(4gz) Leveraging Bottlebrush Polymers to Design Tissue-Specific Synthetic Extracellular Matrix-Mimics

The macromolecular architectures and mechanical properties which define the extracellular matrix (ECM) of cells are tissue-specific and respond to disease in a sex- and age-dependent manner. To study these dynamic transformations with user-directed control, we will need to develop accurate in vitro systems to investigate how biomechanical and biochemical extracellular matrix cues guide cellular functions in healthy and diseased tissues. My graduate and postdoctoral training in polymer chemistry at the University of Minnesota with Profs. Theresa Reineke and Frank Bates and in biomaterials at the University of Colorado Boulder with Prof. Kristi Anseth, respectively, has provided me with an interdisciplinary skill set at the interface of chemistry, chemical and biological engineering, and materials science. This has equipped me to lead a future lab investigating cell-ECM interactions using expertise in polymer and hydrogel synthesis and characterization applied to questions within the musculoskeletal tissue niche. My future research interests focus on 1) harnessing controlled polymerization techniques to engineer synthetic materials which recapitulate the mechanical properties and functions of tissue niche-specific ECM components. In doing so, we will be able to investigate the structure-property relationships of macromolecular architectures, such as graft and bottlebrush polymers, incorporated into hydrogels and networks. These materials can then be applied to 2) study how cells respond to user-defined biomechanical and biochemical cues in 3D culture. Then, 3) we will study how bottlebrush polymers can be used to develop sulfated proteoglycan-mimetic macromolecules to study the role of proteoglycans in tissues and develop therapies for diseases in which proteoglycans breakdown, such as osteoarthritis.

Research Experience:

Nonlinear Elastic Bottlebrush Polymer Hydrogels, Department of Chemical and Biological Engineering, BioFrontiers Institute, CU Boulder (advised by Kristi S. Anseth)

As a postdoctoral fellow, I have been able to harness my abilities as a polymer chemist to synthesize poly(ethylene glycol)-based bottlebrush polymer hydrogels that recapitulate the nonlinear elastic properties of proteinaceous hydrogels such as, collagen and fibrin. I have demonstrated that this thiol-ene photo-crosslinked hydrogel can be used as a 3D culture scaffold for bone marrow-derived human mesenchymal stem/stromal cells (hMSCs). Through rheological characterization, I have proven that we can tune the strain-stiffening mechanical properties of these materials by controlling the length of the bottlebrush dithiol crosslinker in the network while limiting the stress-relaxation. This in turn allowed me to tune the morphological phenotype of hMSCs as a function of the amount of stress needed to be exerted by the cells to elicit a strain-stiffening material response. Through this, we elucidated that the strain-stiffening mechanical response promoted the formation of mature cell-matrix adhesions resulting in contractile cell responses which were measured through 3D traction force microscopy. We identified that the formation of spindle-like protrusions by hMSCs cultured within the most strain-stiffening bottlebrush polymer hydrogels was mediated through Arp 2/3 which is involved in actin branching and focal adhesion maturation. These materials are currently being applied as osteoid-mimetic microenvironments to support the differentiation of hMSCs into osteocytes, which are the most prevalent cell type in bone.

End-Group Modified Bottlebrush Polymers to Deliver Active Pharmaceutical Ingredients, Department of Chemistry, UMN (advised by Frank S. Bates and Theresa M. Reineke)

Oral drug delivery is the most common form of medication administration worldwide, but over 60% of active pharmaceutical ingredients in the pharmaceutical pipeline are impacted by insolubility in the gastrointestinal tract. This work provided the first investigation of bottlebrush polymers for hydrophobic non-covalent sequestration and solubilization of pharmaceuticals and is patented by the US Patent office. Poly(N-isopropylacrylamide-stat-N,N-dimethylacrylamide) bottlebrush copolymers were synthesized using ring-opening metathesis polymerization and the thermoresponsive properties were modulated using post-polymerization end-group modification. The bottlebrushes out-performed linear copolymer analogues at solubilizing model small molecule therapeutics. This was attributed to the fact that bottlebrush copolymers exist in solution as unimolecular nanoparticles with synthetically defined chain densities and are more stable in solution than their linear copolymer analogues. These findings were then expanded to work with an orally administered breast cancer therapeutic, GDC-0810. Next, expanding the scope of deliverable payloads, we demonstrated how cationic bottlebrush polymers could be used to complex and deliver pDNA. Drawing inspiration from the end-group modification work optimized for applications in oral drug delivery, we investigated how systematically increasing the hydrophilicity of the cationic bottlebrush end-groups would impact the complexation, toxicity, and transfection efficiency when delivering both pDNA and CRISPR-Cas9 payloads. It was found that increasing hydrophilicity of bottlebrush polymer end-groups decreased toxicity while also decreasing transfection efficiency. Therefore, we identified the intermediate hydrophilicity needed to optimize the balance of transfection efficiency and toxicity with both payloads.

Successful Proposals:

• NIH T32 Fellowship “Interdisciplinary Training in Musculoskeletal Research” (2022-2024)
• NSF Graduate Research Fellowship Program Awardee (2017-2021)

Selected Publications and Patents (10 peer-reviewed publications total, 6 first/co-first author, 1 US patent, 1 perspective):

1. Ohnsorg, M. L.; Mash, K. M.; Khang, A.; Rao, V. V.; Bera, K.; Kirkpatrick, B. E.; Anseth, K. S. “Nonlinear Elastic Bottlebrush Polymer Hydrogels Modulate Actomyosin Mediated Protrusion Formation in Mesenchymal Stromal Cells,” Mater. 2024, 2403198.
2. Reineke, T.; Bates, F. S.; Ohnsorg, M. L. “Macromonomers and bottle brush polymers for delivery of biological agents” U.S. Patent No. 11,986,533, May 21st, 2024.
3. Ohnsorg, M. L.⁼; Barr, K. E.⁼; Liberman, L.; Corcoran, L.G.; Sarode, A.; Nagapudi, K.; Feder, C. R.; Bates, F. S.; Reineke, T. M. “Drug-Polymer Nanodroplet Formation and Morphology Drive Solubility Enhancement of GDC-0810,” Bioconjugate Chem. 2024, 35, 4, 499-516.
4. Ohnsorg, M. L.;⁼ Dalal, R.J.;⁼ Panda, S.; Reineke, T. M. “Hydrophilic Surface-Modification of Cationic Unimolecular Bottlebrush Vectors Moderate pDNA and RNP Bottleplex Stability and Delivery Efficacy,” Biomacromolecules 2022, 23, 12, 5179-5192.
5. Ohnsorg, M. L.; Prendergast, P. C.; Robinson, L. L.; Bockman, M. R.; Bates, F. S.; Reienke, T. M. “Bottlebrush Polymer Excipients Enhance Drug Solubility: Influence of End-Group Hydrophilicity and Thermoreponsiveness,” ACS Macro Lett. 2021, 10, 375-381.
6. Ohnsorg, M. L.; Ting, J. M.; Jones, S. D.; Jung, S.; Bates, F. S.; Reineke, T. M. “Tuning PNIPAm self-assembly and thermoresponse: roles of hydrophobic end-groups and hydrophilic comonomer,” Polym. Chem. 2019, 10, 3469-3479.

Selected Awards:

• Inaugural ACS Biomacromolecules Early Career Board Member (2023-present)
• RCS Polymer Chemistry Outstanding Peer Reviewer of 2023
• ACS Polymeric Materials Science and Engineering Future Faculty Scholar (2023)
• Best Postdoctoral Research Award – 2^nd place, Society for Biomaterials (2023)
• Best Poster Award – ACS POLY: Polymers in Medicine and Biology (2022)
• Eastman Chemical Student Award in Applied Polymer Science, ACS PMSE (2020)
• CAS Future Leaders (2020)
• Beckman Scholar (2014-2015)

Teaching Interests: Learning alongside students will always be one of the things that brings me the greatest fulfillment as a scientist and educator, and I look forward to a career filled with asking tough questions and discovering the answers together with undergraduate and graduate students. My goal as an educator is to instill the importance of critical thinking skills through the development of dynamic and engaging inquiry-focused lectures. Although my undergraduate and graduate degrees are in Chemistry, I have always been enrolled in courses at the interface of the chemistry and chemical engineering departments. With this interdisciplinary training, I feel confident in my ability to teach a range of courses undergraduate courses such as, Chemical Kinetics, Thermodynamics, Fluid Mechanics, and Transport Phenomena and graduate core/elective courses such as, Polymer Synthesis, Polymer Physics, Biomaterials, Tissue Engineering. As well as service classes coded towards engineers such as General Chemistry or Organic Chemistry for Engineers.

The most rewarding aspect of pursuing a career as a tenure-track professor at an R1 institution is the ability to mentor students and participate in outreach activities that breakdown the scientific language barriers that are often intimidating for younger trainees. I have been fortunate to be mentored by strong scientists throughout my career that wanted me to not only excel in science but also grow as a human being. It is important to me to become a strong female mentor for my future trainees and build a lab with a core culture of inclusion, communication, and respect. With this foundation, I will cultivate an environment in which collaboration, innovation, and creative problem-solving will flourish.

Teaching Experience:

• General Chemistry for Engineers II Lecture, Co-Instructor (Postdoctoral Teaching Fellow)
• General Chemistry I and II Lab, Teaching Assistant
• Polymer Chemistry Lecture, Teaching Assistant

Service/Leadership:

• Peer Reviewer for the following journals: Polymer Chemistry, Chemical Science, Biomacromolecules, Bioconjugate Chemistry, Molecular Pharmaceutics
• Secretary of Postdoctoral Association of Colorado Boulder (2022-2023)
• President of ACS POLY/PMSE Student Chapter, UMN (2018-2020)
• National Graduate Research Polymer Conference Planning Committee (2018)

Outreach:

I am passionate about inspiring the next generation of researchers and their families to engage with science and engineering concepts in tangible ways. I have been committed to outreach efforts throughout my career shown through the select following examples:

• Colorado State Science and Engineering Fair Mentor – high school mentee awarded 1^st prize from ACS and 2^nd Grand Chemistry Prize (2022-2023)
• CU Science Ambassador, developed and presented “Not all plastics are the same!” demo (2022)
• ColorMePhD Coloring Book Page Guest Scientist and Illustrator, “Polymer Parachutes” (2020-2021)
• Open-Door Learning Center, developed science lab supplements for GED curriculum (2016-2018)