Presented by: Team UteRus (Rochester)
Ethan Chen¹, Nello Gu¹, Gabe Isaacson¹, Emily Laskey¹, Isabel Lopez¹, Meghan Martin¹, Helen Shammas¹, Emily Schiller¹, Heather Shi¹, James Tang¹, Zivile Vebraite¹, Nancy Chen³, Ram Gona², Lynn Sidor², Alexis Stein³, Omar Soufan³, Roger White², Anne Meyer³⁴
¹Team member, undergraduate student at the University of Rochester, ²Teaching Assistant, graduate student at the University of Rochester, ³Mentor, faculty at the University of Rochester, ⁴Primary PI, faculty at the University of Rochester
Abstract
The 2020 University of Rochester iGEM Team, Team UteRus, created a novel, non-invasive diagnostic for endometriosis using menstrual effluent (ME). Endometriosis is a chronic disease, affecting 200 million women worldwide, that causes abnormal endometrial-like tissue growth outside of the uterine cavity. The only diagnostic available is exploratory surgery. Our team created lateral flow immunoassays (LFA) that can qualitatively and quantitatively measure the presence of endometriosis biomarkers in ME and built a model to optimize the assay design. Using a Plug and Play approach, our team designed plasmids for antibody production in SHuffle stain Escherichia coli and reduced the cost of our diagnostic. Additionally, we created menstrual cups best suited for the comfort of endometriosis patients as well as inexpensive laboratory equipment for clinics without laboratory access. Our team also built a predictive model that we integrated into a software tool as an endometriosis diagnostic based solely on clinical variables.
Team:Rochester/Poster
Creation of a Novel Diagnostic for Endometriosis using Menstrual Effluent
Project Goals
Our team had four main goals when designing our project
- Spread awareness of endometriosis and women’s reproductive health.
- Create simple and non-invasive sample collection methods tailored to individuals with endometriosis.
- Design reliable, affordable, and accessible diagnostic tools for endometriosis.
- Create user-friendly tools to interpret diagnostic results.
Inspiration
The major inspiration for our project came from a recent research publication detailing the presence of biomarkers for endometriosis in menstrual effluent [1]. Having a team member who had been diagnosed with endometriosis, we were shocked to learn about the lack of research and funding that were allocated to this chronic disease and wanted to improve the quality of care provided to endometriosis patients. We learned that:
Warren et al. not only exposed our team to endometriosis research and unique diagnostics methods but also inspired us to approach this problem with synthetic biology principles.
We were also inspired by a previous iGEM team, MIT 2016, who tried to detect endometriosis using newly characterized parts from mammalian synthetic biology [4]. Their work demonstrated how synthetic gene circuits could be utilized to detect endometriosis biomarkers and inspired our team to not only build upon this work, but find new ways to utilize synthetic biology in endometriosis diagnostics.
References:
[1] Warren, L. A., Shih, A., Renteira, S. M., Seckin, T., Blau, B., Simpfendorfer, K., Lee, A., Metz, C. N., & Gregersen, P. K. (2018). Analysis of menstrual effluent: Diagnostic potential for endometriosis. Molecular Medicine, 24(1), 1–12. https://doi.org/10.1186/s10020-018-0009-6
[2] Cramer, D., & Missmer, S. (2009). The Epidemiology of Endometriosis. Annals of the New York Academy of Sciences, 955(1), 11–22.
[3] Mutter, G. L. (2014). Endometriosis. In Pathology of the Female Reproductive Tract (pp. 487–508).
[4] iGEM Team MIT 2016 (2016). Diagnosing Endometriosis. Retrieved November 08, 2020, from https://2016.igem.org/Team:MIT
- Endometriosis affects 200 million people worldwide [2]
- Symptoms include heavy and painful periods, pain during intercourse, and infertility [3]
- Definite diagnosis currently relies upon expensive, invasive, exploratory surgery [3]
- There currently is a 10 year diagnostic delay [2]
Warren et al. not only exposed our team to endometriosis research and unique diagnostics methods but also inspired us to approach this problem with synthetic biology principles.
We were also inspired by a previous iGEM team, MIT 2016, who tried to detect endometriosis using newly characterized parts from mammalian synthetic biology [4]. Their work demonstrated how synthetic gene circuits could be utilized to detect endometriosis biomarkers and inspired our team to not only build upon this work, but find new ways to utilize synthetic biology in endometriosis diagnostics.
References:
[1] Warren, L. A., Shih, A., Renteira, S. M., Seckin, T., Blau, B., Simpfendorfer, K., Lee, A., Metz, C. N., & Gregersen, P. K. (2018). Analysis of menstrual effluent: Diagnostic potential for endometriosis. Molecular Medicine, 24(1), 1–12. https://doi.org/10.1186/s10020-018-0009-6
[2] Cramer, D., & Missmer, S. (2009). The Epidemiology of Endometriosis. Annals of the New York Academy of Sciences, 955(1), 11–22.
[3] Mutter, G. L. (2014). Endometriosis. In Pathology of the Female Reproductive Tract (pp. 487–508).
[4] iGEM Team MIT 2016 (2016). Diagnosing Endometriosis. Retrieved November 08, 2020, from https://2016.igem.org/Team:MIT
Raising Awareness
Due to a lack of endometriosis awareness among the general public and medical community:
To address this, we created multilingual educational infographics, brochures, and websites specifically targeted people of different ages and educational backgrounds to spread awareness of endometriosis and women’s health.These educational materials are displayed in several clinical settings, posted on social media platforms, and available for download from endometriosis foundations.
[Left] Our infographic displayed at University of Rochester Medical Center
[Right] Part of our brochure for post-menopausal women
Our team also collaborated with the Health Promotion Office at the University of Rochester to hold a panel that focused on addressing current gender disparities in health care and creating a space for students to ask questions about current issues in women’s health. We invited three panelists to our event:
Dr. Elhdad spoke about her experiences both as an endometriosis patient and researcher at our Perspectives in Women’s Health event.
References:
[1] Husby, G., Haugen, R., & Moen, M. (2003). Diagnostic delay in women with pain and endometriosis. Acta Obstetricia et Gynecologica Scandinavica, 82, 649–653.
[2] Endometriosis UK
- 46% of endometriosis patients see five or more doctors before receiving a diagnosis [1]
- 40% of women with endometriosis feel disrespected by their physicians [2]
To address this, we created multilingual educational infographics, brochures, and websites specifically targeted people of different ages and educational backgrounds to spread awareness of endometriosis and women’s health.These educational materials are displayed in several clinical settings, posted on social media platforms, and available for download from endometriosis foundations.
[Right] Part of our brochure for post-menopausal women
Our team also collaborated with the Health Promotion Office at the University of Rochester to hold a panel that focused on addressing current gender disparities in health care and creating a space for students to ask questions about current issues in women’s health. We invited three panelists to our event:
- Dr. Terlazzo from the University of Rochester Philosophy department who teaches courses such as Philosophy in Feminism
- Dr. Gubbels from University of Rochester Medical Center specializing in endometriosis
- Dr. Elhadad from Columbia University who created Phendo, an app that allows patients to track and manage endometriosis symptoms
References:
[1] Husby, G., Haugen, R., & Moen, M. (2003). Diagnostic delay in women with pain and endometriosis. Acta Obstetricia et Gynecologica Scandinavica, 82, 649–653.
[2] Endometriosis UK
Sample Collection
Menstrual Cup Designed for Endometriosis Patients
To collect the menstrual effluent needed for the non-invasive diagnostic panel, we designed an endometriosis-friendly menstrual cup. This cup takes into account the thickening of the vaginal wall and pelvic muscle fatigue that are commonly experienced by women with endometriosis, making menstrual cups painful [1, 2].
It also can collect up to 40 mL of menstrual effluent compared to the average market design of 25 mL to cater to heavy menstruation, a common symptom of endometriosis [6].
Our design for an endometriosis-friendly menstrual cup
UV-C Sterilizer
Bacterial growth on menstrual cups can cause a lethal condition known as Toxic Shock Syndrome [7]. To provide tools for sterilizing our Endo cup in areas with limited access to clean water, we designed a compact portable sterilizer that utilizes UV-C light [7].
3D model of our UV-C Sterilizer for menstrual cups
DIY Centrifuge
Prior to using menstrual effluent in our diagnostic, the sample must be spun to separate out the serum. In order to reduce the cost of sample preparation for our LFA, we designed a replicable DIY centrifuge through recycling toys, computer parts and simple circuitry.
Image of our assembled DIY Centrifuge
References
[1] Yang, N. (n.d.). Endometriosis: Radiology Reference Article. Retrieved from https://radiopaedia.org/articles/endometriosis
[2] Bispo, A. P., Ploger, C., Loureiro, A. F., Sato, H., Kolpeman, A., Girão, M. J., & Schor, E. (2016). Assessment of pelvic floor muscles in women with deep endometriosis. Archives of Gynecology and Obstetrics, 294(3), 519-523. doi:10.1007/s00404-016-4025-x
[3] Put A Cup In It. (2019, October 07). Menstrual Cup Firmness Guide
[4] Menstrual Cup Stems and their purpose: MCA Online Australia (2019, November 18).
[5] Barnhart, K. T., et al. (2006). Baseline dimensions of the human vagina.
[6]Eskenazi, B., & Warner, M. L. (1997). Epidemiology of endometriosis. Obstetrics and gynecology clinics of North America, 24(2), 235-258.
[7] Mitchell, M. A., Bisch, S., Arntfield, S., & Hosseini-Moghaddam, S. M. (2015). A confirmed case of toxic shock syndrome associated with the use of a menstrual cup. Canadian Journal of Infectious Diseases and Medical Microbiology, 26.
To collect the menstrual effluent needed for the non-invasive diagnostic panel, we designed an endometriosis-friendly menstrual cup. This cup takes into account the thickening of the vaginal wall and pelvic muscle fatigue that are commonly experienced by women with endometriosis, making menstrual cups painful [1, 2].
- Thin wall and inward facing rim to reduce pressure [3]
- Rounded stem for easy grip and comfort [4]
- Bent cup body for easy insertion [5]
It also can collect up to 40 mL of menstrual effluent compared to the average market design of 25 mL to cater to heavy menstruation, a common symptom of endometriosis [6].
UV-C Sterilizer
Bacterial growth on menstrual cups can cause a lethal condition known as Toxic Shock Syndrome [7]. To provide tools for sterilizing our Endo cup in areas with limited access to clean water, we designed a compact portable sterilizer that utilizes UV-C light [7].
DIY Centrifuge
Prior to using menstrual effluent in our diagnostic, the sample must be spun to separate out the serum. In order to reduce the cost of sample preparation for our LFA, we designed a replicable DIY centrifuge through recycling toys, computer parts and simple circuitry.
References
[1] Yang, N. (n.d.). Endometriosis: Radiology Reference Article. Retrieved from https://radiopaedia.org/articles/endometriosis
[2] Bispo, A. P., Ploger, C., Loureiro, A. F., Sato, H., Kolpeman, A., Girão, M. J., & Schor, E. (2016). Assessment of pelvic floor muscles in women with deep endometriosis. Archives of Gynecology and Obstetrics, 294(3), 519-523. doi:10.1007/s00404-016-4025-x
[3] Put A Cup In It. (2019, October 07). Menstrual Cup Firmness Guide
[4] Menstrual Cup Stems and their purpose: MCA Online Australia (2019, November 18).
[5] Barnhart, K. T., et al. (2006). Baseline dimensions of the human vagina.
[6]Eskenazi, B., & Warner, M. L. (1997). Epidemiology of endometriosis. Obstetrics and gynecology clinics of North America, 24(2), 235-258.
[7] Mitchell, M. A., Bisch, S., Arntfield, S., & Hosseini-Moghaddam, S. M. (2015). A confirmed case of toxic shock syndrome associated with the use of a menstrual cup. Canadian Journal of Infectious Diseases and Medical Microbiology, 26.
Diagnostic Design
Lateral Flow Assay Test Panel Development
We developed lateral flow assays (LFA) to measure levels of endometriosis biomarkers in menstrual blood as a diagnostic test for endometriosis [1].
A lateral flow assay (LFA) detects the presence of target molecules in a sample using a sandwich-style immunoassay approach.
Normally, these labeled antibodies are labeled using dye. However, our team found that conjugating these antibodies to gold nanoparticles (GNP) significantly increased the sensitivity of our design to the range of our desired concentrations with a detection threshold of less than 0.1 ng/mL [1].
Schematic representation of our lateral flow assay design
Biomarker Selection
We used combined log odds ratios to identify three biomarkers, out of 12 reported in literature, with the highest correlation to endometriosis: IGFBP-1, CA125, and IL-6 [3-9]. We also selected IL-1β, TNF-𝛼, and IL-8 (which are targets of FDA-approved pain and inflammation treatment [10] to inform physicians of potential therapies to relieve endometriosis symptoms.
Top 15 combinations of biomarkers with highest combined log odds ratio
Selection of Antibody Pairs With Non-overlapping Binding Sites
We used molecular modeling software Rosetta to identify antibody pairs with non-overlapping epitopes to capture biomarkers on our assays.
The biomarker IL1B (yellow) with detector-receptor pair Gevokizumab (teal) and Canakinumab (red).
LFA Model for Computational Design Optimization
We created a functional model of our LFAs with partial differential equations (PDE) to optimize assay design [11].
Results from LFA model
References
[1] Teerinen, T., Lappalainen, T., & Erho, T. (2014). A paper-based lateral flow assay for morphine. Analytical and Bioanalytical Chemistry, 406(24), 5955–5965. https://doi.org/10.1007/s00216-014-8001-7
[2] Sajid, M., Kawde, A.-N., & Daud, M. (2015). Designs, formats and applications of lateral flow assay: A literature review. Journal of Saudi Chemical Society, 19(6), 689–705. https://doi.org/https://doi.org/10.1016/j.jscs.2014.09.001
[3] Bedaiwy, M. A., Falcone, T., Sharma, R. K., Goldberg, J. M., Attaran, M., Nelson, D. R., & Agarwal, A. (2002). Prediction of endometriosis with serum and peritoneal fluid markers: a prospective controlled trial. Human reproduction, 17(2), 426-431.
[4] Galo, S., Zubor, P., Szunyogh, N., Kajo, K., Machalekova, K., Biringer, K., & Visnovský, J. (2005). TNF-alpha serum levels in women with endometriosis: prospective clinical study. Ceská Gynekologie, 70(4), 286-290.
[5] Kitawaki, J., Ishihara, H., Koshiba, H., Kiyomizu, M., Teramoto, M., Kitaoka, Y., & Honjo, H. (2005). Usefulness and limits of CA-125 in diagnosis of endometriosis without associated ovarian endometriomas. Human reproduction, 20(7), 1999-2003.
[6] Malutan, A. M., Drugan, T., Costin, N., Ciortea, R., Bucuri, C., Rada, M. P., & Mihu, D. (2015). Pro-inflammatory cytokines for evaluation of inflammatory status in endometriosis. Central-European journal of immunology, 40(1), 96.
[7] Mosbah, A., Nabiel, Y., & Khashaba, E. (2016). Interleukin-6, intracellular adhesion molecule-1, and glycodelin A levels in serum and peritoneal fluid as biomarkers for endometriosis. International Journal of Gynecology & Obstetrics, 134(3), 247-251.
[8] Ohata, Y., Harada, T., Miyakoda, H., Taniguchi, F., Iwabe, T., & Terakawa, N. (2008). Serum interleukin-8 levels are elevated in patients with ovarian endometrioma. Fertility and sterility, 90(4), 994-999.
[9] Vodolazkaia, A., Yesilyurt, B. T., Kyama, C. M., Bokor, A., Schols, D., Huskens, D., ... & Lambrechts, D. (2016). Vascular endothelial growth factor pathway in endometriosis: genetic variants and plasma biomarkers. Fertility and sterility, 105(4), 988-996.
[10] Animal Cell Technology Industrial Platform. (2020). Monoclonal Antibodies Approved by the EMA and FDA for Therapeutic Use. Retrieved November 08, 2020, from https://www.actip.org/products/monoclonal-antibodies-approved-by-the-ema-and-fda-for-therapeutic-use/
[11] Liu, Z., Hu, J., Li, A., Feng, S., Qu, Z., & Xu, F. (2017). The effect of report particle properties on lateral flow assays: a mathematical model. Sensors and Actuators B: Chemical, 248, 699-707.
We developed lateral flow assays (LFA) to measure levels of endometriosis biomarkers in menstrual blood as a diagnostic test for endometriosis [1].
A lateral flow assay (LFA) detects the presence of target molecules in a sample using a sandwich-style immunoassay approach.
- Add menstrual effluent to the sample pad that contains a fixed concentration of labeled antibodies on the membrane
- Sample flows and the labeled antibodies bind to their target molecules
- The target molecule is bound to by a second immobilized antibody at the test line
- Free labeled antibodies continue to travel down the strip to bind to immobilize antibodies at the control line [2]
Normally, these labeled antibodies are labeled using dye. However, our team found that conjugating these antibodies to gold nanoparticles (GNP) significantly increased the sensitivity of our design to the range of our desired concentrations with a detection threshold of less than 0.1 ng/mL [1].
Biomarker Selection
We used combined log odds ratios to identify three biomarkers, out of 12 reported in literature, with the highest correlation to endometriosis: IGFBP-1, CA125, and IL-6 [3-9]. We also selected IL-1β, TNF-𝛼, and IL-8 (which are targets of FDA-approved pain and inflammation treatment [10] to inform physicians of potential therapies to relieve endometriosis symptoms.
Selection of Antibody Pairs With Non-overlapping Binding Sites
We used molecular modeling software Rosetta to identify antibody pairs with non-overlapping epitopes to capture biomarkers on our assays.
LFA Model for Computational Design Optimization
We created a functional model of our LFAs with partial differential equations (PDE) to optimize assay design [11].
References
[1] Teerinen, T., Lappalainen, T., & Erho, T. (2014). A paper-based lateral flow assay for morphine. Analytical and Bioanalytical Chemistry, 406(24), 5955–5965. https://doi.org/10.1007/s00216-014-8001-7
[2] Sajid, M., Kawde, A.-N., & Daud, M. (2015). Designs, formats and applications of lateral flow assay: A literature review. Journal of Saudi Chemical Society, 19(6), 689–705. https://doi.org/https://doi.org/10.1016/j.jscs.2014.09.001
[3] Bedaiwy, M. A., Falcone, T., Sharma, R. K., Goldberg, J. M., Attaran, M., Nelson, D. R., & Agarwal, A. (2002). Prediction of endometriosis with serum and peritoneal fluid markers: a prospective controlled trial. Human reproduction, 17(2), 426-431.
[4] Galo, S., Zubor, P., Szunyogh, N., Kajo, K., Machalekova, K., Biringer, K., & Visnovský, J. (2005). TNF-alpha serum levels in women with endometriosis: prospective clinical study. Ceská Gynekologie, 70(4), 286-290.
[5] Kitawaki, J., Ishihara, H., Koshiba, H., Kiyomizu, M., Teramoto, M., Kitaoka, Y., & Honjo, H. (2005). Usefulness and limits of CA-125 in diagnosis of endometriosis without associated ovarian endometriomas. Human reproduction, 20(7), 1999-2003.
[6] Malutan, A. M., Drugan, T., Costin, N., Ciortea, R., Bucuri, C., Rada, M. P., & Mihu, D. (2015). Pro-inflammatory cytokines for evaluation of inflammatory status in endometriosis. Central-European journal of immunology, 40(1), 96.
[7] Mosbah, A., Nabiel, Y., & Khashaba, E. (2016). Interleukin-6, intracellular adhesion molecule-1, and glycodelin A levels in serum and peritoneal fluid as biomarkers for endometriosis. International Journal of Gynecology & Obstetrics, 134(3), 247-251.
[8] Ohata, Y., Harada, T., Miyakoda, H., Taniguchi, F., Iwabe, T., & Terakawa, N. (2008). Serum interleukin-8 levels are elevated in patients with ovarian endometrioma. Fertility and sterility, 90(4), 994-999.
[9] Vodolazkaia, A., Yesilyurt, B. T., Kyama, C. M., Bokor, A., Schols, D., Huskens, D., ... & Lambrechts, D. (2016). Vascular endothelial growth factor pathway in endometriosis: genetic variants and plasma biomarkers. Fertility and sterility, 105(4), 988-996.
[10] Animal Cell Technology Industrial Platform. (2020). Monoclonal Antibodies Approved by the EMA and FDA for Therapeutic Use. Retrieved November 08, 2020, from https://www.actip.org/products/monoclonal-antibodies-approved-by-the-ema-and-fda-for-therapeutic-use/
[11] Liu, Z., Hu, J., Li, A., Feng, S., Qu, Z., & Xu, F. (2017). The effect of report particle properties on lateral flow assays: a mathematical model. Sensors and Actuators B: Chemical, 248, 699-707.
Interpreting Results
Clinical Predictive Software
To further noninvasive diagnostics, we used Random Forest machine learning algorithm [1] to create a model that predicts endometriosis risk with 85% accuracy using only symptoms and clinical history. We integrated this model into an endometriosis risk assessment software tool.
Screenshots of our drafts of a user-interface that integrates our predictive model into a user-friendly web server.
LFA Imaging Station
To lower the cost of imaging, we designed a visual imaging station that can be created from our instructions using Lego blocks, a lens, and simple circuitry. The imaging station holds a smartphone, which can run imaging software to quantify the colorimetric signal from our LFA.
Multiple images showing various angles of our LFA imaging station
Image of a phone capturing an image of a test strip on our LFA imaging station
References
[1] Breiman, L. (2001), Random Forests, Machine Learning 45(1), 5-32
To further noninvasive diagnostics, we used Random Forest machine learning algorithm [1] to create a model that predicts endometriosis risk with 85% accuracy using only symptoms and clinical history. We integrated this model into an endometriosis risk assessment software tool.
LFA Imaging Station
To lower the cost of imaging, we designed a visual imaging station that can be created from our instructions using Lego blocks, a lens, and simple circuitry. The imaging station holds a smartphone, which can run imaging software to quantify the colorimetric signal from our LFA.
References
[1] Breiman, L. (2001), Random Forests, Machine Learning 45(1), 5-32
Synthetic Biology
Plug and Play Antibody Production
Our project makes use of synthetic biology by synthesizing proteins and antibodies through the creation of genetically modified organisms. Genetically engineering organisms to produce our required antibodies would help lower the cost of an immunoassay design for both ourselves and future iGEM teams. Previous literature research has shown the efficacy of using a specially engineered E. coli strain, SHuffle, for the production of antibodies [1, 2]. Our team created BioBricks for the variable and constant regions of a therapeutic antibody used in our design, Siltuximab, including a mutated constant chain for efficient use in E. coli SHuffle. This produced a novel, plug-and-play method for others to produce monoclonal antibodies in an easy and efficient manner.
Workflow for producing plug and play antibodies
As a proof of concept, we collaborated with Linkoping’s 2020 iGEM Team who utilized our Plug and Play design to create antibodies
Our science manager, Emily Laskey, and modeling manager, Linh Hoang, working on our partnership with the Project Leader of the Linköpings Universitet team, Alexander Johansson, in a Zoom meeting.
Estrogen Biosensor
We created a biosensor gene circuit for estrogen to inform physician diagnosis and appropriate dosage of estrogen-targeting therapeutics for endometriosis [3]. We used Hill function to model gene expression of this circuit to optimize design and found that the use of a weak promoter with only one activating site and a high plasmid copy number produced the best results.
Schematic representation of our estrogen biosensor
References
[1] Lobstein, J., Emrich, C. A., Jeans, C., Faulkner, M., Riggs, P., & Berkmen, M. (2012). SHuffle, a novel Escherichia coli protein expression strain capable of correctly folding disulfide bonded proteins in its cytoplasm. Microbial Cell Factories, 11, 56. https://doi.org/10.1186/1475-2859-11-56
[2] Robinson, M.-P., Ke, N., Lobstein, J., Peterson, C., Szkodny, A., Mansell, T. J., Tuckey, C., Riggs, P. D., Colussi, P. A., Noren, C. J., Taron, C. H., DeLisa, M. P., & Berkmen, M. (2015). Efficient expression of full-length antibodies in the cytoplasm of engineered bacteria. Nature Communications, 6(1), 8072. https://doi.org/10.1038/ncomms9072
[3] Chantalat, E., Valera, M. C., Vaysse, C., Noirrit, E., Rusidze, M., Weyl, A., ... & Arnal, J. F. (2020). Estrogen Receptors and Endometriosis. International Journal of Molecular Sciences, 21(8), 2815.
Our project makes use of synthetic biology by synthesizing proteins and antibodies through the creation of genetically modified organisms. Genetically engineering organisms to produce our required antibodies would help lower the cost of an immunoassay design for both ourselves and future iGEM teams. Previous literature research has shown the efficacy of using a specially engineered E. coli strain, SHuffle, for the production of antibodies [1, 2]. Our team created BioBricks for the variable and constant regions of a therapeutic antibody used in our design, Siltuximab, including a mutated constant chain for efficient use in E. coli SHuffle. This produced a novel, plug-and-play method for others to produce monoclonal antibodies in an easy and efficient manner.
As a proof of concept, we collaborated with Linkoping’s 2020 iGEM Team who utilized our Plug and Play design to create antibodies
Estrogen Biosensor
We created a biosensor gene circuit for estrogen to inform physician diagnosis and appropriate dosage of estrogen-targeting therapeutics for endometriosis [3]. We used Hill function to model gene expression of this circuit to optimize design and found that the use of a weak promoter with only one activating site and a high plasmid copy number produced the best results.
References
[1] Lobstein, J., Emrich, C. A., Jeans, C., Faulkner, M., Riggs, P., & Berkmen, M. (2012). SHuffle, a novel Escherichia coli protein expression strain capable of correctly folding disulfide bonded proteins in its cytoplasm. Microbial Cell Factories, 11, 56. https://doi.org/10.1186/1475-2859-11-56
[2] Robinson, M.-P., Ke, N., Lobstein, J., Peterson, C., Szkodny, A., Mansell, T. J., Tuckey, C., Riggs, P. D., Colussi, P. A., Noren, C. J., Taron, C. H., DeLisa, M. P., & Berkmen, M. (2015). Efficient expression of full-length antibodies in the cytoplasm of engineered bacteria. Nature Communications, 6(1), 8072. https://doi.org/10.1038/ncomms9072
[3] Chantalat, E., Valera, M. C., Vaysse, C., Noirrit, E., Rusidze, M., Weyl, A., ... & Arnal, J. F. (2020). Estrogen Receptors and Endometriosis. International Journal of Molecular Sciences, 21(8), 2815.
Human Practices
Advice from Endometriosis Researchers and Physicians
Team members Zivile Vebraite, Heather Shi, Meghan Martin, and James Tang in a Zoom meeting with Dr. Gubbels
Team members Isabel Lopez-Molini, Meghan Martin, Emily Laskey, Gabe Isaacson, and Emily Schiller in a Zoom meeting with Dr. Flores
- Dr. Gubbels emphasized the need for a non-invasive diagnostic for endometriosis and an educational program for both the general public and medical professionals.
- Dr. Flores, Dr. Metz, and Dr. Gregsen taught us about current research into endometriosis and helped us select biomarkers that are sensitive and accurate enough for an in-clinic test.
- Representatives from Pixie Cup & RITVA, an NGO focusing on promoting sustainable female hygienic products in India, recommended the use of menstrual cup as the initial menstrual effluent collection method, and helped us confirm the potential of using UVC sterilization as an alternative for cleaning the cups.
- Ms. Zivku, -Diva Cup representative, confirmed the potential to use menstrual cups for initial menstrual blood collection and the accessibility aspect of using a UV Sterilizer as an alternative to the traditional sanitizing method.
Supporting the Synbio Community
Synthetic Biology Education
In collaboration with pedagogy expert Dr. Nicholas Hammond, we created remote synthetic biology courses for high school and elementary students in the Rochester City School District and Sonshine Summer Camp.
[Left] Team member Helen Shammas teaching our online course about biosensors and bioreporters
[Right] Team member Emily Schiller teaching an activity via Zoom with Mr. Birthwright’s class in the Rochester City School District
SAA and Inclusivity
Our Science as Art (SAA) initiative encourages science without language barriers. We also translated our project to over 10 languages including American Sign Language (ASL), to facilitate the Deaf community’s involvement in synthetic biology.
Rochester Institute of Technology student, Rose, signing our promotional video in ASL
[Left] iGEM Team iBowu’s submission to SAA
[Middle] One of our infographics translated to Swedish
[Right] An elementary school student’s submission to SAA
Biomarker database
We created an online biomarker database where iGEM teams can store their information about biomarkers as a resource for future iGEM teams who work on the same topic. So far, four iGEM [1-4] teams have contributed 32 biomarkers to the database.
References
[1] Linkopings 2020 iGEM Team
[2] MIT 2020 iGEM Team
[3] Thelassy 2020 iGEM Team
[4] Queen’s University at Kingston’s iGEM Team
In collaboration with pedagogy expert Dr. Nicholas Hammond, we created remote synthetic biology courses for high school and elementary students in the Rochester City School District and Sonshine Summer Camp.
[Right] Team member Emily Schiller teaching an activity via Zoom with Mr. Birthwright’s class in the Rochester City School District
SAA and Inclusivity
Our Science as Art (SAA) initiative encourages science without language barriers. We also translated our project to over 10 languages including American Sign Language (ASL), to facilitate the Deaf community’s involvement in synthetic biology.
[Middle] One of our infographics translated to Swedish
[Right] An elementary school student’s submission to SAA
Biomarker database
We created an online biomarker database where iGEM teams can store their information about biomarkers as a resource for future iGEM teams who work on the same topic. So far, four iGEM [1-4] teams have contributed 32 biomarkers to the database.
References
[1] Linkopings 2020 iGEM Team
[2] MIT 2020 iGEM Team
[3] Thelassy 2020 iGEM Team
[4] Queen’s University at Kingston’s iGEM Team
Acknowledgements and Sponsors
Sponsors
