About

Systems glycobiology enabled by innovations in mass spectrometry and chemical biology. Glycosylation is a dominant feature of extracellular phenotypes. Glycoproteins present distinct combinations of glycans and amino acids that create unique molecular surfaces to relay biological information in a variety of biological processes.

Research topics

• Biology
• Biochemistry
• Chemistry
• Cell biology
• Computational biology
• Immunology
• Organic chemistry
• Genetics
• Chromatography

Selected publications

• Understanding <i>m</i> / <i>z</i> Range Settings for MS/MS Scans: A Case Study with Intact Glycopeptides

  Journal of the American Society for Mass Spectrometry · 2026-02-24

  articleSenior authorCorresponding

  Effective glycopeptide identification with tandem mass spectrometry (MS/MS) often relies on both low mass-to-charge (m/z) ions derived from glycan-specific oxonium ions and higher m/z peptide fragment ions that retain glycan modifications. Thus, glycoproteomic experiments benefit from a wider MS/MS scan range, i.e., the breadth of m/z values measured in fragmentation spectra, than those typically used in nonmodified peptide analyses. Here, we explore the implications of breaking a common axiom for scan range settings called the “5–10–15 rule” used to maximize transmission of ion populations of interest. This 5–10–15 rule, which defines the upper m/z value for a scan as a multiple of the first m/z value, comes from fundamental requirements for stable ion trajectories, where voltage settings must balance retention of low m/z ions while also generating effective pseudopotential wells to trap high m/z ions. Adhering to this calculation for MS/MS scan range settings can reduce glycopeptide ion coverage by excluding the analysis of either low m/z oxonium ions or high m/z fragment ions. We use a quadrupole-Orbitrap-linear ion trap Tribrid MS system (Orbitrap Ascend) to investigate the implications of following or breaking the 5–10–15 rule in MS/MS scans for glycopeptide characterization with higher-energy collisional dissociation (HCD), electron-transfer dissociation (ETD), and electron-transfer/higher-energy collision dissociation (EThcD). For scans with a first m/z value around m/z 120 (i.e., capturing most common glycan-specific oxonium ions), we show that breaking the 5–10–15 rule does not lead to a significant loss of fragment ion transmission at either extreme of the m/z range. We use this case study to discuss the concepts important to using the 5–10–15 rule wisely and when it can be practically ignored, such as using large scan ranges to improve glycopeptide characterization.

  PublisherDOI

• MassIVE MSV000100989 - Colorectal Cancer Proteomics, Phosphoproteomics, and Glycoproteomics

  California Digital Library · 2026-01-01

  datasetOpen access1st authorCorresponding
  OA PDFDOI

• Collaboration between labs of Neus Agell and Nick Riley

  Open MIND · 2026-01-01

  other1st authorCorresponding

• Profiling Glycoproteins Enriched by Multinanoparticle Protein Corona

  Analytical Chemistry · 2026-01-08 · 1 citations

  articleSenior authorCorresponding

  Biofluids, such as plasma and cerebrospinal fluid (CSF), present significant challenges for proteome and glycoproteome analysis due to their complex protein composition and wide dynamic range of protein abundances. Nanoparticle (NP)-based enrichment methods that rely on the formation of protein coronas to compress dynamic range have emerged as promising approaches to improve the proteomic sampling depth in biofluids, yet implications of these methods for glycoproteome characterization remain underexplored. In this study, we investigate enrichment via multinanoparticle protein corona (abbreviated here as NP enrichment) with and without subsequent mixed-mode anion exchange (MAX) enrichment for characterizing N- and O-linked glycopeptides in human plasma and CSF. Using an automated commercial platform, we demonstrate that NP enrichment significantly enhances N-glycoproteome coverage, particularly in plasma, and increases detection of N-glycopeptides from low abundant glycoproteins. These N-glycoproteome improvements are further enhanced when combined with MAX enrichment. For the O-glycoproteome, NP enrichment alone outperformed NP+MAX, suggesting that MAX may preferentially enrich N-glycopeptides. We also generated the first application of multinanoparticle protein corona enrichment to analyze CSF, another biofluid of interest for biomarker discovery, and found that NP enrichment increased proteome and glycoproteome depth, with notable identification of extracellular vesicle protein biomarkers. Overall, this study highlights the synergy of protein level enrichment with MNPPC and glycopeptide enrichment for enhancing the glycoproteome coverage of human biofluids and provides a valuable framework for studying glycoproteins in complex biological samples.

  PublisherDOI

• TMPRSS2-mediated coronavirus spike activation and inhibition

  Nature Structural & Molecular Biology · 2026-04-28

  articleOpen access

  Abstract The protease TMPRSS2 facilitates coronavirus infections, yet its mechanism of viral glycoprotein recognition remains unclear. Here we show that, following ACE2 engagement of the SARS-CoV-2 spike (S) inducing the early fusion intermediate conformation (E-FIC), TMPRSS2 cleaves the R815 S 2 ′ site and promotes fusogenic conformational changes leading to viral entry. We unveil TMPRSS2 recognition of S 2 ′, identify key residues modulating binding specificity and demonstrate that S 2 ′ site-directed broadly neutralizing antibodies target E-FIC and inhibit viral entry by blocking TMPRSS2 access. We computationally designed stabilized E-FIC as a vaccine candidate, overcoming the transient nature of this state. We describe a TMPRSS2-directed monoclonal antibody inhibiting several coronaviruses, including SARS-CoV-2 variants and protecting mice against SARS-CoV-2 challenge. These results outline the mechanistic role of TMPRSS2 and S 2 ′ site-directed antibodies in coronavirus entry.

  PublisherDOI

• P-349. Using Participatory Design to Establish a Workflow for Administration of Long-Acting Cabotegravir + Rilpivirine in Community Pharmacies

  Open Forum Infectious Diseases · 2026-01-01

  articleOpen access

  Abstract Background Long acting cabotegravir and rilpivirine (LA CAB/RPV), the first fully injectable antiretroviral therapy for people with HIV (PWH), is administered intramuscularly typically by a nurse in a clinic. Limitations to this approach include staff availability, office hour restrictions, travel distance, and HIV stigma. Alternate models of care are needed to address these obstacles expanding access to LA CAB/RPV. We used a human-centered participatory approach to co-design a workflow adapting in clinic administration of LA CAB/RPV to community pharmacies. Workflow for Long-acting Cabotegravir/Rilpivirine Administration in Community Pharmacies Methods Our design team consisted of 12 members, including HIV providers, pharmacists, nurses, informaticists, and PWH receiving LA CAB/RPV. The team met weekly for 5 sessions to delineate current practices and iteratively produce a modified workflow. We assessed barriers and facilitators of co-design through semi-structured interviews with design team members. A phenomenologically informed codebook thematic analysis identified key themes across interviews. Results Factors identified as essential to the in clinic model of LA CAB/RPV administration included care coordination utilizing electronic health record reports to track patient follow-up, communication among the team and with the patient, and injection visit experience. The figure shows the workflow adapted to community pharmacies. Next, from interviews with 6 design team members, we identified 4 themes of successful co-design. First, meeting modality created essential infrastructure for collaboration with in-person interactions enabling more effective engagement. Second, strategic leadership shaped the process with atomization of a complex workflow into manageable steps which involved work between meetings to frame and organize discoveries. Third, team composition and inclusivity enriched the process where multidisciplinary representation contributed critical insights. Last, collaborative humility fostered participation across power differentials aiding the integration of expertise from all participants. Conclusion Using co-design, we were able to adapt the workflow for in clinic administration of LA CAB/RPV to community pharmacies. Structural, compositional and interpersonal team factors contributed to the successful co-design process. Disclosures Melissa O. Jenkins, Meliisa Jenkins, Gilead Sciences, Inc.: Grant/Research Support Corrilynn O. Hileman, MD, MS, Gilead: Grant/Research Support|ViiV: Grant/Research Support

  PublisherDOI

• Abstract 7655: Defining molecular interfaces of glyco-immune checkpoint ligands using tandem mass spectrometry and ion-ion reactions.

  Cancer Research · 2026-04-03

  articleSenior author

  Abstract The Siglec (sialic acid-binding immunoglobulin-like lectin) family of glycan-binding immune receptors are emerging as attractive targets for cancer immunotherapy. Siglecs recognize cell surface glycoproteins through complex interactions with various glycans containing sialic acid residues, which are upregulated in numerous malignancies. Engagement of Siglecs by their sialoglyco-ligands elicits an inhibitory signaling cascade using the same immunoreceptor tyrosine-based inhibitory motif (ITIM) signaling domains shared by established immune checkpoint receptors (e.g., PD-1, CTLA-4, and SIRPα). Our understanding of the mechanisms behind Siglec-mediated immune evasion has been rapidly improving, largely due to efforts to define transcriptional and metabolic programs that promote Siglec ligand expression. Even so, unknown combinations of glycan and protein epitopes (i.e., the glycocode) that guide Siglec ligand recognition are a major hurdle for exploiting Siglecs as therapeutic targets. Molecular details of specific glycocode patterns that contribute to Siglec ligand recognition are needed to target immune modulatory glycoforms in tumor-specific microenvironments while minimizing off-target effects, but these heterogenous modifications states challenge traditional analytical approaches. Here, we describe our efforts to develop cutting-edge mass spectrometry (MS) data acquisition technologies that incorporate electron transfer dissociation (ETD) and proton transfer charge reduction (PTCR) ion-ion reactions to profile highly complex Siglec ligand glycoproteins. We use online and offline MS workflows in combination with bottom-up glycoproteomics to profile highly heterogeneous intact glycoproteoforms of Siglec ligands that have emerged as attractive targets for next-generation glyco-immunotherapies. PTCR readily isolated and charge-reduced proteoforms of interest, generating pseudo-native spectra. PTCR scans of interest were identified, and spectra were cleaned using in-house software, PTCRcleaner. Analysis of both intact approaches was supported by bottom-up mass spectrometry data collected using a sceHCD-pd-EThcD approach. We also applied this workflow to characterize other glycosylated immune checkpoint ligands, e.g., PD-L1. Altogether, our novel tandem MS approaches that uniquely leverage ion-ion reactions enable the characterization of highly heterogeneous glycoproteoform repertoires, contributing to a deeper understanding of the complexity of the glycocode and how these proteins contribute to heterogeneity in the tumor-immune synapse. Citation Format: Tim S. Veth, Haley M. Schramm, Nicholas M. Riley, . Defining molecular interfaces of glyco-immune checkpoint ligands using tandem mass spectrometry and ion-ion reactions [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 7655.

  PublisherDOI

• The protease cathepsin K can debulk the cancer glycocalyx

  Journal of Biological Chemistry · 2026-01-23 · 1 citations

  articleOpen access

  The cancer glycocalyx is characterized by the overexpression of large glycoconjugates, including mucins, proteoglycans, and polysialic acid, which collectively increase glycocalyx thickness and stiffness to promote tumor survival, metastasis, and progression. Previous work demonstrated that enzymatic degradation of cancer cell-surface mucins reduced tumor burden and metastasis in mouse models of breast cancer, thus validating glycocalyx remodeling as a therapeutic strategy. However, this work relied on an engineered bacterial mucin-selective protease, or mucinase, which does not degrade other bulky glycoconjugates and raises immunogenicity concerns because of its bacterial origin. A human enzyme that can degrade cell-surface mucins and other bulky glycoconjugates would address both limitations. We screened all 15 human cathepsins for their ability to degrade purified and cell-surface mucins, since multiple cathepsins have previously been shown to degrade mucins within regions of dense glycosylation. We found that cathepsin K (CTSK) uniquely degrades cell-surface mucins, proteoglycans, and polysialylated glycoproteins, and we demonstrated that CTSK reduces total glycocalyx thickness. These findings establish CTSK as a promising starting point for the development of a glycocalyx-debulking enzyme for cancer therapeutics.

  PublisherOA PDFDOI

• Extracting Informative Glycan-Specific Ions From Glycopeptide MS/MS Spectra With GlyCounter

  Molecular & Cellular Proteomics · 2025-10-15 · 2 citations

  articleOpen accessSenior author

  Glycopeptide tandem mass spectra typically contain numerous glycan-specific fragments that can inform several features of glycan modifications, including glycan class, composition, and structure. While these fragment ions are often straightforward to observe by eye, few tools exist to systemically explore these common glycopeptide spectral features or explore their relationships with each other. Instead, most studies rely on manual inspection to understand glycan-informative ion content in their data, or they are restricted to evaluating the presence of these ions only in the small fraction of spectra that are identified by glycopeptide search algorithms. Here we introduce GlyCounter as a freely available, open-source tool to rapidly extract oxonium, Y-type, and custom ion information from raw data files. We highlight GlyCounter's utility by evaluating glycan-specific fragments in a diverse selection of publicly available datasets to demonstrate how others in the field can make immediate use of this software. In several cases, we show how conclusions drawn in these publications are evident simply through GlyCounter's extracted ion information without requiring database searches or experiment-specific programs. Although one of our goals is to decouple spectral evaluation from glycopeptide identification, we also show that evaluating oxonium ion content with GlyCounter can supplement a database search as valuable spectral evidence to validate conclusions. In all, we present GlyCounter as a user-friendly platform that can be easily incorporated into most glycoproteomic workflows to refine sample preparation, data acquisition, and post-acquisition identification methods through straightforward evaluation of the glycan content of glycoproteomic data. Software and instructions are available at https://github.com/riley-research/GlyCounter.

  PublisherOA PDFDOI

• Antibody-lectin chimeras for glyco-immune checkpoint blockade

  Nature Biotechnology · 2025-12-16 · 4 citations

  articleOpen access

  Despite the curative potential of checkpoint blockade immunotherapy, many patients remain unresponsive to existing treatments. Glyco-immune checkpoints, which involve interactions of cell-surface glycans with lectin, or glycan-binding, immunoreceptors, have emerged as prominent mechanisms of immune evasion and therapeutic resistance in cancer. Here, we describe antibody-lectin chimeras (AbLecs), a modular system for glyco-immune checkpoint blockade. AbLecs are bispecific antibody-like molecules comprising a cell-targeting antibody domain and a lectin 'decoy receptor' domain that directly binds glycans and blocks their ability to engage inhibitory lectin receptors. AbLecs potentiate cancer cell destruction by primary human immune cells in vitro and reduce tumour burden in a humanized, immunocompetent mouse model, outperforming most existing therapies and combinations tested. By targeting a distinct axis of immunological regulation, AbLecs synergize with blockade of established immune checkpoints. AbLecs can be readily designed to target numerous tumours and immune cell subsets as well as glyco-immune checkpoints, thus representing a potential modality for cancer immunotherapy.

  PublisherDOI

Recent grants

• Uniting Mass Spectrometry and Glycoscience to Investigate Cancer Biology

  NIH · $88k · 2018–2023

• Capturing the Holistic Glycocode through Systems Glycobiology

  NIH · $498k · 2022–2027

• Uniting Mass Spectrometry and Glycoscience to Investigate Cancer Biology

  NIH · $65k · 2016–2018

• Capturing the Holistic Glycocode through Systems Glycobiology

  NIH · $100k · 2022–2023

Frequent coauthors

• Carolyn R. Bertozzi

  Stanford University

  147 shared

• Joshua J. Coon

  Morgridge Institute for Research

  106 shared

• Michael S. Westphall

  Quantitative BioSciences

  43 shared

• Kayvon Pedram

  Janelia Research Campus

  30 shared

• Sharon J. Pitteri

  Stanford University

  24 shared

• D. Judy Shon

  California Institute of Technology

  24 shared

• Amato J. Giaccia

  CRUK/MRC Oxford Institute for Radiation Oncology

  23 shared

• Stacy A. Malaker

  Yale University

  22 shared

Labs

• Riley Research Group at UWPI

Education

• Ph.D. in Analytical Chemistry, Department of Chemistry, Joshua J. Coon Group

  University of Wisconsin Madison

  2018

• B.S. in Chemistry and Psychology, with Honors from the South Carolina Honors College, College of Arts and Sciences

  University of South Carolina

  2012

Awards & honors

• American Society for Mass Spectrometry (ASMS) Research Award…
• Scialog Fellow, Automating Chemical Laboratories, 2024
• Human Proteome Organization (HUPO) Rising Star Award, 2023
• Rising Star in Glycoscience, Translational Glycomics Center,…
• Distinguished Young Honors Alumni Award, South Carolina Hono…