Description

Overview

Cataracts, the leading cause of reversible blindness globally, affect approximately 94 million people worldwide. At the core of this condition lies oxidative damage to lens proteins: excessive accumulation of reactive oxygen species (ROS) attacks lens fibers, triggering protein denaturation, aggregation, and ultimately loss of transparency. While phacoemulsification combined with intraocular lens implantation remains the clinical gold standard, its high cost and uneven distribution of medical resources restrict accessibility—especially in underserved regions. Superoxide dismutase (SOD), a key enzyme in the endogenous antioxidant network, effectively neutralizes ROS in the lens and slows oxidative stress-induced protein damage. However, natural SOD has application defects of poor thermal stability and easy degradation. Therefore, our project leverages computational biology-driven protein engineering to design and optimize SOD variants with enhanced performance. By improving stability and activity, these engineered enzymes will serve as the core component of a novel eye drop formulation. This approach aims to provide a cost-effective, accessible solution for cataract prevention and early-stage intervention, with potential to alleviate the global burden of this growing disease.

1 Our Inspiration

Our project inspiration began with a quiet, personal observation: the world slowly fading from the vibrant clarity we knew into a persistent haze for our grandparents. We witnessed cataracts not only dimmed their sight but also isolated them from life's precious details. This deeply human experience compelled us to investigate deeper. We learned that cataracts were the number one cause of blindness worldwide. At that time, invasive surgery was still the mainstream method of cataract treatment. Sadly, we discovered that many people were not incurable, but were trapped by financial barriers, inadequate surgical accessibility, and the risk of postoperative complications. In biology class, we had studied the scientific roots of age-related cataracts. The lens of the eye functioned similarly to a camera lens and required perfect transparency. As people aged, reactive oxygen species (ROS) accumulated and destroyed lens proteins, leading to a series of visual disturbances such as blurred vision, glare, and reduced color perception. Although the human body naturally produced superoxide dismutase (SOD), an antioxidant guardian that neutralized these harmful compounds, its activity declined with age, causing the eyes to lose their natural protection.However, we found that existing antioxidant products showed little effect on the treatment of cataracts.

In addition, we explored the innovative work of previous iGEM teams who had long recognized the therapeutic potential of SOD in combating oxidative stress across various diseases. Their pioneering research provided valuable insights for our subsequent project scenarios. The iGEM 2024_NJTech-China developed an impressive project that focused on using engineered bacteria to sense and treat inflammatory bowel disease (IBD). Their system specifically expressed three therapeutic proteins at inflammation sites: an immunomodulatory protein (PDL1), a tissue repair protein (MFP), and an antioxidant enzyme (Cu/Zn SOD). The Cu/Zn SOD component was designed to efficiently remove excess reactive oxygen species (ROS) from the intestinal environment, directly addressing the key oxidative stress problem in IBD pathology. Their approach illuminated both the promises and challenges of employing SOD in therapeutic applications, demonstrating how synthetic biology could create intelligent, targeted therapies with built-in safety mechanisms. Their work also highlighted the crucial need to overcome biological barriers for effective drug delivery. At that time, no team had directly addressed the unique challenges of ocular delivery for cataract prevention. This gap became our opportunity. We were inspired to harness the creativity of synthetic biology for ocular therapeutics. Thus began our synthetic biology journey.

2 Cataracts Background

2.1 Definition and Global Burden of Cataracts

Cataract, a leading cause of visual impairment worldwide, is characterized by the opacification of the eye's natural lens (Figure 1). This clouding occurs when proteins within the lens undergo structural changes, disrupting the normal passage of light and leading to symptoms such as blurred vision, increased sensitivity to glare, and diminished color perception . As the foremost cause of global blindness, cataracts account for approximately 33.4% of all blindness cases and 18.4% of moderate to severe visual impairment worldwide. It is estimated that around 94 million people are affected by cataract-induced moderate to severe vision loss or blindness . Advancing age is the most significant risk factor, with incidence rising substantially after the age of 50 [1].

Recent epidemiological studies indicate that the global years lived with disability (YLD) due to cataracts reached 6.55 million in 2021, marking a 91.8% increase from 1990 (3.42 million). Social barriers and unequal access to health care are major contributors to the burden of disease. Geographically, regions near the equator—including Southeast Asia, South Asia, and Sub-Saharan Africa—bear the highest burden. These areas experience intense ultraviolet radiation, a known risk factor for cataract development, coupled with limited medical resources, resulting in elevated age-standardized prevalence rates (ASPR) and YLD [2-3].

Figure1. Comparison of normal eye and Cataract-affected eye.

2.2 Major Pathological Mechanisms of Cataracts

The pathogenesis of cataracts is actively investigated under various hypotheses, including lens protein denaturation, apoptosis, genetic factors, and metabolic disorders [4]. Among these, oxidative stress and free radical damage are considered the most central and universal mechanism, serving as the common pathway through which multiple risk factors ultimately induce lens opacification. Oxidative stress occurs when the production of reactive oxygen species (ROS) overwhelms the body's antioxidant defenses, leading to harmful accumulation of radicals and subsequent tissue damage [5]. Key radicals involved include the superoxide anion (O2⁻) and hydrogen peroxide (H2O2). The lens, particularly its epithelial and fiber cells, contains high mitochondrial density [6], which is essential for maintaining energy metabolism and transparency. Although the lens possesses an intrinsic antioxidant system, including enzymes such as superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT), and glutathione reductase (GR). This protective capacity declines with age or under persistent stressors like UV exposure and diabetes. As the balance tilts toward oxidation, ROS attack soluble crystallin proteins, causing their denaturation, cross-linking, and eventual aggregation into insoluble light-scattering precipitates [7]. This process ultimately leads to lens clouding. Therefore, Targeting this oxidative damage mechanism is crucial for developing novel strategies to prevent cataracts.

Figure2. Schematic illustration of the oxidative stress in Cataractogenesis.

2.3 Limitations of Existing treatments

Currently, the clinical management of cataracts relies primarily on surgical and pharmacological interventions, both of which are constrained by significant limitations. Although surgical treatment, particularly phacoemulsification with intraocular lens (IOL) implantation [4], represents the most effective approach with success rates exceeding 95% and rapid visual recovery, its global coverage remains unsatisfactory This observation is consistent with the feedback we received from our interviews with cataract patients. However, surgical accessibility is especially limited in many developing regions due to economic constraints, inadequate medical infrastructure, and a shortage of trained ophthalmologists, as evidenced (Figure 3) by a nearly 30-fold disparity in the cataract surgical rate (CSR) between high-income nations (e.g., 11,000 per million in the USA and Netherlands) and some regions (e.g., 400 per million in South Africa). Furthermore, even successful procedures carry risks of serious complications, such as infectious endophthalmitis and IOL dislocation, which can compromise vision post-operatively [8].

Pharmacological approaches, aimed at slowing disease progression in early stages, are incapable of reversing existing lens opacification. The commonly used agents, including pirenoxine sodium and glutathione eye drops, function primarily as antioxidants or by supporting lens metabolic function [4]. However, their efficacy is generally modest, varies significantly among individuals, and fails to address the underlying pathogenesis. Consequently, cataract treatment faces a pronounced imbalance. while surgery is effective but often inaccessible, available drugs are merely palliative and insufficient. This pronounced treatment gap underscores the critical need for novel, accessible, and proactive strategies.

Figure3. Cataract surgical rate (CSR) across several different countries. Data referenced from [8].

3 Superoxide dismutase (SOD) Background

3.1 Key Roles and Mechanistic Insights of SOD

Superoxide dismutase (SOD) is a crucial metalloenzyme that serves as the primary defense against oxidative stress by catalyzing the dismutation of the superoxide anion radical (O2⁻) into hydrogen peroxide (H2O2) and molecular oxygen (O2). The resulting H2O2 is subsequently detoxified into water and oxygen by enzymes such as catalase and glutathione peroxidase, collectively safeguarding cellular components from oxidative damage and maintaining the intracellular redox balance [9]. SOD exists in multiple isoforms, including Cu/Zn-SOD, Mn-SOD, and Fe-SOD. Each SOD exhibits distinct subcellular localization and functional profiles (Table 1). Among these, Mn-SOD is particularly critical due to its specific strategic localization within the mitochondria, the primary site of cellular respiration and a major source of reactive oxygen species (ROS). It plays an indispensable role in preserving mitochondrial integrity and preventing oxidative damage that can propagate throughout the cell [10].

Within the specific context of cataract pathogenesis, mitochondrial dysfunction in lens epithelial cells and fiber cells is a key contributor to oxidative stress accumulation and protein aggregation, the hallmarks of lens opacification [11]. Compounding this issue, studies indicate that Mn-SOD activity declines with age and under conditions of heightened oxidative stress, thereby accelerating cataract progression [12]. Therefore, our project is dedicated to engineering novel Mn-SOD variants with enhanced structural stability and catalytic activity. By targeting the mitochondrial source of ROS within the lens, our bioengineered enzyme aims to provide a potent defense against oxidative insult, and represents a proactive strategy to delay or even reverse early-stage lens opacification.

Table1. Current Superoxide dismutase (SOD) classification information.

SOD Type	Metal Cofactor	Primary Location
Cu/Zn-SOD (Copper and Zinc-containing SOD)	(Cu) and Zinc (Zn)	Cytoplasm of eukaryotes
Mn-SOD (Manganese-containing SOD)	Manganese (Mn)	Mitochondrial matrix of eukaryotes
Fe-SOD (Ferrum-containing SOD)	Ferrum (Fe)	Mainly in prokaryotes
Ni-SOD (Nickel-containing SOD)	Nickel (Ni)	Certain bacteria

3.2 Current Challenges of SOD production

The clinical translation of superoxide dismutase (SOD) is constrained by significant bottlenecks in both production and ocular application. Regarding production, conventional methods face considerable challenges: natural extraction (e.g., from animal blood) raises concerns about viral contamination and involves complex purification processes; chemical synthesis proves cost-prohibitive and often yields products with suboptimal catalytic activity; while microbial biosynthesis, though promising, is hampered by complex fermentation control and difficulties in ensuring the stability and activity of the recombinant product [13].

For ocular therapeutics, the challenges are even more pronounced. As a macromolecular protein, SOD is highly susceptible to degradation by gastrointestinal enzymes upon oral administration, resulting in extremely low systemic bio-availability and ineffective delivery to ocular tissues. Even when formulated as topical eye drops, SOD exhibits poor corneal permeability due to the eye’s natural barriers (e.g., the corneal epithelium), and undergoes rapid clearance from the administration site, preventing effective accumulation at the target site. Furthermore, formulating a stable and effective eye drop that preserves SOD’s enzymatic activity over the long term remains a major hurdle [14]. Consequently, overcoming these limitations necessitates not only optimizing production strategies (e.g., using advanced fermentation and synthetic biology techniques to enhance yield and stability), but also the development of novel ocular drug delivery systems designed to enhance corneal penetration, prolong ocular residence time, and maintain enzymatic integrity. Therefore, we further optimized our ideas about eye drop formula in human practice and literature research during the iGEM project. For more information, please go to the Human Practice page.

Figure4. SOD Intervention allevates the Progression of Lens Opacification in Cataracts.

4 Our Solution

To overcome the limitations of scarce surgical access and ineffective pharmacological treatments in cataract, our iGEM 2025 project seeks to develop a novel eye drop formulation centered on a engineered superoxide dismutase (SOD). By proactively preventing and delaying early-stage cataracts, our solution aims to lower treatment costs and alleviate the global burden of this disease. Our project leverages computational biology methods such as machine learning (ML) to engineer thermostable and high-activity SOD variants. These variants will be expressed in Escherichia coli BL21(DE3) under optimized conditions, and then affinity chromatographic purification and enzymatic characterization (Figure 5).

For more information, please go to the Design page.

Figure5. The design schematic of our project.

5 References

​ [1] Chen SP, Woreta F, Chang DF. Cataracts: A Review. JAMA. 2025 Jun 17;333(23):2093-2103.

​ [2] Vision Loss Expert Group of the Global Burden of Disease Study; GBD 2019 Blindness and Vision Impairment Collaborators. Global estimates on the number of people blind or visually impaired by cataract: a meta-analysis from 2000 to 2020. Eye (Lond). 2024 Aug;38(11):2156-2172.

​ [3] Li M, Jia W, Song J, Ma J, Zhou Y, Han Y, Peng M, Zhou J, Chen X, Li X. Global prevalence and years lived with disability (YLDs) of cataract in 204 countries and territories: findings from the Global Burden of Disease Study 2021. Eye (Lond). 2025 Jun;39(9):1737-1743.

​ [4] 徐靖杰,张颖,姚克,等.白内障发病机制与防治策略的研究进展[J].中国科学:生命科学,2022,52(12):1807-1814.

​ [5] Mishra D, Kashyap A, Srivastav T, Yadav A, Pandey S, Majhi MM, Verma K, Prabu A, Singh V. Enzymatic and biochemical properties of lens in age-related cataract versus diabetic cataract: A narrative review. Indian J Ophthalmol. 2023 Jun;71(6):2379-2384.

​ [6] Babizhayev MA. Mitochondria induce oxidative stress, generation of reactive oxygen species and redox state unbalance of the eye lens leading to human cataract formation: disruption of redox lens organization by phospholipid hydroperoxides as a common basis for cataract disease. Cell Biochem Funct. 2011 Apr;29(3):183-206.

​ [7] Lee B, Afshari NA, Shaw PX. Oxidative stress and antioxidants in cataract development. Curr Opin Ophthalmol. 2024 Jan 1;35(1):57-63.

​ [8] Wang, W., Yan, W., Fotis, K., Prasad, N. M., Lansingh, V. C., Taylor, H. R., Finger, R. P., Facciolo, D., & He, M. Cataract Surgical Rate and Socioeconomics: A Global Study. Invest. Ophthalmol. Vis. Sci. 2017;57(14):5872-5881.

​ [9] 张旭,张蕾,许鹏琳,等.锰超氧化物歧化酶的催化原理与酶活性调节机制[J].生物化学与生物物理进展,2024,51(01):20-32.

​ [10] Zelko IN, Mariani TJ, Folz RJ. Superoxide dismutase multigene family: a comparison of the CuZn-SOD (SOD1), Mn-SOD (SOD2), and EC-SOD (SOD3) gene structures, evolution, and expression. Free Radic Biol Med. 2002 Aug 1;33(3):337-49.

​ [11] Zou X, Ratti BA, O'Brien JG, Lautenschlager SO, Gius DR, Bonini MG, Zhu Y. Manganese superoxide dismutase (SOD2): is there a center in the universe of mitochondrial redox signaling? J Bioenerg Biomembr. 2017 Aug;49(4):325-333.

​ [12] Younus H. Therapeutic potentials of superoxide dismutase. Int J Health Sci (Qassim). 2018 May-Jun;12(3):88-93.

​ [13] Ohrloff C, Hockwin O. Superoxide dismutase (SOD) in normal and cataractous human lenses. Graefes Arch Clin Exp Ophthalmol. 1984;222(2):79-81.

​ [14] Noor R, Mittal S, Iqbal J. Superoxide dismutase--applications and relevance to human diseases. Med Sci Monit. 2002 Sep;8(9):RA210-5.