Publications
48 results found
Broomfield J, Kalofonou M, Franklin S, et al., 2023, Handheld ISFET Lab-on-Chip detection of TMPRSS2-ERG and AR mRNA for prostate cancer prognostics, IEEE Sensors Letters, Vol: 7, Pages: 1-4, ISSN: 2475-1472
Ion-sensitive field-effect transistors (ISFETs) in combination with unmodified complementary metal oxide semiconductors present a point-of-care platform for clinical diagnostics and prognostics. This work illustrates the sensitive and specific detection of two circulating mRNA markers for prostate cancer, the androgen receptor and the TMPRSS2-ERG fusion using a target-specific loop-mediated isothermal amplification method. TMPRSS2-ERG and androgen receptor RNA were detected down to 3x10 1 and 5x10 1 copies respectively in under 30 minutes. Administration of these assays onto the ISFET Lab-on-chip device was successful and the specificity of each marker was corroborated with mRNA extracted from prostate cancer cell lines.
Tripathi P, Gulli C, Broomfield J, et al., 2023, Classification of nucleic acid amplification on ISFET arrays using spectrogram-based neural networks., Computers in Biology and Medicine, Vol: 161, Pages: 1-11, ISSN: 0010-4825
The COVID-19 pandemic has highlighted a significant research gap in the field of molecular diagnostics. This has brought forth the need for AI-based edge solutions that can provide quick diagnostic results whilst maintaining data privacy, security and high standards of sensitivity and specificity. This paper presents a novel proof-of-concept method to detect nucleic acid amplification using ISFET sensors and deep learning. This enables the detection of DNA and RNA on a low-cost and portable lab-on-chip platform for identifying infectious diseases and cancer biomarkers. We show that by using spectrograms to transform the signal to the time-frequency domain, image processing techniques can be applied to achieve the reliable classification of the detected chemical signals. Transformation to spectrograms is beneficial as it makes the data compatible with 2D convolutional neural networks and helps gain significant performance improvement over neural networks trained on the time domain data. The trained network achieves an accuracy of 84% with a size of 30kB making it suitable for deployment on edge devices. This facilitates a new wave of intelligent lab-on-chip platforms that combine microfluidics, CMOS-based chemical sensing arrays and AI-based edge solutions for more intelligent and rapid molecular diagnostics.
Broomfield J, Kalofonou M, Pataillot-Meakin T, et al., 2022, Detection of YAP1 and AR-V7 mRNA for prostate cancer prognosis using an ISFET lab-on-chip platform, ACS Sensors, Vol: 7, Pages: 3389-3398, ISSN: 2379-3694
Prostate cancer (PCa) is the second most common cause of male cancer-related death worldwide. The gold standard of treatment for advanced PCa is androgen deprivation therapy (ADT). However, eventual failure of ADT is common and leads to lethal metastatic castration-resistant PCa. As such, the detection of relevant biomarkers in the blood for drug resistance in metastatic castration-resistant PCa patients could lead to personalized treatment options. mRNA detection is often limited by the low specificity of qPCR assays which are restricted to specialized laboratories. Here, we present a novel reverse-transcription loop-mediated isothermal amplification assay and have demonstrated its capability for sensitive detection of AR-V7 and YAP1 RNA (3 × 101 RNA copies per reaction). This work presents a foundation for the detection of circulating mRNA in PCa on a non-invasive lab-on-chip device for use at the point-of-care. This technique was implemented onto a lab-on-chip platform integrating an array of chemical sensors (ion-sensitive field-effect transistors) for real-time detection of RNA. Detection of RNA presence was achieved through the translation of chemical signals into electrical readouts. Validation of this technique was conducted with rapid detection (<15 min) of extracted RNA from prostate cancer cell lines 22Rv1s and DU145s.
Wormald BW, Moser N, deSouza NM, et al., 2022, Lab-on-Chip assay of tumour markers and Human Papilloma Virus for cervical cancer detection at the point-of-care, Scientific Reports, Vol: 12, ISSN: 2045-2322
Cervical cancer affects over half a million people worldwide each year, the majority of whom are in resource-limited settingswhere cytology screening is not available. As persistent human papilloma virus (HPV) infections are a key causative factor,detection of HPV strains now complements cytology where screening services exist. This work demonstrates the efficacy ofa handheld Lab-on-Chip (LoC) device, with an external sample extraction process, in detecting cervical cancer from biopsysamples. The device is based on Ion-Sensitive Field-Effect Transistor (ISFET) sensors used in combination with loop-mediatedisothermal amplification (LAMP) assays, to amplify HPV DNA and human telomerase reverse transcriptase (hTERT) mRNA.These markers were selected because of their high levels of expression in cervical cancer cells, but low to nil expression innormal cervical tissue. The achieved analytical sensitivity for the molecular targets resolved down to a single copy per reactionfor the mRNA markers, achieving a limit of detection of 102for hTERT. In the tissue samples, HPV-16 DNA was present in 4/5malignant and 2/5 benign tissues, with HPV-18 DNA being present in 1/5 malignant and 1/5 benign tissues. hTERT mRNA wasdetected in all malignant and no benign tissues, with the demonstrated pilot data to indicate the potential for using the LoC incervical cancer screening in resource-limited settings on a large scale.
Allsopp R, Alexandrou G, Toumazou C, et al., 2022, A comparison between Mini-loop mediated isothermal amplification and polymerase spiral reaction for selective amplification of short template DNA, bioRxiv
Alexandrou G, Moser N, Mantikas K-T, et al., 2021, Detection of Multiple Breast Cancer ESR1 mutations on an ISFET based Lab-on-Chip Platform., IEEE Trans Biomed Circuits Syst, Vol: PP
ESR1 mutations are important biomarkers in metastatic breast cancer. Specifically, p.E380Q and p.Y537S mu- tations arise in response to hormonal therapies given to patients with hormone receptor positive (HR+) breast cancer (BC). This paper demonstrates the efficacy of an ISFET based CMOS integrated Lab-on-Chip (LoC) system, coupled with variant- specific isothermal amplification chemistries, for detection and discrimination of wild type (WT) from mutant (MT) copies of the ESR1 gene. Hormonal resistant cancers often lead to increased chances of metastatic disease which leads to high mortality rates, especially in low-income regions and areas with low healthcare coverage. Design and optimization of bespoke primers was carried out and tested on a qPCR instrument and then benchmarked versus the LoC platform. Assays for detection of p.Y537S and p.E380Q were developed and tested on the LoC platform, achieving amplification in under 25 minutes and sensitivity of down to 1000 copies of DNA per reaction for both target assays. The LoC system hereby presented, is cheaper and smaller than other standard industry equivalent technologies such as qPCR and sequencing. The LoC platform proposed, has the potential to be used at a breast cancer point-of-care testing setting, offering mutational tracking of circulating tumour DNA in liquid biopsies to assist patient stratification and metastatic monitoring.
Li X, Kalofonou M, 2021, Predicting cancer drug response using an adapted deep neural network model, ISSN: 0271-4310
Recent advancements in biotechnology have contributed to the concept of precision oncology through the application of machine learning algorithms. The proposed work focuses on the improvement of a novel Deep Learning model, known as Reference drug-based Deep Neural Network (RefDNN), applied to the prediction of cancer drug response. The model utilizes drug's structure similarity profiles (SSP) to describe the similarity between different reference cancer drugs and uses an SSP vector to weigh the pre-predicted drug response probability obtained by the use of Elastic Net (EN), with the weighted response to be the input of the Deep Neural Network. The prediction performance of RefDNN has been improved by adding a t-distributed stochastic neighbor embedding (t-SNE) based feature extraction estimator, through the integration of gene expression, copy number variants (CNV) and mutation data. This adaptation was used to characterise the model and customize the prediction procedure based on cell line data to provide more precise and time-efficient results. The performance of the proposed system was based on a 5-fold cross validation and was compared to the original RefDNN model, showing significant improvements in accuracy and reduction of the computational processing time.
Jahin M, Fenech-Salerno B, Moser N, et al., 2021, Detection of MGMT methylation status using a Lab-on-Chip compatible isothermal amplification method, 43rd Annual International Conference of the IEEE-Engineering-in-Medicine-and-Biology-Society (IEEE EMBC), Publisher: IEEE, Pages: 7385-7389, ISSN: 1557-170X
Zhang J, Alexandrou G, Toumazou C, et al., 2021, Automating the Design of Cancer Specific DNA Probes Using Computational Algorithms, 43rd Annual International Conference of the IEEE-Engineering-in-Medicine-and-Biology-Society (IEEE EMBC), Publisher: IEEE, Pages: 1852-1856, ISSN: 1557-170X
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Dimitrakopoulos F-ID, Antonacopoulou AG, Kottorou AE, et al., 2021, Genetic Variations of CD40 and LTβR Genes Are Associated With Increased Susceptibility and Clinical Outcome of Non-Small-Cell Carcinoma Patients., Front Oncol, Vol: 11, ISSN: 2234-943X
BACKGROUND: Immune system-related receptors CD40 (tumor necrosis factor receptor superfamily member 5), BAFFR (tumor necrosis factor receptor superfamily member 13C), and LTβR (tumor necrosis factor receptor superfamily member 3) play a pivotal role in non-small-cell lung cancer (NSCLC). To further evaluate their role in NSCLC, CD40 rs1883832 (T>C), BAFFR rs7290134 (A>G), and LTβR rs10849448 (A>G) single-nucleotide polymorphisms (SNPs) were investigated regarding their impact in risk and clinical outcome of NSCLC patients. METHODS: The three selected SNPs were evaluated in 229 NSCLC patients and 299 healthy controls, while CD40, BAFFR, and LTβR protein expression was assessed by immunohistochemistry in 96 tumor specimens from NSCLC patients. RESULTS: In total, CD40 rs1883832 was associated with NSCLC risk, with the T allele, after adjusting for cofactors, being related to increased risk (p = 0.007; OR 1.701). Moreover, the CT genotype was associated with increased risk (p = 0.024; OR 1.606) and poorer 5-year overall survival (OS) after adjusting for cofactors (p = 0.001, HR 1.829), while CC was associated with higher CD40 expression in tumorous cells (p = 0.040) and in stromal cells (p = 0.036). In addition, AA homozygotes for the LTβR rs10849448 had increased risk for NSCLC in multivariate analysis (p = 0.008; OR, 2.106) and higher LTβR membranous expression (p = 0.035). Although BAFFR rs7290134 was associated with BAFFR membranous expression (p = 0.039), BAFFR rs7290134 was not associated with neither the disease risk nor the prognosis of NSCLC patients. CONCLUSIONS: In conclusion, CD40 rs1883832 and LTβR rs10849448 seem to be associated with increased risk for NSCLC, while CD40 rs1883832 is also associated with OS of patients with NSCLC.
Dimitrakopoulos F-ID, Kottorou AE, Kalofonou M, et al., 2020, The fire within: NF-κB involvement in non–small cell lung cancer, Cancer Research, Vol: 80, Pages: 4025-4036, ISSN: 0008-5472
Thirty-four years since its discovery, NF-κB remains a transcription factor with great potential for cancer therapy. However, NF-κB–targeted therapies have yet to find a way to be clinically translatable. Here, we focus exclusively on the role of NF-κB in non-small cell lung cancer (NSCLC) and discuss its contributing effect on cancer hallmarks such as inflammation, proliferation, survival, apoptosis, angiogenesis, epithelial–mesenchymal transition, metastasis, stemness, metabolism, and therapy resistance. In addition, we present our current knowledge of the clinical significance of NF-κB and its involvement in the treatment of patients with NSCLC with chemotherapy, targeted therapies, and immunotherapy.
Alexandrou G, Moser N, Rodriguez-Manzano J, et al., 2020, Detection of breast cancer ESR1 p.E380Q mutation on an ISFET lab-on-chip platform, IEEE International Symposium on Circuits and Systems (ISCAS), Publisher: IEEE, Pages: 1-5, ISSN: 0271-4302
This paper presents a method for detection of ESR1 p.E380Q, a common Breast Cancer (BC) mutation, using an ISFET (Ion-Sensitive Field-Effect Transistor) based Lab-on-Chip (LoC) platform. The LoC contains an ISFET array that can detect pH changes during DNA amplification, specifically Loop-Mediated Isothermal Amplification (LAMP). Synthetic ESR1 DNA was detected in a comparison pH-LAMP assay, carried out on the LoC platform as well as a conventional qPCR instrument. Positive detection of the allele arises due to bespoke allele-specific primers that target one base-pair difference between the wild-type and mutant alleles. The LoC and qPCR demonstrate comparable results detecting the mutant allele with mutant primers in around 25 minutes. The sensing microchip technology coupled with the molecular methods of isothermal chemistries and primer design allow this platform to be tested at a Point-of-Care setting for breast cancer patients, offering mutational tracking platform of circulating tumour DNA in liquid biopsies to assist patient stratification and allow tailored treatments.
Kalofonou M, Malpartida-Cardenas K, Alexandrou G, et al., 2020, A novel hotspot specific isothermal amplification method for detection of thecommon PIK3CA p.H1047R breast cancer mutation, Scientific Reports, Vol: 10, ISSN: 2045-2322
Breast cancer (BC) is a common cancer in women worldwide. Despite advances in treatment, up to 30% of women eventually relapse and die of metastatic breast cancer. Liquid biopsy analysis of circulating cell-free DNA fragments in the patients’ blood can monitor clonality and evolving mutations as a surrogate for tumour biopsy. Next generation sequencing platforms and digital droplet PCR can be used to profile circulating tumour DNA from liquid biopsies; however, they are expensive and time consuming for clinical use. Here, we report a novel strategy with proof-of-concept data that supports the usage of loop-mediated isothermal amplification (LAMP) to detect PIK3CA c.3140 A > G (H1047R), a prevalent BC missense mutation that is attributed to BC tumour growth. Allele-specific primers were designed and optimized to detect the p.H1047R variant following the USS-sbLAMP method. The assay was developed with synthetic DNA templates and validated with DNA from two breast cancer cell-lines and two patient tumour tissue samples through a qPCR instrument and finally piloted on an ISFET enabled microchip. This work sets a foundation for BC mutational profiling on a Lab-on-Chip device, to help the early detection of patient relapse and to monitor efficacy of systemic therapies for personalised cancer patient management.
Alexandrou G, Rodriguez-Manzano J, Malpartida-Cardenas K, et al., 2020, In-silico automated allele-specific primer design for loop-mediated isothermal amplification, IEEE International Symposium on Circuits and Systems (ISCAS), Publisher: IEEE, ISSN: 0271-4302
Dimitrakopoulos F-ID, Antonacopoulou AG, Kottorou AE, et al., 2019, Expression of intracellular components of the NF-κB alternative pathway (NF-κB2, RelB, NIK and Bcl3) is associated with clinical outcome of NSCLC patients, Scientific Reports, Vol: 9, Pages: 14299-14310, ISSN: 2045-2322
A growing number of studies has shed light on the role of the NF-κΒ in non-small-cell lung cancer (NSCLC). To address the significance of major effectors of the NF-κΒ alternative pathway, we investigated the relationship between NF-κΒ2, RelB, NIK and Bcl3 expression (mRNA and protein) and the clinical outcome of NSCLC patients. NF-κΒ2, RelB, NIK and Bcl3 protein expression levels were assessed by immunohistochemistry in tissue samples from 151 NSCLC patients who had curative resection. mRNA levels were also evaluated in 69 patients using quantitative real-time PCR. Although all studied proteins were overexpressed in NSCLC (P < 0.001 for all), only RelB mRNA levels were strongly increased in cancerous specimens compared to tumor-adjacent non-neoplastic tissues (P = 0.009). Moreover, NF-κB2, RelB and Bcl3 expression was associated with overall survival (OS). In particular, cytoplasmic and mRNA expression of RelB was related to 5-year OS (P = 0.014 and P = 0.006, respectively). Multivariate analysis also showed that Bcl3 expression (nuclear and cytoplasmic) was associated with increased 5-year OS (P = 0.002 and P = 0.036, respectively). In addition, higher Bcl3 mRNA levels were associated with inferior OS in stages I & II and improved OS in stages III and IV after 5-year follow-up (P = 0.004 and P = 0.001, respectively). Furthermore, stage I patients with lower NF-κB2 mRNA levels had better 5-year survival in univariate and multivariate analysis (P = 0.031 and P = 0.028, respectively). Interestingly, RelB expression (cytoplasmic and mRNA) was inversely associated with relapse rates (P = 0.027 and P = 0.015, respectively), while low NIK cytoplasmic expression was associated with lower relapse rates (P = 0.019). Cytoplasmic NIK ex
Dimitrakopoulos F-I, Kottorou A, Antonacopoulou A, et al., 2019, Expression of Immune System-Related Membrane Receptors CD40, RANK, BAFFR and LTβR is Associated with Clinical Outcome of Operated Non-Small-Cell Lung Cancer Patients, Journal of Clinical Medicine, Vol: 8, Pages: 741-741, ISSN: 2077-0383
An increasing number of studies implicates the NF-κB (Nuclear Factor of kappa light chain gene enhancer in B cells) alternative pathway in non-small-cell lung cancer (NSCLC). We assessed the clinical significance of CD40 (Tumor necrosis factor receptor superfamily member 5, TNFRSF5), BAFFR (B-cell activating factor receptor), RANK (Receptor activator of NF-κB) and LTβR (lymphotoxin β receptor) receptors, which activate the alternative pathway of NF-κB, in NSCLC. Evaluation of CD40, BAFFR, RANK and LTβR expression was performed based on the Cancer Genome Atlas (TCGA) and the Genotype-Tissue Expression (GTEx) datasets, while protein expression was assessed by immunohistochemistry in specimens from 119 operated NSCLC patients. CD40 gene overexpression was correlated with improved five-year overall survival (OS) (p < 0.001), while increased BAFFR and LTβR mRNA levels were associated with worse OS in patients with adenocarcinomas (p < 0.001 and p < 0.001, respectively). Similarly, patients with adenocarcinomas exhibited a negative correlation between membranous BAFFR protein expression in carcinoma cells and three- and five-year survival (p = 0.021; HR, 4.977 and p = 0.030; HR, 3.358, respectively) as well as between BAFFR protein overexpression in cancer-associated fibroblasts (CAFs) and two-year survival (p = 0.036; HR, 1.983). Patients with increased LTβR nuclear protein staining or stage II patients with lower cytoplasmic LTβR protein expression had worse five-year OS (p = 0.039 and p = 0.008, respectively). Moreover, CD40 protein expression in tumor infiltrating lymphocytes (TILs) and CAFs was positively associated with metastatic spread while BAFFR protein expression in CAFs was negatively associated with bone metastasis (p = 0.041). Our data suggests that CD40, BAFFR, RANK and LTβR play an important role in NSCLC and further supports the role of NF-κB alternative pathway in NSCLC.
Dimitrakopoulos F-I, Kottorou A, Antonacopoulou A, et al., 2019, Association of BAFFR expression in CAFs with overall survival and response to platinum-based chemotherapy in NSCLC, American Society of Clinical Oncology, Publisher: American Society of Clinical Oncology, Pages: 8537-8537, ISSN: 0732-183X
Background: Β-cell activating factor receptor (BAFFR) is a surface receptor, which leads to activation of the Nuclear Factor-kappaB (NF-κB) alternative pathway, a pathway with an important role in non-small cell lung cancer (NSCLC). In addition, cancer associated fibroblasts (CAFs) are major players of the tumor microenvironment promoting NSCLC. The aim of this study was to assess the possible associations of BAFFR expression in CAFs with response to first-line chemotherapy doublet and clinical outcome of NSCLC patients. Methods: Immunohistochemical analysis of BAFFR expression on CAFs was performed on tumor and tumor-adjacent formalin fixed and paraffin embedded tissue samples from 124 operated patients with NSCLC. Patients were under follow-up for at least 60 months, while response to chemotherapy was evaluated in patients who relapsed during this period. Results: BAFFR expression, which was noted exclusively in the cytoplasm of CAFs, was associated with OS only in patients with no infiltration of regional lymph nodes. Higher expression levels of BAFFR in CAFs were related to worse 2-, 3- and 5-year survival (P = 0.015, P = 0.027 and P = 0.040, respectively). This finding persisted after multivariate analysis with age, gender, histological subtype, histological differentiation and disease stage as coefficients (P = 0.009; HR, 2.734; 95% CI, 1.283-5.828). In addition, response to first line chemotherapy was associated with BAFFR expression in CAFs (P = 0.025). Patients who progressed had lower BAFFR levels. Furthermore, BAFFR expression in CAFs was associated with patients’ age. In particular, older patients had higher expression of BAFFR compared to patients younger than 55 years (P = 0.010). Additionally, carcinomas with better differentiation had lower expression of BAFFR in CAFs (P = 0.005). Finally, BAFFR expression in CAFs was related to development of metastatic disease (P = 0.033) and particularly in liver (P = 0,017) and in bones (P = 0.00
Dimitrakopoulos FID, Antonacopoulou A, Kottorou A, et al., 2019, 37P Genetic variation of lymphotoxin beta receptor (LTβR) rs10849448 (A/G) is associated with risk for lung cancer and metastatic spread to adrenals, Annals of Oncology, European Lung Cancer Congress 2019, Publisher: Oxford University Press, Pages: mdz073. 016-mdz073. 016
BackgroundDuring the last years there is an expansion of our knowledge in lung cancer immunology. In addition, there is a growing number of studies on the role of lymphotoxin beta receptor (LTβR), a member of the tumor necrosis factor (TNF) family, which plays an important role in lymphoid system formation and homeostasis as well as in immune system regulation mainly through NF-κB signaling. The aim of the current study was to investigate the clinical relevance of LTβR single nucleotide polymorphism (SNP) rs10849448 (A/G) with the susceptibility to NSCLC, the clinicopathological parameters, the relapse and the survival rates of NSCLC patients, as well as with the protein expression of LTβR.MethodsLTβR SNP was genotyped in 268 randomly selected NSCLC patients and 279 age- and gender-matched healthy donors. Immunohistochemical analysis for LTβR was performed on 127 NSCLC tumors. The studied cohort was under observation during a five-year period.ResultsGenotype frequencies of rs10849448 (AA, AG, and GG) varied between healthy controls and patients, but the difference did not reach statistical significance (P = 0.054). AA homozygotes were found to have lower risk for NSCLC compared to G allele carriers in univariate as well as in multivariate analysis (both P = 0.016). Moreover, rs10849448 was associated with development of metastases with A allele carriers developing less often metastatic disease in adrenals (P = 0.013). Interestingly, rs10849448 was related to membranous LTβR protein expression (P = 0.035) in malignant cells, with AA homozygotes being associated with higher protein levels.ConclusionsThe present findings suggest that the investigated SNP rs10849448 may be associated with NSCLC initiation as well as with the development of metastatic disease. More association and functional studies are needed in order to further clarify it’s role in NSCLC.
Dimitrakopoulos F-ID, Antonacopoulou A, Kottorou A, et al., 2019, Genetic variation of lymphotoxin beta receptor (LTβR) rs10849448 (A/G) is associated with risk for lung cancer and metastatic spread to adrenals., Ann Oncol, Vol: 30 Suppl 2
Zhao Z, Li K, Toumazou C, et al., 2019, A computational model for anti-cancer drug sensitivity prediction, IEEE Biomedical Circuits and Systems Conference (BioCAS), Publisher: IEEE, ISSN: 2163-4025
Khwaja M, Kalofonou M, Toumazou C, 2018, A Deep Autoencoder System for Differentiation of Cancer Types Based on DNA Methylation State, arXiv preprint arXiv:1810.01243
Ma D, Rodriguez-Manzano J, Lopez SDM, et al., 2018, Adapting ISFETs for Epigenetics: An Overview, IEEE Transactions on Biomedical Circuits and Systems, Vol: 12, Pages: 1186-1201, ISSN: 1932-4545
This paper gives an overview of how CMOS design methods can be applied to ion-sensitive field effect transistor (ISFETs) for pH-based DNA methylation and miRNA detection. Design specifications are fundamentally defined by the choice of analysis. As such, the focus for DNA methylation was on developing front-end analogue circuits to carry out Methylation-specific PCR (MSP) for Point-of-Care applications, and sequencing for detailed analysis. The use of MSP prompted the design of an ISFET weak inversion current mirror topology for differential sensing and reduction of drift and temperature sensitivities. The primary limitation in ion-semiconductor sequencing is base calling of repeated nucleotides known as homopolymers. Implementation of a switched current integrator can potentially increase both accuracy and window for detection, within the frequency region of DNA reactions. For quantifying miRNAs, digital back-end processing circuits were considered toward a fully portable platform that can carry out real-time monitoring of DNA amplification reactions. Two systems to evaluate threshold cycles were developed, based on the Derivative method and a new proposed 3-point exponential evaluation aim to reduce detection time simultaneously. Both implementations were tested with datasets from fluorescent qPCR reactions, as well as pH-LAMP experiments that have been optimized for on-chip amplifications. All designs were fabricated in unmodified CMOS with performance assessed based on functionality as well as pH-resolution required in practice.
Hall DA, Kalofonou M, 2018, Guest Editorial ISCAS 2017 Special Issue, IEEE Transactions on Biomedical Circuits and Systems, Vol: 12, Pages: 449-451, ISSN: 1932-4545
Gantier M, Kalofonou M, Toumazou C, 2018, A trapped charge compensation scheme for ISFET based translinear circuits, IEEE International Symposium on Circuits and Systems (ISCAS), Publisher: IEEE
A trapped charge compensation scheme for ISFET based translinear circuits is presented, as part of a system for prediction of cancer risk, based on DNA methylation. Each pixel is able to measure a DNA methylation ratio through pH-based measurements and by using in-pixel comparison to a tunable threshold, to output a result which indicates percentage of methylation used as a cancer score. The developed system was designed in a 0.35 μm CMOS technology and uses a novel trapped charge compensation scheme for ISFETs used in translinear circuits. The output scheme was able to compensate trapped charge of up to 380mV, with a ratio error below 5%, in a range of ratios between 50% and 80% which is generated from pH-based DNA methylation reactions.
Kottorou A, Antonacopoulou A, Dimitrakopoulos F-I, et al., 2018, Deregulation of methylation of transcribed-ultra conserved regions in colorectal cancer and their value for detection of adenomas and adenocarcinomas, Oncotarget, Vol: 9, Pages: 21411-21428, ISSN: 1949-2553
Expression of Transcribed Ultraconserved Regions (T-UCRs) is often deregulated in cancer. The present study assesses the expression and methylation of three T-UCRs (Uc160, Uc283 and Uc346) in colorectal cancer (CRC) and explores the potential of T-UCR methylation in circulating DNA for the detection of adenomas and adenocarcinomas.Expression levels of Uc160, Uc283 and Uc346 were lower in neoplastic tissues from 64 CRC patients (statistically significant for Uc160, p<0.001), compared to non-malignant tissues, while methylation levels displayed the inverse pattern (p<0.001, p=0.001 and p=0.004 respectively). In colon cancer cell lines, overexpression of Uc160 and Uc346 led to increased proliferation and migration rates. Methylation levels of Uc160 in plasma of 50 CRC, 59 adenoma patients, 40 healthy subjects and 12 patients with colon inflammation or diverticulosis predicted the presence of CRC with 35% sensitivity and 89% specificity (p=0.016), while methylation levels of the combination of all three T-UCRs resulted in 45% sensitivity and 74.3% specificity (p=0.013). In conclusion, studied T-UCRs’ expression and methylation status are deregulated in CRC while Uc160 and Uc346 appear to have a complicated role in CRC progression. Moreover their methylation status appears a promising non-invasive screening test for CRC, provided that the sensitivity of the assay is improved.
Dimitrakopoulos F-I, Antonacopoulou A, Kottorou A, et al., 2018, NF-kB2 genetic variations are significantly associated with non-small cell lung cancer risk and overall survival, Scientific Reports, Vol: 8, Pages: 5259-5269, ISSN: 2045-2322
During the last decade, a growing number of publications implicate NF-kB2 in NSCLC pathogenesis. Here, we investigated the clinical relevance of NF-kB2 single nucleotide polymorphisms (SNPs) rs7897947, rs11574852 and rs12769316 in NSCLC and their association with NF-kB2 protein and mRNA levels. Our data show that TT (rs7897947T >G) and AA (rs12769316G >A) genotypes were strongly associated with an increased risk for NSCLC (P = 0.019 and P = 0.003, respectively). Additionally, in multivariate analysis, TT (rs7897947T >G) homozygosity was associated with worse 2- and 3-year survival rates (P = 0.030 and P = 0.028, respectively), especially among patients with stages III/IV, who had worse 2, 3 and 5-year survival (P = 0.001, P = 0.022 and P = 0.035, respectively). In chemotherapy-treated patients, TT (rs12769316G >A) homozygosity was also associated with worse 2- and 3-year survival compared to G allele carriers (P = 0.006 and P = 0.014, respectively). Furthermore, rs12769316 was correlated with survival outcome of stage I and II patients (P = 0.031 and P = 0.006, respectively). Interestingly, amongst the patients who developed metastases, A allele carriers had better 5-year survival (P = 0.020). In addition, rs12769316 was associated with NF-kB2 protein (P = 0.001) and mRNA expression (P = 0.017) as well as with tumor maximum diameter (P = 0.025). Overall, this study suggests that rs7897947 and rs12769316 are involved in NSCLC susceptibility, in treatment response and in clinical outcome.
Khwaja M, Kalofonou M, Toumazou C, 2017, A Deep Belief Network system for prediction of DNA methylation, IEEE Biomedical Circuits and Systems Conference (BioCAS), Publisher: IEEE
A Deep Belief Network architecture is proposed for prediction of DNA methylation characteristics across genetic regions. The proposed system uses an image analogous visualisation of DNA methylation features through an efficient mapping model. Implementation of this method has resulted in an accurate classification of DNA methylation for multiple CpG regions identified in cancer cell lines and has been designed to address variability in patterns found in a given human cell, regardless of their function or disease state. The proposed method is compared to time-tested supervised learning algorithms that include Support Vector Machine and Random Forest classifiers and has been validated using data from cancer cell lines. Using documented features, it achieves differentiation of DNA methylation states, while predicting distinct features with an average value of sensitivity 92%, specificity 99%, accuracy 95% and Matthew's Correlation Coefficient 0.91. The feature set coupled with the deep learning model makes the system efficient for DNA methylation prediction, while being independent of the data set used.
Moser N, Rodriguez-Manzano J, Yu LS, et al., 2017, Live demonstration: A CMOS-based ISFET array for rapid diagnosis of the Zika virus, ISSN: 0271-4310
We demonstrate a diagnostics platform which integrates an ISFET array and a temperature control loop for isothermal DNA detection. The controller maintains a temperature of 63°C to perform nucleic acid amplification which is detected by the on-chip sensors. The 32×32 ISFET array is first calibrated to cancel trapped charge and then measures the change in the pH of the reaction. The sensor data is sent to a microcontroller and the reaction is monitored in real-time using a MATLAB interface. Experiments confirm a change of 0.9 pH when tested for the presence of RNA associated with the Zika virus.
Kottorou A, Antonacopoulou A, Dimitrakopoulos F, et al., 2017, Deregulation of methylation of transcribed-ultra conserved regions in colorectal cancer and their diagnostic and prognostic value, ASCO 2017, Publisher: American Society of Clinical Oncology, Pages: e15130-e15130
Background: Expression of Transcribed Ultra Conserved Regions (T-UCRs) is often deregulated in cancer. We investigated the role of three T-UCRs (Uc160, Uc283 and Uc346) in colorectal adenocarcinomas and their prognostic and diagnostic value. Methods: Expression and methylation levels of the T-UCRs were assessed in neoplastic and paired non-malignant fresh frozen (FF) tissue specimens from 64 colorectal cancer (CRC) patients, as well as in 6 FF adenoma tissue specimens. In addition, T-UCR methylation levels were assessed in FFPE tumor tissues from 80 CRC patients and in plasma from 161 patients (50 CRC, 59 adenoma patients, 40 healthy subjects and 12 patients with colon inflammation or diverticulosis). Results: Expression levels of all three T-UCRs were lower in neoplastic, compared to non-malignant tissues, although at a statistically significant level only for Uc160 (p< 0.001). Also, methylation levels of Uc160, Uc283 and Uc346 were higher in tumors compared to non-malignant tissues (p< 0.001, p= 0.001 and p= 0.004 respectively). Tissue methylation levels of Uc160 were associated with TTP (p= 0.017). The combination of Uc283 and Uc346 methylation levels was related to OS, however without reaching statistical significance (p= 0.066). Methylation status of Uc160 and Uc346 in plasma differed significantly among the four patient groups with CRC patients exhibiting the higher levels. Moreover, a strong correlation was found between Uc160 plasma methylation levels and adenoma or adenocarcinoma size and lymph node infiltration (p< 0.001 and p= 0.024 respectively). When methylation status was used to predict if a subject has CRC, sensitivity and specificity were 35% and 89% respectively, while the values changed to 45% and 74.3% respectively when we combined the sum of the three T-UCR plasma methylation levels. For adenomas, the combination of Uc160 and Uc346 plasma methylation displayed 30.2% (sensitivity) and 80.7% (specificity). Conclusions: T-UCR expression an
Koutsos A, Kalofonou M, Sohbati M, et al., 2016, Epigenetic-IC: A fully integrated sensing platform for epigenetic reaction monitoring, IEEE International Symposium on Circuits and Systems (ISCAS), Publisher: IEEE, Pages: 325-328, ISSN: 2379-447X
This paper presents a pH-based System-on-Chip DNA methylation quantification platform for real time monitoring of DNA methylation ratio in target genes. The architecture forms a novel autonomous system, capable of providing diagnostic information on the progression of a disease, notably cancer. The system is equipped with drift and trapped charge compensation schemes based on differential measurements and an auto-calibration algorithm. The simulated system in 0.35μm CMOS technology achieves a power consumption of 0.997mW, with a DNA methylation ratio output sensitivity of 0.1%. The ISFET-based detection platform occupies a total of 901um2 and allows the calculation of DNA methylation ratio in pH-monitored DNA methylation based reactions.
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