Vaccine Genome Researchers Report 99.7% Colinear Identity Between a U.S. Civil War Era Smallpox Vaccine and Horsepox Virus Findings Published in the Current Issue of the Journal Genome Biology Horsepox is the Vector for

Tonix Pharmaceuticals Holding Corp. 8-K

Vaccine Genome Researchers Report 99.7% Colinear

U.S. Civil War Era Smallpox Vaccine and Horsepox Virus

Findings Published in the Current Issue

of the Journal Genome Biology

Horsepox is the Vector for Tonix's

Experimental TNX-1800 COVID-19 Vaccine

Horsepox Has Been Used as a Smallpox Vaccine

Since at Least the 1860's

Smallpox is the Only Viral Disease Ever

CHATHAM, NJ, December 4, 2020 - Tonix Pharmaceuticals Holding Corp.

(Nasdaq: TNXP) (Tonix or the Company), a clinical-stage biopharmaceutical company, announced today that independent researchers

reported 99.7% colinear identity between a circa 1860 U.S. smallpox vaccine and horsepox virus. Tonix's TNX-801 is a horsepox-based

live virus vaccine being developed as a potential vaccine to prevent smallpox and monkeypox. Tonix's TNX-801 is also the

vector on which Tonix's COVID-19 experimental vaccine is based. Tonix was not involved in the research reported.

The publication in the Journal Genome

Biology, entitled "Re-assembly of 19th-century smallpox vaccine genomes reveals the contemporaneous use of horsepox and

horsepox-related viruses in the United States"1 analyzed sequences of U.S. smallpox vaccines that had been published

recently by a different group also in Genome Biology, entitled "The origins and genomic diversity of American Civil

War Era smallpox vaccine strains"2. The first paper applied modern techniques of sequencing old DNA to smallpox

vaccine specimens stored in the M tter Museum in Philadelpia, and identified that vaccines with horsepox core viral sequences

were used in the 1860's and referenced sequence information from several vaccines that the authors posted in GenBank, the

National Institute of Health genetic sequence database. The new report re-assembled certain genomes from the GenBank sequences

associated with the first paper and found a remarkable degree of identity with the circa 1860 U.S. smallpox vaccine VK05 and the

1976 Mongolian horspox isolate called MNR-76. Tonix's TNX-801 was synthesized3 based on the sequence of the Mongolian

horsepox isolate MNR-764.

"The extent of colinear identity, approximately 99.7%,

is remarkable between the circa 1860 U.S. smallpox vaccine VK05and the 1976 Mongolian horsepox genome," said, Jos

Esparza, MD, PhD, Adjunct Professor at the Instiyute of Human Virology, School of Medicine of the University of Maryland, who was

an author of the Genome Biology letter. "This finding indicates that a true horsepox virus was used as a smallpox vaccine

in the U.S. in the 1860's. It is astonishing that there are so few differences between these viruses despite the separation

of their isolation by over 100 years and from different continents."

"Thes recent discovery is another step in establishing that

what is called 'horsepox today was used to vaccinate against smallpox in the 19th century." said

Seth Lederman, M.D., President and Chief Executive Officer of Tonix Pharmaceuticals. "Dr. Edward Jenner invented vaccination

in 1798 and the procedure was called vaccination because the inoculum material was initially obtained from lesions on the udders

of cows affected by a mild disease known as cowpox. Cow' is vacca' in Latin. However, Dr. Jenner suspected

that cowpox originated from horsepox.5 Subsequently, Dr. Jenner and others immunized against smallpox using material

directly obtained from horses. The use of vaccines from horses was sometimes called equination' from the Latin equus'

which means horse'. Equination and vaccination were practiced side-by-side in Europe6. Several years ago,

we recognized that the 2006 genome sequence of the 1976 Mongolian horsepox4 may be more like the vaccines that Jenner

used than modern vaccinia vaccines, because the modern vaccinia vaccines appear to have undergone both divergent and convergent

evolution since Jenner's time. To recreate a vaccine similar to Jenner's, together with scientists at the University

of Alberta, Canada, we synthesized3 a live horsepox virus based on the 1976 Mongolian horsepox virus sequence and we

are developing it as a potential vaccine for smallpox and monkeypox, called TNX-801. Horsepox is also the vector for our COVID-19

vaccine, TNX-1800, which has been modified to direct the expression of CoV-2 spike protein."

"The new findings are particularly significant, because they

confirm that horsepox has been used as a safe and effective smallpox vaccine since at least 1860, and probably since Edward Jenner's

time in 1798," continued Dr. Lederman. "The new report confirms that modern 'vaccinia'-based smallpox vaccines

from around the world have lost, apparently convergently, two genomic segments in evolution: an 11 Kilobase (kB) segment on the

left side and a 6 kB Segment on the right side relative to the progenitor horsepox-like vaccines. The presence of these genetic

elements in horsepox may contribute to the decreased virulence of horsepox relative to vaccinia in mice and the smaller plaque

size in tissue culture. Later, after approximately 1875, when vaccinia propagation was moved from arm to arm' to growth

on cows in vaccine farms'8, the core viral sequences of modern vaccinia vaccines appear to have diverged

away from the horsepox core viral sequences. A standard for modern vaccine development and manufacturing is to make a master

virus bank' and tie vaccine production to the exact properties of the original master virus bank. Edward Jenner did not keep

a master virus bank, because his work pre-dated understanding of viruses by more than 100 years. However, the recent work on sequencing

and analyzing archaic smallpox vaccines gives us a clearer understanding of the structure and features of Jenner's vaccines.

The more we learn, the more we are led to believe that Jenner's vaccinia was either horsepox or more like horsepox than modern

The recent report is the latest in a series from a team composed

by Dr. Esparza, Clarissa R Damaso, PhD at the Carlos Chagas Filho Biophisics Institute of the Federal University of Rio de Janeiro,

Brazil, and Andreas Nitsche, Ph.D. at the Robert Koch Institute in Berlin, Germany. They have previously reported7 that

the core viral sequence of a 1902 U.S. smallpox vaccine manufactured by Mulford, a vaccine manufacturer subsequently acquired by

Merck, was closely related to the Mongolian horsepox MNR-76 on which TNX-801 is based. The latest report shows that the circa 1860

VK05 vaccine also contains a high degree of colinear identity through the ends of the viruses, including regions where all previously

characterized smallpox vaccines including Mulford 1902 contained deletions of approximately 11 kB on the left side and 6 kB on

Dr. Lederman continued, "Smallpox was eradicated using single-dose

live replicating vaccines that induce durable T cell immunity, prevent serious illness after infection and block forward transmission.

Our hope and our goal is to produce vaccines that will provide long term immunity with a single dose using a proven technology

that can be readily scaled up for manufacturing and that does not require a costly and cumbersome cold chain for distribution and

TNX-801 is a live virus vaccine based on synthesized horsepox3,4.

Horsepox and vaccinia are closely related orthopoxviruses that are believed to share a common ancestor. Molecular analysis suggests

that TNX-801 has relatively "complete" left and right inverted terminal repeats (ITRs) while different vaccinia isolates

have a variety of deletions adjacent to the left and right ITRs including a characteristic approximately 11 kB deletion on the

left side and an approximately 6 kB deletion on the right side. Therefore, TNX-801 has additional genes, relative to vaccinia vaccines,

that may play roles in host immune interactions and one or more of such proteins may serve as antigens for protective immunity.

Molecular analysis also shows that horsepox is closer than modern vaccines in DNA sequence to the vaccine discovered and disseminated

by Dr. Edward Jenner3,4,57,8. The small plaque size in culture of TNX-801 appears identical to the U.S. Centers for

Disease Control publication of the natural isolate10. Relative to vaccinia, horsepox has substantially decreased virulence

in mice3. Tonix's TNX-801 vaccine candidate is administered percutaneously using a two-pronged, or "bifurcated"

needle. The major cutaneous reaction or "take" to vaccinia vaccine was described by Dr. Edward Jenner in 1796 and has

been used since then as a biomarker for protective immunity to smallpox, including in the World Health Organization's (WHO)

accelerated smallpox eradication program that successfully eradicated smallpox in the 1960's. The "take" is a

measure of functional T cell immunity validated by the eradication of smallpox, a respiratory-transmitted disease caused by variola.

Tonix's proprietary horsepox vector is believed to be more closely related to Jenner's vaccinia vaccine than modern

vaccinia vaccines, which appear to have evolved by deletions and mutations to a phenotype of larger plaque size in tissue culture

and greater virulence in mice. TNX-801 vaccinated macaques showed no overt clinical signs after monkeypox challenge11.

TNX-1800 is a live modified horsepox virus vaccine for percutaneous

administration that is designed to express the Spike protein of the SARS-CoV-2 virus and to elicit a predominant T cell response.

TNX-1800 is based on a horsepox vector, which is a live replicating, attenuated virus that elicts a strong immune response. Live

replicating orthopoxviruses, like vaccinia or horsepox, can be engineered to express foreign genes and have been explored as platforms

for vaccine development because they possess; (1) large packaging capacity for exogenous DNA inserts, (2) precise virus-specific

control of exogenous gene insert expression, (3) lack of persistence or genomic integration in the host, (4) strong immunogenicity

as a vaccine, (5) ability to rapidly generate vector/insert constructs, (6) readily manufacturable at scale, and (7) ability to

provide direct antigen presentation. Relative to vaccinia, horsepox has substantially decreased virulence in mice3.

Horsepox-based vaccines are designed to be single dose, vial-sparing vaccines, that can be manufactured using conventional cell

culture systems, with the potential for mass scale production and packaging in multi-dose vials. Like TNX-801, Tonix's TNX-1800

vaccine candidate is administered percutaneously using a two-pronged, or "bifurcated" needle. Tonix recently reported

that immunization with a single dose of TNX-1800 induced "takes" and neutralizing anti-SARS-CoV-2 antibodies in non-human

*TNX-801 is in the pre-IND stage and has not been approved for any