients who received at least one dose of study drug. This ongoing study is registered with ClinicalTrials.gov, number NCT01466660. Findings Between Dec 13, 2011, and Aug 8, 2013, 319 patients were randomly assigned (160 to afatinib and 159 to gefitinib). Median follow-up was 27·3 months (IQR 15·3–33·9). Progression-free survival (median 11·0 months [95% CI 10·6–12·9] with afatinib vs 10·9 months [9·1–11·5] with gefitinib; hazard ratio [HR] 0·73 [95% CI 0·57–0·95], p=0·017) and time-to-treatment failure (median 13·7 months [95% CI 11·9–15·0] with afatinib vs 11·5 months [10·1–13·1] with gefitinib; HR 0·73 [95% CI 0·58–0·92], p=0·0073) were significantly longer with afatinib than with gefitinib. Overall survival data are not mature. The most common treatment-related grade 3 or 4 adverse events were diarrhoea (20 [13%] of 160 patients given afatinib vs two [1%] of 159 given gefitinib) and rash or acne (15 [9%] patients given afatinib vs five [3%] of those given gefitinib) and liver enzyme elevations (no patients given afatinib vs 14 [9%] of those given gefitinib). Serious treatment-related adverse events occurred in 17 (11%) patients in the afatinib group and seven (4%) in the gefitinib group. Ten (6%) patients in each group discontinued treatment due to drug-related adverse events. 15 (9%) fatal adverse events occurred in the afatinib group and ten (6%) in the gefitinib group. All but one of these deaths were considered unrelated to treatment; one patient in the gefitinib group died from drug-related hepatic and renal failure. Interpretation Afatinib significantly improved outcomes in treatment-naive patients with EGFR-mutated NSCLC compared with gefitinib, with a manageable tolerability profile. These data are potentially important for clinical decision making in this patient population.

Clinician perspective on molecular profiling of non-small-cell lung cancer

The article by Meric-Bernstam et al1 that was recently published in Journal of Clinical Oncology raises important questions about the clinical application of large-scale genomic testing. We congratulate the authors for this ambitious study, which successfully profiled 2,000 consecutive patients with advanced cancer. The next-generation sequencing (NGS) platform was used for 1,749 of 2,000 patients (87.5%). Of 789 patients with potentially actionable mutations, 83 (11%, or 4% of screened population) were enrolled in a genomically matched clinical study. As the editorial2 accompanying the article by Meric-Bernstam et al1 pointed out, the 4% figure, albeit disappointing, may be an underestimate because cancers such as lung adenocarcinoma and melanoma, for which ≥ 50% of patients have actionable mutations, were under-represented. ...

Cilengitide combined with cetuximab and platinum-based chemotherapy as first-line treatment in advanced non-small-cell lung cancer (NSCLC) patients: Results of an open-label, randomized, controlled phase II study (CERTO)

Background: This multicentre, open-label, randomized, controlled phase II study evaluated cilengitide in combination with cetuximab and platinum-based chemotherapy, compared with cetuximab and chemotherapy alone, as first-line treatment of patients with advanced non-small-cell lung cancer (NSCLC). Patients and methods: Patients were randomized 1:1:1 to receive cetuximab plus platinum-based chemotherapy alone (control), or combined with cilengitide 2000 mg 1×/week i.v. (CIL-once) or 2×/week i.v. (CIL-twice). A protocol amendment limited enrolment to patients with epidermal growth factor receptor (EGFR) histoscore ≥200 and closed the CIL-twice arm for practical feasibility issues. Primary end point was progression-free survival (PFS; independent read); secondary end points included overall survival (OS), safety, and biomarker analyses. A comparison between the CIL-once and control arms is reported, both for the total cohorts, as well as for patients with EGFR histoscore ≥200. Results: There were 85 patients in the CIL-once group and 84 in the control group. The PFS (independent read) was 6.2 versus 5.0 months for CIL-once versus control [hazard ratio (HR) 0.72; P = 0.085]; for patients with EGFR histoscore ≥200, PFS was 6.8 versus 5.6 months, respectively (HR 0.57; P = 0.0446). Median OS was 13.6 for CIL-once versus 9.7 months for control (HR 0.81; P = 0.265). In patients with EGFR ≥200, OS was 13.2 versus 11.8 months, respectively (HR 0.95; P = 0.855). No major differences in adverse events between CIL-once and control were reported; nausea (59% versus 56%, respectively) and neutropenia (54% versus 46%, respectively) were the most frequent. There was no increased incidence of thromboembolic events or haemorrhage in cilengitide-treated patients. αvβ3 and αvβ5 expression was neither a predictive nor a prognostic indicator. Conclusions: The addition of cilengitide to cetuximab/chemotherapy indicated potential clinical activity, with a trend for PFS difference in the independent-read analysis. However, the observed inconsistencies across end points suggest additional investigations are required to substantiate a potential role of other integrin inhibitors in NSCLC treatment.

VEGF-mediated cell survival in non-small-cell lung cancer: implications for epigenetic targeting of VEGF receptors as a therapeutic approach

Aims: To evaluate the potential therapeutic utility of histone deacetylase inhibitors (HDACi) in targeting VEGF receptors in non-small-cell lung cancer. Materials & methods: Non-small-cell lung cancer cells were screened for the VEGF receptors at the mRNA and protein levels, while cellular responses to various HDACi were examined. Results: Significant effects on the regulation of the VEGF receptors were observed in response to HDACi. These were associated with decreased secretion of VEGF, decreased cellular proliferation and increased apoptosis which could not be rescued by addition of exogenous recombinant VEGF. Direct remodeling of the VEGFR1 and VEGFR2 promoters was observed. In contrast, HDACi treatments resulted in significant downregulation of the Neuropilin receptors. Conclusion: Epigenetic targeting of the Neuropilin receptors may offer an effective treatment for lung cancer patients in the clinical setting.

Mutational analysis of the insulin‑like growth factor 1 receptor tyrosine kinase domain in non-small cell lung cancer patients

The insulin‑like growth factor 1 receptor (IGF1R) pathway plays an important role in the pathogenesis of non‑small cell lung cancer (NSCLC) and also provides a mechanism of resistance to targeted therapies. IGF1R is therefore an ideal therapeutic target and several inhibitors have entered clinical trials. However, thus far the response to these inhibitors has been poor, highlighting the importance of predictive biomarkers to identify patient cohorts who will benefit from these targeted agents. It is well‑documented that mutations and/or deletions in the epidermal growth factor receptor (EGFR) tyrosine kinase (TK) domain predict sensitivity of NSCLC patients to EGFR TK inhibitors. Single‑nucleotide polymorphisms (SNPs) in the IGF pathway have been associated with disease, including breast and prostate cancer. The aim of the present study was to elucidate whether the IGF1R TK domain harbours SNPs, somatic mutations or deletions in NSCLC patients and correlates the mutation status to patient clinicopathological data and prognosis. Initially 100 NSCLC patients were screened for mutations/deletions in the IGF1R TK domain (exons 16‑21) by sequencing analysis. Following the identification of SNP rs2229765, a further 98 NSCLC patients and 866 healthy disease‑free control patients were genotyped using an SNP assay. The synonymous SNP (rs2229765) was the only aberrant base change identified in the IGF1R TK domain of 100 NSCLC patients initially analysed. SNP rs2229765 was detected in exon 16 and was found to have no significant association between IGF1R expression and survival. The GA genotype was identified in 53.5 and 49.4% of NSCLC patients and control individuals, respectively. No significant difference was found in the genotype (P=0.5487) or allele (P=0.9082) frequencies between the case and control group. The present findings indicate that in contrast to the EGFR TK domain, the IGF1R TK domain is not frequently mutated in NSCLC patients. The synonymous SNP (rs2229765) had no significant association between IGF1R expression and survival in the cohort of NSCLC patients.

2nd ESMO Consensus Conference in Lung Cancer: Locally advanced stage III non-small-cell lung cancer

To complement the existing treatment guidelines for all tumour types, ESMO organises consensus conferences to focus on specific issues in each type of tumour. The 2nd ESMO Consensus Conference on Lung Cancer was held on 11–12 May 2013 in Lugano. A total of 35 experts met to address several questions on non-small-cell lung cancer (NSCLC) in each of four areas: pathology and molecular biomarkers, first-line/second and further lines of treatment in advanced disease, early-stage disease and locally advanced disease. For each question, recommendations were made including reference to the grade of recommendation and level of evidence. This consensus paper focuses on locally advanced disease.

Non-Gaussianities and Preheating from N-flation and Magnetogenesis via rotating cosmic string loops

In this thesis we examine multi-field inflationary models of the early Universe. Since non-Gaussianities may allow for the possibility to discriminate between models of inflation, we compute deviations from a Gaussian spectrum of primordial perturbations by extending the delta-N formalism. We use N-flation as a concrete model; our findings show that these models are generically indistinguishable as long as the slow roll approximation is still valid. Besides computing non-Guassinities, we also investigate Preheating after multi-field inflation. Within the framework of N-flation, we find that preheating via parametric resonance is suppressed, an indication that it is the old theory of preheating that is applicable. In addition to studying non-Gaussianities and preheatng in multi-field inflationary models, we study magnetogenesis in the early universe. To this aim, we propose a mechanism to generate primordial magnetic fields via rotating cosmic string loops. Magnetic fields in the micro-Gauss range have been observed in galaxies and clusters, but their origin has remained elusive. We consider a network of strings and find that rotating cosmic string loops, which are continuously produced in such networks, are viable candidates for magnetogenesis with relevant strength and length scales, provided we use a high string tension and an efficient dynamo.