Electrochemical biosensors for analysis of DNA point mutations in cancer research

Abstract

Cancer is a genetic disease induced by mutations in DNA, in particular point mutations in important driver genes that lead to protein malfunctioning and ultimately to tumorigenesis. Screening for the most common DNA point mutations, especially in such genes as TP53, BRCA1 and BRCA2, EGFR, KRAS, or BRAF, is crucial to determine predisposition risk for cancer or to predict response to therapy. In this review, we briefly depict how these genes are involved in cancer, followed by a description of the most common techniques routinely applied for their analysis, including high-throughput next-generation sequencing technology and less expensive low-throughput options, such as real-time PCR, restriction fragment length polymorphism, or high resolution melting analysis. We then introduce benefits of electrochemical biosensors as interesting alternatives to the standard methods in terms of cost, speed, and simplicity. We describe most common strategies involved in electrochemical biosensing of point mutations, relying mostly on PCR or isothermal amplification techniques, and critically discuss major challenges and obstacles that, until now, prevented their more widespread application in clinical settings.

Advanced technologies towards improved HPV diagnostics

“…EC methods provide an option for inexpensive, simple, and miniaturized instrumentation with the possibility of parallel measurements at electrode chips and arrays while allowing rapid and highly sensitive determinations. [76][77][78] For instance, a diagnostic Clinichip HPV test that employed LAMP and electrochemical DNA chip could recognize 13 clinically relevant HPV types in less than 3 h. In that study, 247 Japanese women, including 109 with normal cytology, 43 with cervical intraepithelial neoplasia of grade 1 (CIN1), 60 with CIN2/3, and 35 with invasive cervical cancer were tested for carcinogenic HPV genotypes, reaching good agreement with direct sequencing. 68 Our team has published several research articles reporting fast and simple LAMP-based assays combined with electrochemical detection (EC-LAMP), targeting HPV16/18 genotypes in both cervical cell lines and cervical smears from women with precancerous lesions (Figure 1).…”

“…Electrochemical (EC) end‐point detection is another suitable technique used in combination with LAMP. EC methods provide an option for inexpensive, simple, and miniaturized instrumentation with the possibility of parallel measurements at electrode chips and arrays while allowing rapid and highly sensitive determinations 76–78 . For instance, a diagnostic Clinichip HPV test that employed LAMP and electrochemical DNA chip could recognize 13 clinically relevant HPV types in less than 3 h. In that study, 247 Japanese women, including 109 with normal cytology, 43 with cervical intraepithelial neoplasia of grade 1 (CIN1), 60 with CIN2/3, and 35 with invasive cervical cancer were tested for carcinogenic HPV genotypes, reaching good agreement with direct sequencing 68 .…”

Electrochemical Analysis of Clinical Samples – A Path to Translational Research

“…V genu BRAF je aktuálně nejvíce klinicky významná mutace V600E, způsobující na proteinové úrovni aminokyselinovou záměnu valinu za kyselinu glutamovou. Tato mutace se objevuje u více než 85 % případů maligního melanomu a více než 8 % případů kolorektálního karcinomu a má významný vliv na výběr terapeutického přístupu 18,19 . Jeden z projektů naší výzkumné skupiny je zaměřen na vývoj rychlé a snadno proveditelné metody detekce bodových mutací těchto genů 20 .…”

Strategies and Applications of Graphene and Its Derivatives-Based Electrochemical Sensors in Cancer Diagnosis

“…For protein biomarker detection, graphene-based electrochemical immunosensors have been developed for the sensitive detection of common cancer proteins like prostate specific antigen (PSA) [ 30 ], carcinoembryonic antigen (CEA) [ 31 ], and cancer antigen 125 (CA125) in clinical samples [ 32 ]. Graphene-modified electrochemical genosensors have demonstrated capabilities for detecting cancer-related DNA mutations such as in BRAF [ 33 ], KRAS [ 34 ], and EGFR [ 35 ] genes in liquid biopsy and patient tissue samples [ 36 ]. Aptamer-based graphene biosensors have shown promise in the detection of cancer biomarkers like vascular endothelial growth factor (VEGF) [ 37 ] and thrombin [ 38 ] down to picomolar levels.…”