Project Description

Summary

Hyperion is a synthetic biology project aimed at solving the problem of inaccurate and delayed cancer diagnosis. Despite the advances in modern medicine, cancer remains one of the leading causes of death worldwide, in part due to the limitations of current diagnostic methods. Hyperion offers a novel solution to this problem by utilizing a biosensor that can detect cancer cells with high accuracy and speed. The biosensor works by recognizing specific biomarkers that are present in cancer cells and not in healthy cells. The advantages of Hyperion are numerous: it is cost-effective, non-invasive, and can be used in a wide range of settings, from hospitals to remote areas with limited access to medical facilities (point-of-care). Moreover, the biosensor can be easily customized to detect other diseases and abnormalities, such as aging, neurodegenerative disorders, and autoimmune diseases. Hyperion has the potential to revolutionize cancer diagnosis and improve the quality of life for millions of people around the world.

Description

Cancer is one of the most devastating diseases known to humankind, affecting millions of people every year and causing immeasurable suffering. Despite the advances in modern medicine, cancer remains a formidable foe, in part due to the limitations of current diagnostic methods. The existing tools for cancer detection, such as imaging tests, biopsies, and blood tests, are often inaccurate, costly, and time-consuming. In many cases, the cancer is not detected until it has already metastasized and spread to other parts of the body, making it more difficult to treat and reducing the chances of survival.

Hyperion is a synthetic biology project aimed at solving the problem of inaccurate and delayed cancer diagnosis. Despite the advances in modern medicine, cancer remains one of the leading causes of death worldwide, in part due to the limitations of current diagnostic methods. Hyperion offers a novel solution to this problem by utilizing a biosensor that can detect cancer cells with high accuracy and speed. The biosensor works by recognizing specific biomarkers that are present in cancer cells and not in healthy cells. The advantages of Hyperion are numerous: it is cost-effective, non-invasive, and can be used in a wide range of settings, from hospitals to remote areas with limited access to medical facilities (point-of-care). Moreover, the biosensor can be easily customized to detect other diseases and abnormalities, such as aging, neurodegenerative disorders, and autoimmune diseases. Hyperion has the potential to revolutionize cancer diagnosis and improve the quality of life for millions of people around the world.

Hyperion is a synthetic biology project aimed at addressing this problem by developing a biosensor that can detect cancer cells with high accuracy and speed. The biosensor works by recognizing specific biomarkers that are present in cancer cells and not in healthy cells. By detecting these biomarkers, the biosensor can identify the presence of cancer in its early stages, when it is most treatable.

The advantages of Hyperion over current diagnostic methods are numerous. First and foremost, it offers a non-invasive, cost-effective, and rapid way to detect cancer, making it accessible to a wider range of people and settings. The biosensor can be easily integrated into existing medical equipment, such as CT scanners and MRI machines, allowing doctors and clinicians to obtain accurate and real-time data about a patient's condition. In addition, Hyperion has the potential to be customized to detect other diseases and abnormalities, such as aging, neurodegenerative disorders, and autoimmune diseases, which could significantly improve the quality of life for millions of people around the world.

Hyperion represents a major breakthrough in cancer diagnosis, offering tremendous advantages to the healthcare system and healthcare facilities. By providing a reliable and efficient way to detect cancer in its early stages, Hyperion can help reduce the burden on healthcare providers, allowing them to allocate their resources more effectively. The non-invasive nature of the biosensor also means that it can be used in a variety of settings, from hospitals to remote clinics, making it accessible to patients who may not have had access to traditional diagnostic methods. Furthermore, the biosensor's ability to detect other diseases and abnormalities means that it has the potential to significantly improve the overall quality of care provided to patients. Hyperion has the potential to revolutionize cancer diagnosis and improve the health outcomes of millions of people around the world, making it a project of immense significance and impact.

Future Applications

In addition to the biosensor, Hyperion has the potential for software improvements that could further enhance its capabilities. One such improvement could be the incorporation of artificial intelligence (AI) algorithms that could analyze the biosensor data and provide predictions about the progression and severity of cancer. This could lead to more personalized and effective treatment plans for patients. Another potential improvement could be the development of a mobile application for healthcare providers that could securely transmit and store patient data, allowing for more efficient and streamlined communication between medical professionals. Additionally, automated reports generated by the Hyperion software could be uploaded to government databases, such as the Greek government's health information system, gov.gr, allowing for better tracking of cancer prevalence and outcomes. These software improvements could significantly enhance the impact of Hyperion and provide even greater benefits to patients and healthcare providers.

Background

The development of Project Hyperion draws significant inspiration from a groundbreaking patent that addresses the urgent need for rapid and effective diagnostic tools in the context of public health crises, as exemplified by the SARS-CoV-2 pandemic. In this pivotal study, an antibody-free biosensor was innovatively engineered, centered on the immobilization of ACE2 protein on the surface of gold interdigitated electrodes.

The core objective was to create a diagnostic method capable of swiftly and accurately monitoring the spread of the virus. Remarkably, this biosensor demonstrated exceptional sensitivity, with a reported limit of detection (LOD) of 750 pg/μL/mm² when targeting the virus's structural spike protein, even under laboratory conditions. Such an LOD signifies the biosensor's ability to detect minuscule viral loads.

Furthermore, the practical applicability of this biosensor was validated through its response to clinical samples obtained from hospitalized patients, including swab and saliva specimens. The biosensor's efficacy was confirmed by electrical effective capacitance measurements, which were correlated with real-time PCR results. Impressively, this innovative technology showcased the capability to reliably distinguish between samples that tested positive for the virus and those that were negative, demonstrating its potential as a diagnostic tool.

One of the most significant advantages of this biosensor lies in its speed and simplicity. It is capable of delivering results in less than two minutes, making it a powerful candidate for rapid point-of-care detection. Moreover, its ease of operation enhances its suitability for widespread use in healthcare settings.

In summary, the patent underpinning Project Hyperion represents a significant milestone in the development of biosensors for rapid virus detection. By leveraging the immobilization of ACE2 protein on gold interdigitated electrodes, this innovative technology not only provides rapid and sensitive virus detection but also holds the promise of revolutionizing diagnostic tools for various applications, including the early detection of cancer.

References

  1. Georgas A, Lampas E, Houhoula DP, Skoufias A, Patsilinakos S, Tsafaridis I, Patrinos GP, Adamopoulos N, Ferraro A, Hristoforou E. ACE2-based capacitance sensor for rapid native SARS-CoV-2 detection in biological fluids and its correlation with real-time PCR. Biosens Bioelectron. 2022 Apr 15;202:114021. doi: 10.1016/j.bios.2022.114021. Epub 2022 Jan 21. PMID: 35092924; PMCID: PMC8776344.

Introduction

The Proteome, encompassing all proteins within a cell, plays a fundamental role in the biological functioning of organisms across the spectrum of life forms. Proteins are classified into distinct functional families, with structural proteins maintaining cell morphology and functional proteins, including enzymes, regulating essential cellular processes. Post-translational modifications, such as phosphorylation, dynamically alter protein functionality, and kinases, a class of enzymes, are central to this process. The human cell boasts a remarkable array of 518 different kinases, collectively known as the Kinome, reflecting their pivotal role in cellular regulation.

Phosphorylation involves the addition of phosphate groups to proteins, thereby modulating their activity. This reversible process acts as a cellular switch, turning proteins on or off as required for various cellular functions, including cell division, proliferation, differentiation, and apoptosis. Dysregulation of kinase activity within the Kinome can lead to pathological conditions, including cancer.

Traditional approaches to understanding kinase signaling pathways and their dynamic activity have relied on technologies such as phospho-specific antibody arrays and mass spectrometry. However, these methods have limitations, including the availability of specific antibodies and the time-intensive nature of mass spectrometry.

Project Hyperion introduces an innovative alternative: an array of 518 normal antibodies immobilized in individual wells for the entire Kinome. This approach sidesteps issues of cross-reactivity often seen with phospho-specific antibodies and the resource-intensive nature of mass spectrometry. The methodology involves incubating the array with cellular proteins, detecting antibody-kinase interactions using interdigital capacitors, and subsequently evaluating phosphorylation status through enzymatic reactions. This array technology has the potential to redefine the profiling of kinase activity, offering a rapid, cost-effective, and accessible solution.

Project Hyperion's innovative approach holds the promise of advancing our understanding of kinase-based signaling networks and their role in health and disease, with broad-reaching implications for personalized medicine, drug development, and diagnostics.

Map Kinase

In the context of the iGEM Competition, our team started the Biosensor testing by studying the Map Kinase cascades.

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The Mitogen-Activated Protein Kinase (MAPK) signaling pathway is a fundamental mechanism that orchestrates crucial cellular processes in response to external cues, playing a pivotal role in cell proliferation, differentiation, survival, and apoptosis. This intricate molecular network consists of a series of protein kinases that relay signals from the cell membrane to the nucleus, ultimately influencing gene expression and cell behavior.

MAPK signaling begins with the activation of cell surface receptors, such as receptor tyrosine kinases (RTKs) or G-protein coupled receptors (GPCRs), in response to extracellular stimuli like growth factors, hormones, or stress signals. Upon activation, these receptors initiate a cascade of intracellular events, leading to the activation of the core MAPK components: extracellular signal-regulated kinases 1 and 2 (ERK1/2), c-Jun N-terminal kinases (JNK), and p38 MAPK.

ERK1/2, perhaps the most well-studied MAPKs, are often associated with cell growth and proliferation. Once activated, ERK1/2 translocate to the nucleus, where they phosphorylate transcription factors and other proteins, thereby influencing gene expression and promoting cell cycle progression. JNK and p38 MAPK, on the other hand, are primarily involved in stress responses and can induce apoptosis or cell cycle arrest.

Why is MAPK Signaling Important?

The significance of MAPK signaling lies in its ability to integrate extracellular cues with intracellular responses, allowing cells to adapt to their environment. By modulating gene expression and cellular behavior, MAPK signaling ensures proper development, tissue homeostasis, and efficient responses to threats like DNA damage or oxidative stress.

However, dysregulation of the MAPK pathway is implicated in various diseases, most notably cancer. Cancer cells often exploit MAPK signaling to drive their uncontrolled growth and evade normal cellular checkpoints. Alteration of the RAS-RAF-MEK-ERK-MAPK (RAS-MAPK) pathway has frequently been reported in human cancer as a result of abnormal activation of receptor tyrosine kinases or gain-of-function mutations mainly in the RAS or RAF genes. This hyperactivity can result from mutations in upstream receptors or downstream signaling molecules, allowing cancer cells to proliferate relentlessly.

A member of the Mitogen Activated Protein Kinase (MAPK) family is Extracellular Signal-Regulated Kinase (ERK). [2] There are numerous events that may lead to the subsequent hyperphosphorylation of ERK1/2 molecules that activate certain transcription factors such as the NF-kβ resulting in the deregulation of downstream pathways. Such conditions mainly stem from genetic mutations such as Ras, B-Raf and those that result to the overexpression and/or overactivation of tyrosine kinases located on the cellular membrane, including the Epidermal Growth Factor Receptor (EGFR), enhancing cancer cells’ survivability, proliferation and chemoresistance. [1] As such, it appears as no surprise that this kind of mutations and subsequent conditions are present in over 1/3 of all cancers. [3].

ERK is a signaling protein located in the cytoplasm and, when phosphorylated, it translocates to the nucleus of the cell with its own unique mechanism. It is the final member of the MAPK signaling cascade and promotes cell cycling. [6] Physiologically, ERK is activated via extracellular stimuli as well as under stress inducing conditions such as high concentration of Reactive Oxygen Species and consists a crucial element of the DNA Damage Response as phosphorylation of ERK ½ is positively associated with apoptosis [4], as well as antiapoptotic mechanisms [7] and autophagy [9]. Thereby, it is crucial for the physiology of the cell to maintain and regulate the function of ERK utilizing mechanisms like phosphorylation, conformation-dependence, auto-inhibition and dimerization. [6] However, even though the tight regulation of the mechanism of activation, mutations and other events may alter the ERK into a survival tool for cancers, such as the mutations of RAS and RAF oncogenes that are not normally upstream of ERK [5] and their etiology can be either inheritance or environmental factors. [4] Mutations of ERK itself are relatively rare in cancers as the most usual ones affect factors that lead to its abnormal activation rate. [6]

Apart from the aforementioned, ERK overactivation also enhances chemoresistance, ability to metastasize and survivability in the otherwise unfavorable conditions of the tumor microenvironment. This is a result of mainly the antiapoptotic effects of the protein as well as the induction of cyclins and pro-survival effectors like eIF4E thus granting the tumor cells resistance to molecules such as paclitaxel. Additionally, the deregulation of ERK activity is a significant factor in immune evasion via the excretion of cytokines that dampen immune surveillance as well as the function of cytotoxic lymphocytes and drive regulatory cells towards a state catering immune suppression. [8]

Considering the above, it appears as no wonder that ERK overaction is usually spotted in and characterizes some of the most aggressive types and forms of neoplastic diseases, which affect patients whose survival depends on an early diagnosis and start of intervention. Therefore figuring a solution to a diagnostic method that lies on this condition and promises to deliver fast, early and affordable diagnosis is a significant step towards achieving a future where cancer is not a death sentence.

Detecting Cancer-Associated Changes through Hyperphosphorylation Patterns

Understanding the intricacies of MAPK signaling in cancer has led to the identification of hyperphosphorylation patterns as valuable biomarkers. In cancer cells, aberrant MAPK activation frequently leads to hyperphosphorylation of key proteins involved in cell cycle regulation and apoptosis. These phosphorylation patterns can be detected through advanced biochemical techniques and serve as indicators of abnormal MAPK activity, signaling potential malignancy.

References:

  1. De S, Campbell C, Venkitaraman AR, Esposito A. Pulsatile MAPK Signaling Modulates p53 Activity to Control Cell Fate Decisions at the G2 Checkpoint for DNA Damage. Cell Rep. 2020 Feb 18;30(7):2083-2093.e5. doi: 10.1016/j.celrep.2020.01.074. PMID: 32075732; PMCID: PMC7029415.

  2. Santarpia L, Lippman SM, El-Naggar AK. Targeting the MAPK-RAS-RAF signaling pathway in cancer therapy. Expert Opin Ther Targets. 2012 Jan;16(1):103-19. doi: 10.1517/14728222.2011.645805. Epub 2012 Jan 12. PMID: 22239440; PMCID: PMC3457779.

  3. Liu X, Zhang Y, Wang Y, Yang M, Hong F, Yang S. Protein Phosphorylation in Cancer: Role of Nitric Oxide Signaling Pathway. Biomolecules. 2021 Jul 10;11(7):1009. doi: 10.3390/biom11071009. PMID: 34356634; PMCID: PMC8301900.

  4. Morrison DK. MAP kinase pathways. Cold Spring Harb Perspect Biol. 2012 Nov 1;4(11):a011254. doi: 10.1101/cshperspect.a011254. PMID: 23125017; PMCID: PMC3536342.

  5. Regulatory Roles of MAPK Phosphatases in Cancer,Immune Network 2016; 16(2): 85-98. Published online: 28 April 2016,DOI: https://doi.org/10.4110/in.2016.16.2.85, Heng Boon Low, Yongliang Zhang

  6. Rezatabar, S, Karimian, A, Rameshknia, V, et al. RAS/MAPK signaling functions in oxidative stress, DNA damage response and cancer progression. J Cell Physiol. 2019; 234: 14951–14965. https://doi.org/10.1002/jcp.28334

  7. Lee, S.; Rauch, J.; Kolch, W. Targeting MAPK Signaling in Cancer: Mechanisms of Drug Resistance and Sensitivity. Int. J. Mol. Sci. 2020, 21, 1102. https://doi.org/10.3390/ijms21031102

  8. Christian P. Kratz, Martin Zenker,Inherited Disorders of the Ras-MAPK Pathway,Blood, Volume 132, Supplement 1,2018,Page SCI-41,ISSN 0006-4971,https://doi.org/10.1182/blood-2018-99-109379.

  9. Maik-Rachline, G.; Hacohen-Lev-Ran, A.; Seger, R. Nuclear ERK: Mechanism of Translocation, Substrates, and Role in Cancer. Int. J. Mol. Sci. 2019, 20, 1194. https://doi.org/10.3390/ijms20051194

  10. Degirmenci, U.; Wang, M.; Hu, J. Targeting Aberrant RAS/RAF/MEK/ERK Signaling for Cancer Therapy. Cells 2020, 9, 198. https://doi.org/10.3390/cells9010198

  11. Cagnol S, Chambard JC. ERK and cell death: mechanisms of ERK-induced cell death--apoptosis, autophagy and senescence. FEBS J. 2010 Jan;277(1):2-21. doi: 10.1111/j.1742-4658.2009.07366.x. Epub 2009 Oct 16. PMID: 19843174.

  12. Salaroglio, I.C.; Mungo, E.; Gazzano, E.; Kopecka, J.; Riganti, C. ERK is a Pivotal Player of Chemo-Immune-Resistance in Cancer. Int. J. Mol. Sci. 2019, 20, 2505. https://doi.org/10.3390/ijms20102505

  13. Bryant, K.L., Stalnecker, C.A., Zeitouni, D. et al. Combination of ERK and autophagy inhibition as a treatment approach for pancreatic cancer. Nat Med 25, 628–640 (2019). https://doi.org/10.1038/s41591-019-0368-8