# NET stabilization: mechanistic and therapeutic studies in thromboinflammatory disoders

> **NIH NIH K99** · CHILDREN'S HOSP OF PHILADELPHIA · 2021 · $167,400

## Abstract

ABSTRACT
 In response to inflammatory stimuli, neutrophils (PMN) extrude neutrophil extracellular traps (NETs), webs
of negatively-charged cell-free (cf) DNA complexed with positively-charged histones, which ensnare pathogens
but also damage host tissue, contributing to diseases including sepsis, in which a large burden of NETs is
acutely produced causing endothelial damage and organ dysfunction, and sickle cell disease (SCD), in which
there is a chronic increase in NET production that contributes to vascular inflammation and painful vaso-
occlusive episodes (VOE). Unfortunately, interventions that block NET release increase bacterial
dissemination, while treatments that degrade NETs can release entrapped microbes and toxic NET-
degradation products (NDPs) that contribute to multisystem organ damage. I posit that NET stabilization, in
which NETs are preserved but modified to reduce NDP release and enhance bacterial capture, may be
therapeutic in both sepsis and SCD. Platelet factor 4 (PF4) is a positively-charged chemokine released by
activated platelets that binds to and cross-aggregates polyanions like heparin and DNA. I have found that PF4
physically compacts NETs, increasing their resistance to nucleases. PF4 also binds to negatively-charged
molecules on the bacterial surface and markedly enhances their capture by NETs. KKO, a human (h)
PF4:heparin complex-binding monoclonal antibody (moAb), stabilizes PF4:NETs, further increasing nuclease
resistance. In murine sepsis models, hPF4 and an Fc-modified, deglycosylated KKO (DG-KKO), work in
concert to decrease NDP release, enhance bacterial capture, and improve outcomes. In this proposal, I will
compare the effect of NET stabilization in sepsis and SCD, to clarify its mechanism of action and assess if it is
protective in both acute and chronic NET release. Specific Aim (SA) #1: Define the mechanism(s) by which
hPF4 and DG-KKO stabilize NETs. I will evaluate how hPF4 and DG-KKO modify NETs, using hPF4 variants,
other cations, and anti-hPF4 moAbs in vitro and in murine sepsis models. SA#2: Define the mechanism(s) of
enhanced NET antimicrobial activity. I will define how PF4:NETs bind different classes of bacteria, assess if
NET stabilization enhances bacterial killing, and test whether other cations can replicate these effects. SA#3:
Determine whether NET stabilization is protective in SCD. I will define whether NET-targeted therapies
protective in SA#1, reduce cellular injury in microfluidic channels infused with plasma from SCD patients and in
a murine model of SCD. I will then measure the association between PF4, NDP levels, and disease severity in
SCD patients. I will pursue these studies within the context of a career development plan that combines
didactic courses, multidisciplinary mentorship, and interdepartmental collaboration. This work will enhance our
understanding of NETs and support a potential novel intervention for two distinct inflammatory disorders. It will
also facilitate my grow...

## Key facts

- **NIH application ID:** 10117771
- **Project number:** 1K99HL156060-01
- **Recipient organization:** CHILDREN'S HOSP OF PHILADELPHIA
- **Principal Investigator:** Kandace Gollomp
- **Activity code:** K99 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $167,400
- **Award type:** 1
- **Project period:** 2021-03-05 → 2026-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10117771

## Citation

> US National Institutes of Health, RePORTER application 10117771, NET stabilization: mechanistic and therapeutic studies in thromboinflammatory disoders (1K99HL156060-01). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10117771. Licensed CC0.

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