Ganseerpel
Advisory Board, Senior Moderator
Sialidase Fusion Protein as a Novel Broad-Spectrum Inhibitor
of Influenza Virus Infection
of Influenza Virus Infection
Michael P. Malakhov,
1 Laura M. Aschenbrenner,1 Donald F. Smee,2 Miles K. Wandersee,2
Robert W. Sidwell,
2 Larisa V. Gubareva,3 Vasiliy P. Mishin,3 Frederick G. Hayden,3
Do Hyong Kim,
1 Alice Ing,1 Erin R. Campbell,1 Mang Yu,1 and Fang Fang1*
NexBio, Inc., 6330 Nancy Ridge Dr., Suite 105, San Diego, California 92121
1; Institute for Antiviral Research, Utah State University,
Logan, Utah 843222; and Division of Infectious Diseases and International Health, Department of Internal Medicine,
University of Virginia, Charlottesville, Virginia 229083
Logan, Utah 843222; and Division of Infectious Diseases and International Health, Department of Internal Medicine,
University of Virginia, Charlottesville, Virginia 229083
Received 9 November 2005/Returned for modification 13 January 2006/Accepted 1 February 2006
Influenza is a highly infectious disease characterized by recurrent annual epidemics and unpredictable
major worldwide pandemics. Rapid spread of the highly pathogenic avian H5N1 strain and escalating human
infections by the virus have set off the alarm for a global pandemic. To provide an urgently needed alternative
treatment modality for influenza, we have generated a recombinant fusion protein composed of a sialidase
catalytic domain derived from
major worldwide pandemics. Rapid spread of the highly pathogenic avian H5N1 strain and escalating human
infections by the virus have set off the alarm for a global pandemic. To provide an urgently needed alternative
treatment modality for influenza, we have generated a recombinant fusion protein composed of a sialidase
catalytic domain derived from
Actinomyces viscosus fused with a cell surface-anchoring sequence. The sialidase
fusion protein is to be applied topically as an inhalant to remove the influenza viral receptors, sialic acids, from
the airway epithelium. We demonstrate that a sialidase fusion construct, DAS181, effectively cleaves sialic acid
receptors used by both human and avian influenza viruses. The treatment provides long-lasting effect and is
nontoxic to the cells. DAS181 demonstrated potent antiviral and cell protective efficacies against a panel of
laboratory strains and clinical isolates of IFV A and IFV B, with virus replication inhibition 50% effective
concentrations in the range of 0.04 to 0.9 nM. Mouse and ferret studies confirmed significant in vivo efficacy
of the sialidase fusion in both prophylactic and treatment modes.
fusion protein is to be applied topically as an inhalant to remove the influenza viral receptors, sialic acids, from
the airway epithelium. We demonstrate that a sialidase fusion construct, DAS181, effectively cleaves sialic acid
receptors used by both human and avian influenza viruses. The treatment provides long-lasting effect and is
nontoxic to the cells. DAS181 demonstrated potent antiviral and cell protective efficacies against a panel of
laboratory strains and clinical isolates of IFV A and IFV B, with virus replication inhibition 50% effective
concentrations in the range of 0.04 to 0.9 nM. Mouse and ferret studies confirmed significant in vivo efficacy
of the sialidase fusion in both prophylactic and treatment modes.
Influenza, caused by infection with influenza virus A (IFV
A) and IFV B, carries enormous direct and indirect socioeconomic
impacts. Since 1997, a new avian IFV A virus of the
H5N1 type has been causing epidemics in wild birds, as well as
in domestic poultry. Alarmingly, human infections by this virus
are also on the rise. Thus far, 100 people have been confirmed
to be infected by the virus and 50 of them have died
(5). Evidence has also shown that since 1999, the H5N1 virus
has been evolving rapidly in ducks and has become increasingly
pathogenic in both chicken and mice (7). All of these are
serious warning signs that a pandemic may be imminent. No
vaccine is currently available against the future pandemic virus.
The neuraminidase inhibitor (NAI), oseltamivir, was linked to
a surprisingly high frequency of drug-resistant viruses in children
(26). Oseltamivir-resistant H5N1 virus has also been isolated
from a patient who had been taking the drug as a prophylactic
measure (27). For these reasons, it is imperative to
develop alternative approaches to prevent and treat influenza.
The host cell receptors for influenza A and B viruses are cell
surface sialic acids (20). The predominant type of sialic acids is
A) and IFV B, carries enormous direct and indirect socioeconomic
impacts. Since 1997, a new avian IFV A virus of the
H5N1 type has been causing epidemics in wild birds, as well as
in domestic poultry. Alarmingly, human infections by this virus
are also on the rise. Thus far, 100 people have been confirmed
to be infected by the virus and 50 of them have died
(5). Evidence has also shown that since 1999, the H5N1 virus
has been evolving rapidly in ducks and has become increasingly
pathogenic in both chicken and mice (7). All of these are
serious warning signs that a pandemic may be imminent. No
vaccine is currently available against the future pandemic virus.
The neuraminidase inhibitor (NAI), oseltamivir, was linked to
a surprisingly high frequency of drug-resistant viruses in children
(26). Oseltamivir-resistant H5N1 virus has also been isolated
from a patient who had been taking the drug as a prophylactic
measure (27). For these reasons, it is imperative to
develop alternative approaches to prevent and treat influenza.
The host cell receptors for influenza A and B viruses are cell
surface sialic acids (20). The predominant type of sialic acids is
N
-acetylneuraminic acid (Neu5Ac), which is the biosynthetic
precursor for most of the other types. In nature, Neu5Ac is
mostly linked to the penultimate galactose residues of carbohydrate
side chains via (2,3)- or (2,6)-linkages. Both
Neu5Ac (2,3)-Gal and Neu5Ac (2,6)-Gal molecules can be
recognized as a receptor by influenza viruses (44), but human
viruses prefer (2,6)-linked sialic acid, whereas avian and
equine viruses predominantly recognize (2,3)-linked sialic
precursor for most of the other types. In nature, Neu5Ac is
mostly linked to the penultimate galactose residues of carbohydrate
side chains via (2,3)- or (2,6)-linkages. Both
Neu5Ac (2,3)-Gal and Neu5Ac (2,6)-Gal molecules can be
recognized as a receptor by influenza viruses (44), but human
viruses prefer (2,6)-linked sialic acid, whereas avian and
equine viruses predominantly recognize (2,3)-linked sialic
acid (20). The human respiratory
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