Abstract
In the last century, the knowledge of molecular biology has increased, opening new opportunities for addressing different diseases that have been affecting all populations. Before, genome and proteomic studies were seen as complicated field due to inability to get significance understanding of disease components at the molecular level. However, with the expansion of molecular biology, different techniques and tools have been developed to aid diagnosis and treatment of diseases. In this study, these tools have been investigated to understand how the knowledge on molecular diagnosis helps in detection and treatment of diseases. This study focused on the application of genomics and proteomics in the detection of chronic diseases such as cancer and sick cell anemia, and infectious diseases such as immunodeficiency syndrome whose emergence and development is determined by molecular metabolisms. Finally, the application of both genomic and proteomic components in the diagnosis of inherited diseases is also investigated. In brief, this study found molecular diagnosis highly useful in clinical settings to address medical conditions. Also, various DNA sequencing techniques have been useful in studying the proteins and other substances that express the existence of disease-causing pathogens in the human body. In conclusion, the study found that more research is necessary to increase not only the knowledge of the genome and proteome but also discover new diseases whose molecular interventions can provide treatment and cure. As a result, the mortality rate due to infectious, inherited and chronic diseases will reduce while and patient’s experience will improve simultaneously.
Medical Diagnostic
In the recent years, molecular biology has opened opportunities for detection and healing wide range of diseases. It recognizes that every organism has species-specific DNA properties, and when manipulated in clinical settings can treat diseases (Schumacher et al., 2016). For this reason, molecular diagnostic as a group of techniques based on the molecular biology and clinical testing has been successful in study and treatment of genetic and acquired diseases (Peng, 2012). Consequently, these techniques have been recognized in the treatment of all kind of diseases for they aid medical isolation, characterization, and manipulation of cells’ molecular structure and functions (Fischer et al., 2016). Specifically, it helps to focus on the DNA, and mRNA components in diagnosing various diseases. Molecular biology present limitless opportunities for research for prevention and treatment of not only acquired but also genetically determined diseases.
Molecular diagnosis of the disease involves various DNA sequencing and analysis technologies. Each of technologies involves a wide range of tools that are used in the study of diseases. One technology is the Next Level Sequencing (NLS) that allows the study of DNA segments, up to including the entire genome (Kurkela and Brown, 2010). Unlike the initial approaches that focused on sequencing a single DNA at a time, NLS allows multiple sequencing in a rapid manner.
The other technology used in genomic and proteomic studies of diseases is Whole Genome Sequence (WGS). In this technology, the DNA content that encloses the genome of an individual is usually sequenced in studies of personalized treatment. The technology also helps interpret the mutation that may occur in clinical phenotypes. Although this technique helps researchers to focus on the entire genome and provides accurate results, it does not include all components of the genome (Fischer et al., 2016). For this reason, the next technology, the Whole Exon Sequencing (WES) is used in exceptional cases.
The Whole-Exon Sequencing is a component of the genome responsible for coding some proteins. The proteins in this techniques are referred to as the exon and have about 1 percent genome useful in the detection of diseases (Kurkela and Brown, 2010). In all cases, WES involve DNA sequencing to detect the encoding proteins (exon) and can comprise the DNA regions. Unlike the WGS, WES helps sequence the mRNA molecules that are not associated with synthesis of the proteins. In both research and clinical settings, all these techniques and the tools developed on their principles have been largely used to diagnose diseases.
Application in Diagnosis of Chronic Diseases
One application of molecular biology is in the diagnosis of diseases such as cancer, whereby identification and analysis of disease markers help to reveal patterns about diseases. Unlike other diagnostic technologies such as nanotechnology which are under development and its potential not well documented, molecular diagnostic using genome and proteomic analysis have demonstrated success in detection of diseases through manipulation of protein markers.
Specifically, in Female’s ovarian cancer, proteomic studies have helped researchers identify a serum protein pattern that differentiates between patients with the disease among the unaffected female’s population. Remarkably, such detection has come with a predictive value of 94 percent (Toss et al., 2013). Similarly, genomic sequencing and profiling have helped understand cancer development. Evidently, fundamental genetic mechanisms driving cancer initiation, development and maintenance has been achieved, leading to treatment and curative interventions.
A comparison between normal ovarian tissue (A and B) and a tumor tissue C and D with cancer cells) both from a normal patient and their corresponding Representative Gel (Adapted from Toss et al., (2013)
One tool used in diagnosing cancer is the DNA microarray that consists of a two dimension matrix of molecules usually applied on plastic, synthetic or nylon membrane to detect their nucleic acids and proteinase antibodies (Peng, 2012). One advantage of microarrays is that it detects variation in antibodies, nucleic acids, and pathogens simultaneously. For this reason, it is very sensitive, thus appropriate for different cancer diagnosis. Moreover, it can work in high samples.
Microarray, however, is expensive and complex for routine use in diagnosis. Also, it depends on amplification of nucleic acid before the profiling. Nonetheless, the use of microarray particularly DNA and serological tools continue to expand, taking over important roles in future microbiology laboratories. Evidently, several future application has been projected including tuberculosis detection.
Despite the effectiveness of using these microarrays in measuring the relative concentration of nucleic acid and significance disease events, their future seems to be overtaken by the new and upcoming DNA sequence technologies. The baseline of this rapid replacement is the limitations in using microarrays such as their indirect measure of relative concentration thereby influencing the complex or inaccurate detection of diseases.
In another study, Chang and Chan (2010) reported on the use of molecular biology to detect sickle cell anemia. In previously developed diagnosis technology for sickle cell anemia such as Gel electrophoresis has been found effective. However, obtaining results is a slow process and DNA analysis. Also, visualization dyes may involve toxic contamination of the DNA under study. For these reasons, genomic and proteomic diagnostic analysis in anemia detection has been reported most effective with rapid results and reduced toxicity.
In this study by Chang and Chan (2010), DNA analysis revealed a point mutation occurring in the Beta-globin chain of the hemoglobin. Also, the spleen was found to remove aggregation of abnormal hemoglobin. The tool used in this study is the Ddel gene analysis that detects the concentration of certain proteins influencing diseases.
In particular, the study found that the Glu 6, GAG order in the DNA has been transformed to GTG that is responsible for encoding the valine (Chang and Chan, 2010; Fischer et al., 2016). The change from A to T is a mutation that disrupts the recognition sequence that restricts the endonuclease Ddel:C/TGAG. The Ddel tool help detects the sickle cell mutation by digesting DNAs of both normal genomic and for sickle cell (Chang and Chan, 2010). The Ddel is used to conduct a DNA hybridization that uses a 5’specific fragment of the normal Beta-globin allele and one for the normal globin, thereby producing two fragments that are 175 and 201 base pairs (Chang and Chan, 2010). In the study, however, the DNA for sickle cell anemia (Allele S) was found to produce large single fragments with 376 base pairs (Chang and Chan, 2010). Also, this DNA was found to be resistant to Ddel digestion and polymerase reaction was used to simplify the detection of mutation in the sickle cell DNA. Other disease found to be visible using genome analysis include ‘Huntington, Stroke, and Coronary Artery Disease.
One advantage of Ddel is that it is sensitive in identifying the protein concentrations and variations events occurring to anemia patients. Also, the tools are cheap and can focus on large volume of units per concentrations. However, the tool’s result are highly dependent on the solvents or dye used. In this case, low quality or contamination can lead to misleading results. Due to the ability of this tool to factor in other conditions such as temperature, the volume of enzymes, and time required to digest the DNA during testing, it has become a most realistic tool and thus widely used. The future involves advanced developments to eliminate the flaws while at the same time increasing the sensitivity to the DNA proteins.
Application in Diagnosis of Genetic Conditions
Another use of molecular biology specifically the genomics and proteomic is in the diagnosis of inherited diseases. In a study by Jamuar and Tan (2015), Mendelian disease was investigated using the Next Generation Sequencing (NGS), a process in which the DNA is broken down and the fragments amplified using PCR. The study found that NGS was effective in the diagnosis of Mendelian disease and can be used in different clinical settings (Renkema et al., 2014; Cummings et al., 2016). The diagnostic yield achieved in the study was 25 percent and therefore did not attract significance use in medical settings. Unlike in NGS, the use of Whole Genome Sequencing or Whole Exon Sequencing has been reported with a success rate between 25- 50 percent. However, according to Raje et al. (2015), further research studies have projected increased effectiveness of the NGS as the molecular biology and sequencing tools continue to increase.
The stages of the Next Generation Sequence (Adapted from Renkema et al. (2014)
Another technique for genomic testing is the Direct to Consumer testing (DTC), and it mainly helps to study different disease markers, therefore, accurate detection of any underlying disease. Also, the DTC has been useful in investigating the ways the complexity of a disease such as cancer and anemia develop (Cummings et al., 2016). Consequently, practitioners can intervene by providing factors for reducing the disease regeneration or replace the organic elements depleted in some patients such as hemoglobin in sickle cell anemia with synthetic solutions.
Although DTC, is accurate and effective in revealing conditions most people never thought they have, it has expensive and involves complex procedures (Kannan et al., 2016). Also, some studies have revealed that some research reports are over-promising when compared with the evidence-based case studies done in the past few years.
Application in Diagnosis of Infectious Disease
Another application of molecular biology is in the diagnosis of infectious diseases such as drug-resistant Mycobacterium tuberculosis. In general, a screening test of the human serum can detect the pre-clinical conditions of infections thereby facilitating early treatment (Kurkela, and Brown, 2010). As a result, the transmission of such diseases to the public is reduced. In studies of M. tuberculosis, by Liu et al. (2016) proteomic techniques have helped identification of rRv3369 and rRv3874 Vitro proteins that reveal potential influence as serodiagnostic antigens. The tests have come with high sensitivity that ranges between 60 and 74 percent and accuracy of 96-97 percent in clinical settings. Such success rate makes the use of proteomic analysis more effective than the normal medical, cultural testing specimens such as sputum or biopsied tissues from a sick patient. Although the cultural testing provides results even in 30 minutes, they only detect about 10 percent to 70 percent of TB cases (Garberi et al., 2011). In this case, the genomic and proteomic analysis is the most effective in the diagnosis of Tuberculosis.
One genomic and proteomic technique used on molecular diagnostic of Tuberculosis is the Polymerase Chain Reaction (PCR) (Kurkela, and Brown, 2010; Riccio et al., 2016). It works in amplification of nucleic acid to surface the properties of the virus and other pathogens (Cho, & Yu, 2012). As a result, the proteins markers such as rRv3369 can be detected and manipulated through medications. The PCR also help track changes in the concentration of antigens due to the presence of M. tuberculosis that causes the disease.
On advantage of using PCR in diagnosis of infectious disease is that it can detect a wide range of pathogens despite their complex modifications (Yousef & Jothy, 2014). Also, it is more sensitive and specific when in comparison with other techniques that work on the antigen and antibody detection (Kannan et al., 2016). However, there are high chances of contamination when using this technique, leading to positive results that are false or illogical (Kitano, Zelanis, & Iwai, 2016). Also, it requires special facilities, instrumentation and quality control that render the entire project expensive. Moreover, medical institution sin developing nations with a high prevalence of the disease cannot afford the technology.
Most research institutes have engaged in research seeking future development of PCR and the overall diagnostic technique of diagnosis. For instance, the study by Garberi et al. (2011) recommends the development of Orange G3TB, an integrated system for detecting tuberculosis successfully even in samples with low bacillus loads. Several studies have focused on the development of the PCR technique, meaning that the molecular diagnosis of Tuberculosis using genome and proteomics is expanding. The key objectives in the development of molecular diagnostic are to achieve rapid and positive results from both low or high concentration of antigens and other proteins. Also, such discovery should enable detection and treatment of Tuberculosis in countries with high prevalence.
Conclusion
Molecular biology has enabled increased understanding of various diseases with the genetic, infectious or chronic basis that could not be achieved in other existing means. The increased knowledge in this field has resulted in the development of effective techniques for diagnosing chronic diseases such as cancer, heart diseases, leukemia, tuberculosis and some immunodeficiency disease. In particular, the genomic and proteomic-based research have resulted in the development of tools that have are widely used to detect various metabolically changes at the molecular level, and consequently revealing underlying diseases. More importantly, DNA analysis has yielded adequate information to determine the complexity of the disease and the possible interventions. When applied in clinical settings, patients receive care that has reduced their suffering and prolonged their life as well. However, while the potential application of this knowledge is expanding as more research finding are reported, the challenges surrounding the use of these molecular diagnostic approaches tend to evolve. In this case, further research and evidence-based experiments are required not only to overcome these challenges but also to increase the existing body of knowledge and new application of molecular diagnosis in a wide range of diseases.
References
Abdallah, B, Horne, S, Kurkinen, M, Stevens, J, Liu, G, Ye, C, Barbat, J, Bremer, S, & Heng, H 2014, 'Ovarian cancer evolution through stochastic genome alterations: defining the genomic role in ovarian cancer', Systems Biology In Reproductive Medicine, 60, 1, pp. 2-13
Chandramouli, K. and Qian, P. (2009). Proteomics: Challenges, Techniques, and Possibilities to Overcome Biological Sample Complexity.
Chang, A. and Chan, L. (2010). Clinical applications of molecular biology. Wllley Online Library.
Chen, G, Yang, Z, Eshleman, J, Netto, G, & Lin, M 2016, 'Molecular Diagnostics for Precision Medicine in Colorectal Cancer: Current Status and Future Perspective', Biomed Research International, 2016, pp. 1-12, Academic Search Premier, EBSCOhost, viewed 29 December 2016.
Cho, C.-H., & Yu, J. (2012). From inflammation to cancer: advances in diagnosis and therapy for gastrointestinal and hepatological diseases. Singapore, World Scientific. http://www.123library.org/book_details/?id=114823.
Cummings, B., Marshall, J., Tukiainen, T., Lek, M., Donkervoort, S., Foley, A., Bolduc, V., Waddell, L., Sandaradura, S., O'Grady, G., Estrella, E., Reddy, H., Zhao, F., Weisburd, B., Karczewski, K., O'Donnell-Luria, A., Birnbaum, D., Sarkozy, A., Hu, Y., Gonorazky, H., Claeys, K., Joshi, H., Bournazos, A., Oates, E., Ghaoui, R., Davis, M., Laing, N., Topf, A., Consortium, G., Beggs, A., Kang, P., North, K., Straub, V., Dowling, J., Muntoni, F., Clarke, N., Cooper, S., Bonnemann, C. and MacArthur, D. (2016). Improving genetic diagnosis in Mendelian disease with transcriptome sequencing.
Fischer, M, Renevey, N, Thür, B, Hoffmann, D, Beer, M, & Hoffmann, B 2016, 'Efficacy Assessment of Nucleic Acid Decontamination Reagents Used in Molecular Diagnostic Laboratories', Plos ONE, 11, 7, pp. 1-9, Academic Search Premier, EBSCOhost, viewed 29 December 2016.
Garberi, J., Labrador, J., Garberi, F., Garberi, J., Peneipil, J., Garberi, M., Scigliano, L. and Troncoso, A. (2011). Diagnosis of Mycobacterium tuberculosis using molecular biology technology.
Hsu, A., Fleisher, T. and Niemela, J. (2009). Mutation analysis in primary immunodeficiency diseases: case studies. National Institutes of Health
Jamuar, S. and Tan, E. (2015). Clinical application of next-generation sequencing for Mendelian diseases.
Jimenez, J. (2016). Genomic approaches can provide answers to undiagnosed primary immunodeficiency diseases. [Online] Baylor College of Medicine. Available at: https://www.bcm.edu/news/genome-sequencing/genomic-approach-to-immunodeficiency-diseases [Accessed 28 Dec. 2016].
Kannan, L, Ramos, M, Re, A, El-Hachem, N, Safikhani, Z, Gendoo, D, Davis, S, Gomez-Cabrero, D, Castelo, R, Hansen, K, Carey, V, Morgan, M, Culhane, A, Haibe-Kains, B, & Waldron, L 2016, 'Public data and open source tools for multi-assay genomic investigation of disease', Briefings In Bioinformatics, 17, 4, pp. 603-615, Business Source Complete, EBSCOhost, viewed 29 December 2016.
Kitano, E, Zelanis, A, & Iwai, L 2016, 'Proteomics and drug discovery in cancer,' Drug Discovery Today, 21, 2, pp. 264-277, Academic Search Premier, EBSCOhost, viewed 29 December 2016.
Kurkela, S. and Brown, D. (2010). Molecular Diagnostic Techniques. [Online] Academia.edu. Available at: https://www.academia.edu/3577369/Molecular_Diagnostic_Techniques [Accessed 28 Dec. 2016].
Lecuit, M. and Eloit, M. (2014). The diagnosis of infectious diseases by whole genome next-generation sequencing: a new era is opening.
Liu, Y, Wang, L, Feng, Y, He, C, Liu, D, Cai, X, Jiang, L, Chen, H, Liu, C, Wu, H, & Mei, L 2016, 'A New Genetic Diagnostic for Enlarged Vestibular Aqueduct Based on Next-Generation Sequencing', PLoS ONE, 11, 12, pp. 1-13,
Peng, X. (2012). Developing and evaluating genomics- or proteomics-based diagnostic tests: statistical perspectives. - PubMed - NCBI. [Online] Ncbi.nlm.nih.gov. Available at: https://www.ncbi.nlm.nih.gov/pubmed/17085803 [Accessed 28 Dec. 2016].
Raje, N., Soden, S., Swanson, D., Ciaccio, C., Kingsmore, S., and Dinwiddie, D. (2015). The utility of Next Generation Sequencing in Clinical Primary Immunodeficiency.
Renkema, K., Stokman, M., Giles, R. and Knoers, N. (2014). Next-generation sequencing for research and diagnostics in kidney disease.
Riccio, A, De Ferrari, L, Chiappori, A, Ledda, S, Passalacqua, G, Melioli, G, & Canonica, G 2016, 'Molecular diagnosis and precision medicine in allergy management', Clinical Chemistry & Laboratory Medicine, 54, 11, pp. 1705-1714, Academic Search Premier, EBSCOhost, viewed 29 December 2016
Schumacher, S, Sohn, H, Qin, Z, Gore, G, Davis, J, Denkinger, C, & Pai, M 2016, 'Impact of Molecular Diagnostics for Tuberculosis on Patient-Important Outcomes: A Systematic Review of Study Methodologies', PLoS ONE, 11, 3, pp. 1-21, Academic Search Premier, EBSCOhost, viewed 29 December 2016.
Taylor, B, Barretina, J, Maki, R, Antonescu, C, Singer, S, & Ladanyi, M 2011, 'Advances in sarcoma genomics and new therapeutic targets,' Nature Reviews Cancer, 11, 8, pp. 541-557, Academic Search Premier, EBSCOhost, viewed 29 December 2016
Toss, A., De Matteis, E., Rossi, E., Casa, L., Iannone, A., Federico, M. and Cortesi, L. (2013). Ovarian Cancer: Can Proteomics Give New Insights for Therapy and Diagnosis.
Yousef, G. M., & Jothy, S. (2014). Molecular testing in cancer. http://public.eblib.com/choice/publicfullrecord.aspx?p=1730873.
Zhang HM, e. (2014). Application of genomics and proteomics in drug target discovery. - PubMed - NCBI. [Online] Ncbi.nlm.nih.gov. Available at: https://www.ncbi.nlm.nih.gov/pubmed/24446303 [Accessed 29 Dec. 2016].
