Key discovery may enable development of vaccines, therapeutics
BOSTON, Jan. 25, 2007 — Viruses whose genomes constantly mutate, such as the H5N1 "bird flu," are driving a race to find relatively unchanging segments of their genomes; successful identification of these segments may allow more successful vaccines and therapeutics to be developed.
Replikins are short fragments of the genomes of infectious organisms, which have been found to be related quantitatively to rapid replication and epidemic outbreaks (1,2). Scientists at Replikins, LLC have just identified a specific site in the human H5N1 virus genome which contains a dramatically higher concentration of replikins than the rest of the virus, and named it the Replikin Peak Gene(TM) (RPG).
Two of the replikin components of RPG have been found to be conserved over 88 years in the following high-mortality and pandemic virus strains: H1N1, H2N2, H3N2, H5N1, and H7N7. This discovery makes possible very specific targeting with vaccines and other treatments; these Replikin peptides also form the basis for a pan-strain influenza vaccine, for which trials are underway.
New quantitative virus protein sequence search software, FluForecast(R), available as a service from Replikins Ltd., was central to this work. RPG was isolated by comparing the replikin concentration (Replikin Count(TM)) of RPG to seven other defined areas of the H5N1 genome (Figure). The Replikin Count(TM) of RPG, was found to be associated with the pB1 area of the human H5N1 virus genome, and was increased tenfold from 2003 to 2006, during the current bird flu epidemics, when it was 4 to 10 times greater than that of the other genomic sections of the virus (hemagglutinin, neuraminidase, pA, pB2, ns, matrix, nucleocapsid)(p<0.001). In contrast to 'in vivo' or 'in vitro' localization of a gene, this method may be thought of as "in silico" identification or isolation of a gene.
References: 1. BogochS, BogochES. Replikins: The Chemistry of Rapid Replication. Begell Press, New York, 2005. 2. http://www.replikins.com
About Replikins LLC:
Replikins, based in Boston and founded in 2006, is a biotechnology company dedicated to unlocking the secrets of infectious disease virulence. The company's technologies, FluForecast(TM) software and core Replikins genetic elements, have led to a novel and significant forecasting capability, and the initial development of several vaccine candidates. The company is privately funded.
Contact:
Ann Borsanyi, CEO, Replikins Ltd. and LLC.
Email
617-536-0220
Scientific Background
In order to detect significant changes, the FluForecast(R) software calculates the mean, standard deviation and p values for each data point compared with all other data points for the strain or genes annually. The isolation of this new gene, by specific quantitative software methods, that is in silico, is analogous to the isolation of the peak concentration of a protein in vitro by column chromatography, mass spectroscopy, or radioisotope labelling. The in silico data is then correlated with in vitro and correlated with in vivo data, including a) epidemiological data: influenza epidemics and dormant periods were examined back to 1918 for the common strains (data kindly supplied by the Centers for Disease Control (CDC) Atlanta, GA). and b) laboratory experimental data: eg. an independent laboratory 'proof of concept' study showing the relationship of increase in Replikin Count(TM) to the increase in host mortality will be presented at a forthcoming scientific meeting (3).
A. Gene, Virus Strain, Specific Host, and Geographic Localization
Gene Localization
The Replikin Peak Gene(TM) in human and other host H5N1 was found to be concentrated in one segment of the genome, and to contain four to ten times the replikin concentration found in the other areas of the H5N1 genome (Figure 1). The mean Replikin Count(TM) (i.e the number of replikin peptide sequences per 100 amino acids of virus protein) of the Replikin Peak Gene(TM) in human H5N1 has risen from 2.0 (+/-0.1) in 2004 to 16.1 (+/-5.7) in 2006 (Figure 1). This increase correlates with the increased morbidity, the spread of H5N1 outbreaks to several dozen countries, and the increased mortality in fowl and man during the same period (WHO). In the 1997-98 Hong Kong H5N1 epidemic, approximately 8 of the approximately 30 human cases reported (26%) died; in 2006, the mortality rate was 69%) (WHO).
Virus Strain Localization and Replikin Peak Gene History
The Replikin Count of the Replikin Peak Gene of high mortality strains H1N1, H2N2, H3N2, H7N7 and H5N1, was compared to low mortality Influenza B, searched by FluForecast(R) software for each year between 1918 and 2006 for which sequence data for each strain was published on PubMed (Figure 2). Unlike the high mortality influenza A strains, influenza B does not have high mortality and as Figure 2 shows, it lacks any evidence of Replikin Peak Gene activity. H2N2 shows a declining Count in the Replikin Peak Gene in accord with its absence from virus isolates in recent years. H3N2 may be following a similar pattern of decline, as is H7N7. However, there appears to be a trend to an increasing Replikin Count in the Replikin Peak Gene in the H5N1 compared to H1N1 in 1918. In 1918, despite the huge morbidity, the human mortality rate was thought to have been less than 5%; at present, the human mortality rate of H5N1 is 69% (WHO).
Host Localization
Examination of whole H5N1 virus total replikins concentration in four different species, goose, duck, chicken and man, shows that in the last two epidemic years, 2005 and 2006 there has been a marked increase (p<0.001) in the Replikin Count (number of replikins per 100 amino acids) in three of the four species, and that the human H5N1 Count has now overtaken that of the chicken (Figure 3). When the Replikin Peak Gene is examined in the same four hosts, the dominance of the human H5N1 rise is even more clearly shown (Figure 4). Geographic Localization In addition to host localization in the human, geographic localization of the Replikin Peak Gene(TM) during 2006 indicated Indonesia and Thailand are leading countries for likely recurrence of human cases, a forecast now being confirmed by outbreaks in these countries in January of 2007 (Figure 5). B. Fine Structure and Conservation of the Replikin Peak Gene(TM) The Replikin Peak Gene(TM) also provides constituent specific replikin structures to target. We have learned that two replikins occur in H1N1(1918), H2N2(1957), and H3N2(1968) (the last three fatal pandemics) in H7N7 (human fatal Netherlands 2003) as well as in H5N1 (Indonesia, China 2006) , with only minor substitutions which do not involve the basic replikin structure nor the Replikin Count. This remarkable conservation of replikin structure across time, country, host, and virus strain makes it possible for specific pan-influenza synthetic replikin vaccines for which safety and efficacy trials are forthcoming.