Safety and Pharmacokinetics of Intranasally Administered Heparin (preprint)

Purpose Intranasally administered unfractionated heparin (UFH) and other sulfated polysaccharides are potential prophylactics for COVID-19. The purpose of this research was to measure the safety and pharmacokinetics of clearance of intranasally administered UFH solution from the nasal cavity. Methods Double-blinded daily intranasal dosing in C57Bl6 mice with four doses (60 ng to 60 ug) of UFH was carried out for fourteen consecutive days, with both blood coagulation measurements and subject adverse event monitoring. The pharmacokinetics of fluorescent-labeled UFH clearance from the nasal cavity were measured in mice by in vivo imaging. Intranasal UFH at 2000 U/day solution with nasal spray device was tested for safety in a small number of healthy human subjects. Results UFH showed no evidence of toxicity in mice at any dose measured. No significant changes were observed in activated partial thromboplastin time (aPTT), platelet count, or frequency of minor irritant events over vehicle-only control. Human subjects showed no significant changes in aPTT time, international normalized ratio (INR), or platelet count over baseline measurements. No serious adverse events were observed. In vivo imaging in a mouse model showed a two-phase clearance of UFH from the nasal cavity. After 12 hours, 2.1% of the initial administered UFH remained in the nasal cavity, decaying to background levels after 24 hours. Conclusions UFH showed no toxic effects for extended daily intranasal dosing in mice as well as humans. The clearance kinetics of intranasal heparin solution from the nasal cavity indicates potentially protective levels for up to 12 hours after dosing.

Effective Inhibition of SARS-CoV-2 Entry by Heparin and Enoxaparin Derivatives (preprint)

Severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) has caused a pandemic of historic proportions and continues to spread globally, with enormous consequences to human health. Currently there is no vaccine, effective therapeutic or prophylactic. Like other betacoronaviruses, attachment and entry of SARS-CoV-2 is mediated by the spike glycoprotein (SGP). In addition to its well-documented interaction with its receptor, human angiotensin converting enzyme 2 (hACE2), SGP has been found to bind to glycosaminoglycans like heparan sulfate, which is found on the surface of virtually all mammalian cells. Here, we pseudotyped SARS-CoV-2 SGP on a third generation lentiviral (pLV) vector and tested the impact of various sulfated polysaccharides on transduction efficiency in mammalian cells. The pLV vector pseudotyped SGP efficiently and produced high titers on HEK293T cells. Various sulfated polysaccharides potently neutralized pLV-S pseudotyped virus with clear structure-based differences in anti-viral activity and affinity to SGP. Concentration-response curves showed that pLV-S particles were efficiently neutralized by a range of concentrations of unfractionated heparin (UFH), enoxaparin, 6-O-desulfated UFH and 6-O-desulfated enoxaparin with an IC50 of 5.99 {micro}g/L, 1.08 mg/L, 1.77 {micro}g/L, and 5.86 mg/L respectively. The low serum bioavailability of intranasally administered UFH, along with data suggesting that the nasal epithelium is a portal for initial infection and transmission, suggest that intranasal administration of UFH may be an effective and safe prophylactic treatment.