Ricin 1

Ricin:

Evaluation of Ricin Therapies

Katy Lunger

0902BSBD6409040

University of Maryland University College

4/12/2009
 Ricin 2

Table of Contents

Abstract……………………………………………………………………………………3
Introduction………………………………………………………………………………..4
Historical background……………………………………………………………………..4
Weaponization…………………………………………………………………………….5
Level of threat……………………………………………………………………..6
Future use in an attack…………………………………………………………….7
Biological properties of ricin……………………………………………………………...8
Mechanism of action………………………………………………………………8
Toxicity …………………………………………………………………………...9
Clinical presentations…………………………………………………………….10
Detection and diagnosis………………………………………………………….11
Prevention………………………………………………………………………………..11
Traditional and current preventions……………………………………………...12
Vaccines………………………………………………………………………….13
Treatment……………………………………………………………….………………..15
Traditional and current therapies………………………………………………...15
Post-exposure treatment………………………………………………………….16
Government/Medical response action plan………………………………………………17
Weaknesses of current prevention and treatment………………………………………..19
Conclusions and future approaches…….………………………………………………..20
References………………………………………………………………………………..23
Appendix…………………………………………………………………………………29
Table 1. Attacks and incidents involving ricin. …………………………………29
Figure 1. Structure of ricin toxin (3Dchem.com, 2003). ………………………..30
Figure 2. Mechanism of ricin (Audi et al., 2005). ………………………………30
Figure 3. Castor beans (Audi et al., 2005). ……………………………………...31
Figure 4. Countermeasure communication diagram (DHHS, 2006) …………...31
Glossary……………………………………………………………….…………………32
 Ricin 3

Abstract

With the growing awareness of the threat of biological and chemical weapons as

potentially devastating tools of terror, agencies within the United States government have

identified ricin toxin as one for which therapies are virtually non-existent. As a weapon

that is known to have been produced by enemies of the United States, as well as used on

at least two occasions, prevention and therapy of ricin poisoning must be addressed.

Although ricin itself is not considered a “weapon of mass destruction,” it can be readily

produced and obtained, and is therefore a viable biological threat. Current detection,

prevention, and therapeutic protocols for ricin exposure are rudimentary at best.

Antitoxin and vaccination research is showing some positive results with animal models,

however, there is not yet a Food and Drug Administration (FDA) approved treatment or

prophylactic for humans.

 Ricin 4

Introduction

Ricin is a highly hazardous toxin. It is a protein that is found in Ricinus

communis, also known as castor beans, and when extracted and isolated it can be used as

a deadly bioterror agent. Ricin is highly toxic, and when employed as a biological

weapon, it is difficult to diagnose; a lack of current treatments and preventions make ricin

a desirable biological weapon. One million tons of castor beans are processed each year

worldwide, and of the waste mash, 5% is ricin toxin by weight (Pratt et al., 2007). Ricin

toxin is classified as a Category B bioterrorism agent by the Centers for Disease Control

and Prevention (2003) because it is moderately easy to disseminate, would result in

moderate rates of morbidity and mortality, because the CDC is not yet sufficiently

equipped to detect ricin as a biological agent, and further enhancements to the CDC’s

diagnostic capabilities are required to handle a ricin attack (CDC, 2003).

Because of the potential risk of an attack with biological agents, including ricin,

countermeasure development has been taken very seriously by agencies within the United

States, such as the Department of Homeland Security and the Centers for Disease Control

and Prevention. This paper is concerned with the therapeutic value of current and/or

experimental treatments for ricin poisoning in the event of a bioterrorism attack.

Historical background

Ricinus communis was originally grown in Africa and Asia (Maman &

Yehezkelli, 2005). Since antiquity, the bean oil has been extracted and used for the

treatment of many diseases (Franz & Jaax, 1997), such as sores and abscesses in India,

Egypt, and China (Kirby, 2004). Today, castor oil is an important industrial feedstock for

numberous manufacturing processes and also is used as a lubricant and a laxative (Kirby,
 Ricin 5

2004). However, after the extraction of castor oil, the fibrous waste is left containing

toxic ricin protein. It is documented that ricin was developed as a weapon by the United

States, United Kingdom, and Canada in the time period between the two world wars

(Maman & Yehezkelli, 2005). And although the production of ricin as an offensive

weapon has been prohibited through the agreement reached in 1972 at the Biological and

Toxin Weapons convention, it is believed that as recent as 2003 both Paris and London

have developed and kept a stockpile of weaponized ricin (Maman & Yehezkelli, 2005).

One most commonly repeated stories regarding the use of ricin as a weapon is that

of the assassination of Georgi Markov in 1978. It is said that an assassin injected a

poisonous ricin protein pellet into his thigh using a weapon in the shape of an umbrella

(Maman & Yehezkelli, 2005). See table 1 for more information about historical ricin

attacks and incidents.

Weaponization

In the US between 1995 and 1997, several people were arrested for possession of

homemade ricin (Maman & Yehezkelli, 2005). In 1991, 4 members of an extremist

group in Minnesota were convicted of formulating a solution of ricin toxin and dimethyl

sulfoxide (DMSO) in an attempt to kill a US marshal (Franz & Jaax, 1997). In 2003 in

Britain, a domestic laboratory was found by police with the capability to produce

weaponized ricin (Mayor, 2003). Iraq is known to have produced ricin toxin and

stockpiled it from 1985 to 1991; 10 liters of concentrated ricin solution was allegedly

produced, with some of it loaded into artillery shells for field-testing (Maman &

Yehezkelli, 2005). All such incidents suggest that, with the right homemade equipment

and some fairly basic knowledge, a criminal or terrorist could easily obtain ricin toxin
 Ricin 6

and weaponize it.

Ken Alibek, a former top official involved in Russia’s biological weapons

program who defected to the United States in 1991, claims that Russia developed ricin

toxin as a weapon; he claims the ricin toxin used against Markov, as well as another

individual in Bulgaria, was concocted in Russian laboratories (Maman & Yehezkelli,

2005).

“Ricin can be prepared and remain stable in various forms: crystalline, liquid, and

even aerosol” (Maman & Yehezkelli, 2005). A study regarding weaponization of ricin in

the crystalline form (to place in a bomb) was done between the two world wars; the

results indicated that ricin is thermally sensitive (Kirby, 2004). This in turn means that

in the case of use in a bomb, or any form of delivery that would emit large amounts of

heat, much of the ricin toxin would become ineffective. If a liquid form of agent is used,

the liquid would dilute the ricin, requiring the production of larger amounts of the toxin

in order to create havoc. Aerosolized fine powder formed of particles smaller than 10um

in size was determined as the most effective form for use in an attack, since the smaller

the particle, the deeper it can be deposited in the respiratory system causing disease

(Audi, Belson, Patel, & Schier, 2005). A limiting factor for aerosolization is that the

toxin’s intensity fades as the aerosol cloud disperses.

Level of threat

The toxin produced by the ricin protein has been found to cause the “agglutination

of erythrocytes and precipitation of serum proteins” (Olsnes & Pihl, 1976). Without

treatment, this would lead to cell death followed by organ failure, and eventually death of

the individual (Shea & Gottron, 2004). According to data compiled by Maman and
 Ricin 7

Yehezkelli (2005), the routes of exposure found to be most lethal in humans are the

inhalational and intravenous routes, at a median lethal dosage (LD50) of 3ug/kg in

humans. Ingestion of ricin toxin is reported as second in threat level of toxicity with an

LD50 of 30ug/kg. Subcutaneous exposure has an LD50 of 500ug/kg according to the case

of Markov’s assassination (Maman & Yehezkelli, 2005). Person-to-person transfer of

toxin does not take place (Audi et al., 2005). Regardless of route, death within 36-48

hours has been observed after exposure to ricin, while more often a full recovery is seen

in patients (Maman & Yehezkelli, 2005).

According to the CDC (2003), this agent is a Category B bioweapon agent,

because it is a moderate risk having moderate morbidity rates, and is not necessarily a

good candidate for use as a weapon of mass destruction. However, the ease of finding

castor beans and extracting the ricin toxin makes it a credible choice as a biological

weapon. Also, a direct treatment of the agent is not available, only supportive treatment

can be administered, and therefore if not diagnosed properly the victim will be at an even

higher risk of developing deleterious effects (Maman & Yehezkelli, 2005).

Future use in an attack

It is believed that up to as recent as 2003, private laboratories within Paris and

London have dedicated time and money for the weaponization of ricin (Maman &

Yehezkelli, 2005). In 2004, US Senator Frist was targeted by mail contaminated with the

ricin toxin (Shea & Gottron, 2004). This indicates that the possibility of an attack using

ricin continues to exist. But as many experts and scientists have concluded thus far, the

successful use of ricin as a weapon of mass destruction is less likely. Ricin would most

likely be used in an attack targeting specific persons, as has been mainly done in past
 Ricin 8

attacks, and thus continues to be considered a biological weapon used mainly for the

purpose of causing terror more so than death (Shea & Gottron, 2004).

Biological properties of ricin

Mechanism of action

Research has been conducted on ricin not only for weaponization purposes, but

also for defensive, therapeutic purposes in the event of an attack with ricin. The toxic

ricin protein is stable in both powder and liquid forms (Audi et al., 2005). Ricin protein

is extracted from castor beans and purified as a white powder that is highly toxic to

humans. The purified ricin protein is soluble in water and stable throughout a wide pH

range. Ricin can be denatured, and therefore inactivated, at 80 ºC for 1 hour in aqueous

solution (Audi et al., 2005). The toxic protein’s stability as both a liquid and powder

allow it to be disseminated in air, food, or via the public water supply. Therefore, victims

of a ricin attack could inhale or ingest the toxin.

Ricin protein, with a molecular weight of 60-65 kDa, consists of 2 chain proteins,

A and B. The heterodimeric structure is 2.5Å across, and these attributes are very

important for dissemination as a toxin (see figure 1) (Audi et al., 2005). The A chain

protein is connected to the B chain protein by disulfide bridges, and each protein chain

has biological function (Audi et al., 2005). The B chain binds lectin to glycoprotein in the

cell membrane, allowing endocytosis of the ricin protein into the cytosol of the cell; the A

chain has RNA N-glycosidase activity, which inhibits protein synthesis of 28 S rRNA of

the 60S ribosomal subunit (see figure 1) (Audi et al., 2005; Maman & Yehezkelli, 2005).

Once the ricin toxin enters the endosome, it is transported through the golgi apparatus to

the endoplasmic reticulm (ER) and the A chain binds to the 28S rRNA ribosome and
 Ricin 9

inactivates protein synthesis which causes apoptosis in cell (Audi et al., 2005). The ricin

toxin can inactivate ribosomes at a rate of 1500-2000 ribosomes per minute. Ricin toxin

is a type 2 ribosome inactivating protein (RIP-II), like botullinum, and anthrax, that has

the capability of being a very effective toxin; once it enters, the human body is extremely

difficult to detoxify (Audi et al., 2005).

Toxicity

Information regarding the toxicity of ricin is important for determining hazardous

dosage levels in the event o a public emergency. The degree of toxicity depends on the

route of administration. According to Maman and Yehezkelli (2005), toxicity levels of

ricin poisoning vary by route of exposure, as do prognosis and mortality. The mean

lethal dosages (LD50) differ by the administration route as well, such as ingestion,

inhalation, intravenous and subcutaneous injections (Audi et al., 2005). The LD50 for

ingestion, in the case of mice, is 20 mg/kg for 85 hours and 30 ug/kg for 6-8 days for

humans (Audi et al., 2005). However, intake of ricin through the respiratory tract in mice

has a much lower LD50 at 3-5 ug/kg for 36-72 hours, and 3 ug/kg for 36-72 hours in

humans (Audi et al., 2005).

Intravenous administrations of ricin toxin match the median lethal dosages for

inhalation of toxin. Moreover, subcutaneous injection of ricin in mice has a median

lethal dosage of 24 ug/kg for 100 hours, but for humans it is 500 ug/kg for 3- 6 days

(Audi et al., 2005). Inhalation and intravenous injection of ricin can result in life-

threatening prognoses with very small dosages of ricin (Maman & Yehezkelli, 2005).

Griffiths, Phillips and Holley (2007) also found similar results in a study with Green

monkeys, corroborating other reports that symptoms and prognosis will depend on the
 Ricin 10

route of administration.

Clinical presentations

The study of the biological properties, the mechanism of action, and degree of

toxicity, of ricin has clearly shown that different types of symptoms occur via different

administrating routes. The first route is the gastrointestinal tract in animals and humans.

Ingestion of ricin may cause nonspecific symptoms, including abdominal pain, vomiting,

diarrhea, fever and dehydration, which can result in electrolyte imbalances, hypotension,

leukocytosis, elevated transaminase levels, hyperbilirubinemia, and anemia. Moreover,

necrosis of the intestinal tract due to hemorrhagic lesions may occur as well (Audi et al.,

2005).

The second route of ricin poisoning is inhalation via the respiratory tract. The

symptoms are respiratory distress, nausea, fever, cough, edema, hypotension, and

respiratory failure (Audi et al., 2005). Particle size can make a difference in where

particles accumulate in the respiratory tract. For example, smaller particles may

accumulate deeper in the lung, in the lower respiratory tract, resulting in higher mortality

rates than larger particles (Audi et al., 2005).

Intramuscular injection is the third method of ricin poisoning. Symptoms are flu-

like, with nausea, local muscle pain, and swelling. However, high doses result in the

spread of the toxin to the lymph nodes, where necrosis can occur; further, ricin toxin can

spread to other organs through the blood stream where apoptosis can cause the organs to

die (Audi et al., 2005).

Detection and diagnosis

Because human bodies differ somewhat, for example weight and predisposition to
 Ricin 11

respiratory tract dysfunction, clinical presentation may vary, especially if different

dosages and administration routes occur in an attack (though this is unlikely). Because

many of the symptoms of ricin poisoning mimic other diseases, physicians may have

difficulty in identifying ricin as the cause (CDC, 2003).

Clinical detection and diagnosis of ricin poisoning is critical in the event of an

attack. After exposure, clinical diagnostic tests and other types of laboratory tests must

be able to detect small amounts of ricin in the human body. Immunologically based

methods, such as Enzyme-Linked ImmunoSorbent Assay (ELISA), may be able to

measure very low concentrations of ricin in animals, but no clinically validated methods

for humans are currently available (Audi et al., 2005; Franz, 1997).

According to Audi and colleagues (2005), the US Army Medical Research

Institute of Infectious Disease (USAMRIID) and the CDC are working on tailoring

animal-based diagnostics to humans. Matrix-assisted laser desorption-ionization mass

spectrometry (MALDI-MS) may be a useful method for ricin detection in urine and

serum samples. Interestingly, Kalb and Barr (2009) at the CDC were successful in

detecting ricin in clinical specimens and food using analytical methods, such as MALDI-

MS for analyzing a DNA substrate, and liquid chromatography tandem mass

spectrometry (LC-MS/M) for analyzing the ricin protein; however, there is no ricin-

specific commercial assay available (Audi et al., 2005).

Prevention

Ricin toxin poisoning is highly lethal and can kill a victim within a short period of

time. Diagnosis of ricin poisoning is difficult, and the progress of the toxin within the

human body will render delayed detection and treatment futile. Obviously, prevention of
 Ricin 12

infection is ideal, however, currently there are no specific medical prevention or

treatment plans available to the public for natural infection, let alone for a ricin attack.

The need for treatment and prevention plans has resulted in research for ricin

poisoning therapy and prevention. In addition, development of a ricin vaccine would

benefit not only patients, but government as well. A vaccine has the capability of long-

and short-term prevention from infectious and toxic agents using natural human

immunogenic molecules to prevent respiratory tract injury and high mortality rates, and

reduce extraordinary treatments such intensive care and long-term care (DHHS, 2006).

Governments, individual citizens, and physician will benefit from preventing high health

care costs as well as economical damage from these agents.

Traditional and current preventions

Ricin vaccine as a medical prevention is not available. As a result of that, the US

Army has established that the best physical protection from toxin aerosols is a mask and

cloth. The mask is the best practical solution for active duty military personnel as it can

be donned when appropriate (Audi, et al., 2005; Maman & Yehezkelli, 2005). However,

public citizens could not predict such a situation, even if the government has indicated a

high threat level. In this situation, early detection of the toxin is the best potential

prevention. For example, according to Maman and Yehezkelli (2005), US government

agencies, including the US Postal Service, have utilized chemical and biological sensors

for early detection. Theoretically, as soon as a biosensor detects a chemical agent, CDC

should confirm it. Presently, the US biosensor program, BioWatch, is under scrutiny

regarding the efficiency with which it can detect chemical or biological agents (Shea &

Lister, 2003).
 Ricin 13

Vaccines

Although a vaccine as a ricin countermeasure is not yet available, vaccine

research and development have been reported by the Griffiths group (2007) in the United

Kingdom, Smallshaw, Richardson, and Vitetta (2007), and the Carra group (2007) in the

United States; each are using different and progressive technical approaches. Griffiths’

group has used a traditional vaccine production method. Pentacel is a toxoid vaccine,

that is, it contains bacterial components that have been altered so that it does not induce

the same level of toxicity as would naturally occurring toxic bacteria, yet it is still able to

invoke antibody production in the body (FDA, n.d.). For the Pentacel vaccine against

diphtheria and tetanus, proteins with toxic attributes are deactivated by soaking in 10%

formaldehyde for 28 days at 37 ºC. This procedure is very common and safe for

production of vaccines, and has been approved by the FDA.

Griffiths and colleagues (2007) have done several studies on ricin toxoids. In

1995, they used the toxoid method to develop an experimental ricin vaccine; they showed

that it acted as a prophylactic and protected rats when injected with ricin toxin, but tissue

histopathology was abnormal (Griffiths et al., 1995). They also tried to improve efficacy

with an adjuvant and a different inoculation site. In 1997, they published a study in

which they used ricin toxoid, with aluminum hydroxide as an adjuvant, within liposomes;

they found the liposome-encapsulated toxoids were effective against lethal ricin attacks,

but increasing vaccine dosage intratracheally resulted in abnormal histopathology in lung

tissue (Griffiths et al., 1997). The research data implied that the vaccine efficacy was

affected by the inoculation process and route; this data is very important regarding

prevention from aerosolized ricin; local pulmonary protection may be necessary for
 Ricin 14

establishing a vaccine (Griffiths et al, 2007).

Kende et al. (2007) demonstrated an intranasal vaccine, deglycosylated ricin A

chain (rRV), when used by itself induced a very poor antibody response. Thus, they used

mucosal adjuvant LTK63 or LTR72 with rRV. The LTK63 or LTR72 refers Escherichia

coli LT enterotoxin that has two mutations on the toxin using as the vaccine adjuvant.

The adjuvant improved the efficacy, with survival rate and safety significantly improved

when compared with the rRV itself; however, lung injury was not reduced.

Another ricin prophylactic has been explored by Carra and colleagues (2007).

They developed a recombinant ricin A-chain vaccine (RTA1-33/44-198) in which the

toxic portions of the ricin A-chain sequence were deleted (Carra et al, 2007). The group

found that B-cell epitopes bind more tightly to ricin protein when the protein is adhered

to aluminum hydroxide. Not only that, but the aluminum hydroxide helps to stabilize the

protein conformation in the recombinant vaccine so that freezing and thawing multiple

times will not alter the efficacy of the vaccine in mice (Carra et al., 2007). However, it

needs to be confirmed that it is safe and does not cause tissue damage.

The Smallshaw group’s vaccine is also a recombinant ricin A-chain vaccine

(RiVax) (Smallshaw et al., 2007). In RiVax the toxicity sites in the A and B chains have

been deleted, thereby maintaining the molecule’s antigenicity but removing its toxicity

(Smallshaw et al., 2007). Further, this vaccine has been used in mice without aluminum

hydroxide adjuvant and was shown to protect against ricin exposure via gavage or

inhalation of aerosol (Smallshaw et al., 2007). However, tissue damage was found in the

lungs of the mice used in their experiments (Smallshaw et al., 2007). From the results of

the data, optimal dosing needs to be determined for the vaccine; efficacy over time also
 Ricin 15

needs to be determined, as a point mutation could lead to protein aggregation over time

(Carra, et al, 2007).

According to a DOR BioPharma, Inc., (2008) news report, the company

announced they had achieved vaccine stability for up to 2 years in storage; the

Biomedical Advanced Research and Development Authority (BARDA), requires proven

stability of vaccines in order to be considered for the Strategic National Stockpile. DOR

BioPharma, Inc. (2008) also reported that RiVax has completed Phase 1 clinical trials

with good results. Vaccine stability will improve efficacy and allow the optimal dose to

be determined. Although there are examples of vaccines in different stages of

development, RiVax is a vaccine that is a potential candidate for ricin vaccine.

Treatment

Traditional and current therapies

While FDA approved antitoxin therapies exist for some other toxins, such as

horse IgG for botulinum toxin, currently there are no ricin antitoxin therapies available,

and clinicians can only offer supportive care for ricin poisoning symptoms (Audi et al.,

2005; Maman & Yehezkelli, 2005). Because of a lack of treatment options, in the event

of a mass-casualty scenario, most people would die if they were exposed to an adequate

dosage of ricin. Moreover, the patient may require long-term treatments, leading to high

medical costs for patients (DHHS, 2006). According to Pratt and colleagues (2007), the

only countermeasure that could provide immediate immunity against ricin exposure,

including post-exposure protection, is passive antibody administration. Pratt and

colleagues (2007) developed an oropharyngeal aspiration model for ricin-induced lung

toxicity and showed that passive vaccination with certain antibodies could offer full
 Ricin 16

protection up to 18 hours after ricin exposure in animal models. According to Griffiths

and colleagues (2007), the passive immunity approach has high potential for both pre-

and post- ricin exposure protection.

Post-exposure treatment

There is much research and development for post-exposure treatment which has

been published. Without a vaccine, the therapeutic post-treatment for lethal inhalation is

very critical as a function of time, becoming more critical as time passes after exposure

(Griffiths, et al, 2007). The level of dosage plays a role in the necessity of treatment as

well, because a high dose is more likely to cause death and/or progressive lung damage.

According to Pratt and colleagues (2007), passive immunity experiments in animals have

been done against ricin since 1972. The only countermeasure that could provide

immediate immunity against ricin exposure, including post-exposure protection, is

passive antibody administration.

In 1995, the Wellner group demonstrated that passive anti-Ricin IgG polyclonal

antibody introduced 1 hour after ricin exposure in mice resulted in lung injury; with 2 of

6 mice dead and the other four never fully recovering. Pratt and colleagues (2007)

developed an oropharyngeal aspiration model for ricin-induced lung toxicity and showed

that passive vaccination with certain antibodies could offer full protection up to 18 hours

after ricin exposure in animal models, but the survival rate at 24 hours was only 30%. In

addition, the polyclonal antibody doesn’t have specificity and so efficacy is low. In

contrast, monoclonal antibodies have specificity; therefore, the window period will be

long, however, the production is very low. Griffiths and colleagues (2007) found that

anti-A-chain and B-chain IgG allowed mice to extend time to death. Moreover, Pratt and
 Ricin 17

colleagues (2007) pointed out the apoptosis pathway involves p38 Mark activation in the

cascade of the cell, which is very important.

Another example of a new antitoxin therapy was reported by Fan, et al., (2009).

They created a 31RNA ligand (31RA aptamer) that binds on ricin A-chain (RTA) and

inhibits ricin toxicity as assayed with cell based luiciferase translation in Chinese hamster

ovary (CHO) cells. In the experiment, the 31RA aptamer blocked the RTA and reduced

ricin toxicity based on the cell. However, the activity of the 31-mer RNA ligand in mouse

experiments for different post-exposure times is also very interesting.

Passive antibody therapy could be used to treat exposed patients within 18 hours.

The other new method is too new to evaluate whether it is efficacious. However, ricin

intoxication is necessary to investigate in research and development because current

treatments only extend life and do not cure ricin poisoning. Intoxication with ricin

therefore necessitates long-term medication and suffering.

Government/medical response action plan

Since “white powder” cases were reported at Dirksen Senate Office Building in

Washington, D.C. in 2004, the Office of Public Health Emergency Preparedness

(OPHEP) in the Department of Health and Human Services (DHHS) was established

which developed ricin response plans (DHHS, 2006). OPHEP delineated an emergency

response to ricin attack at the federal, state, and local levels. It includes medical response,

treatment and lab testing, diagnostics, health care worker safety and preparedness,

hospital preparedness, surveillance, epidemiology, and environmental assessment and

cleanup (DHHS, 2006). The FBI, DHHS, DHS, CDC, USDA, and OSHA were involved

in the emergency planning after the Dirksen ricin incident. DHHS and CDC prepared
 Ricin 18

medical response plans, which included major ricin medical information in order to

establish awareness of ricin in our public.

The major action plan from the CDC (2008): 1) established risk analysis of

biological and chemical threats against the public, and 2) established emergency

preparedness and response plans for emerging heath threats so as to prevent public health

disasters at the national, state and local levels; and 3) established surveillance systems

domestically and internationally for monitoring infectious or noninfectious diseases

which include biological and chemical weapons. The Environmental Protection Agency

(EPA) has responsibility for environmental issues, including decontamination and

cleanup of agents (CDC, 2008). OSHA is involved with workplace health, including

laboratory safety for lab workers. USDA is responsible for agriculture and animal health.

Department of Homeland Security (2006) is responsible for the management of

the emergency response which involves public health, state, and FBI responses. The

comprehensive communication diagram is shown in figure 4. The collaboration among

the federal and state agencies will enable quick detection and efficient response to disease

outbreaks and implementation of measures to minimize the health, social, and economic

consequences of such outbreaks (DHHS, 2006). Because of several ricin incidents, the

US government determined that preparedness and response plans for ricin terrorism must

be enacted in order to efficiently minimize economic damage and public health disasters.

Weaknesses of current prevention and treatment

Current ricin exposure prevention methods include wearing gas masks and the

BioWatch program. While gas masks hold utility against an attack with ricin aerosol, it
 Ricin 19

is unlikely that all American civilians will have access to these types of protective

equipment when an attack occurs. Government programs are not in place to train the

public on the types of filters necessary against biological agents, where to purchase

masks, how to wear them, or how to maintain them.

The BioWatch program began in 2003, and was set up to provide early detection

and early warning of chemical and biological agents in major US cities (Shea & Lister,

2003). Although the locations of coverage are classified, it is known that sensor stations

are set up in conjunction with Environmental Protection Agency (EPA) air-quality sensor

sites. The BioWatch program is expensive, and many have criticized its efficacy and

efficiency. For each city the program is set up in, the initial cost is $1 million, and then

$1 million per year thereafter (Shea & Lister, 2003). The “sensors” are literally filters

that catch debris in the air; they are collected every 24 hours by technicians who extract

DNA and run PCR tests for known pathogens (Shea & Lister, 2003).

The labor-intensive aspects of the BioWatch program have come into question,

because many argue that the sensor sites have not been adequately studied in relation

with air-flow dynamics or even the likelihood of attack (Shea & Lister, 2003). Further

complications arise from state and federal government responsibilities and how to share

them as pertains to BioWatch; even the legitimacy of actual detection has raised

eyebrows. For example, natural background levels of a toxin could set off a very

expensive false alarm if government agencies responded (Shea & Lister, 2003).

Research for vaccines and antitoxin therapies is showing promise in animal

models, however, as Griffiths and others (2007) have pointed out, differences in the

fundamental morphology in animal models could complicate research efforts. For

 Ricin 20

example, mice are obligate nasal breathers, while primates can breath through the nose

and mouth; these differences could result in different patterns of particle deposition, and

hence differences in pathogenesis of disease (Griffiths et al., 2007). Also, vaccines that

are showing promise, such as RiVax, still show residual toxicity in animal tissue

(Smallshaw et al., 2007).

For cases where ricin toxin is ingested, clinicians are trained to prevent absorption

of ricin toxin in the body by gastric emptying and rinsing, or by absorbing the toxin with

activated charcoal (Maman & Yehezkelli, 2005). These methods have not been clinically

verified or standardized, and are basically rudimentary treatment methods. The

likelihood of an attack of the food or water supply is lower than an aerosol attack because

much more toxin is needed to cause death via ingestion; however, this scenario should

not be overlooked because ricin toxin is readily available and large quantities could be

concentrated and used to infect a population in this way.

Conclusions and future approaches

As has been discussed, treatment and prevention of ricin poisoning is not

currently adequate for protection from a terror attack, or even for accidental ingestion of

ricin toxin. Governmental agencies have focused resources toward research for antidotes,

treatments, and prophylaxis for ricin poisoning, however, there is still a long way to go in

terms of offering viable, standardized protection. The United States government has only

recently become aware of the credibility of a biological weapons attack, and since then,

appropriate countermeasures have begun to take shape, including the initiation of the

BioWatch program, and the founding of the Department of Homeland Security and

Biomedical Advanced Research Development Authority.

 Ricin 21

Most research that has been done thus far has been done on rodent models, and

because of differences in morphology, research needs to be focused on primate models.

Research needs to be adequately funded, even though ricin is considered a Category B

agent and therefore not as high a priority as Category A agents, because ricin toxin can be

acquired quite easily. Large amounts of ricin toxin could easily be disseminated in the

water or food supplies, and more research should be focused on the ingestion of the toxin.

There is not much solid evidence in the open literature regarding the history of

weaponization of ricin toxin. While intelligence data may exist as classified material,

there should be more education and awareness among the public regarding the likelihood

of a ricin attack. Programs need to be enacted so that clinicians are equipped with up-to-

date information regarding treatments and protocols for reporting cases of ricin

poisoning. Communication between state and federal government agencies regarding

biological attacks must be streamlined with standard operating procedures. The swift

response to a biological attack by local, state, and federal governments will only be

successful if the entities work in concert.

Experimental treatments should be considered for production and stockpiling as

emergency-only treatments for ricin attacks. While some deleterious effects have been

shown in some of the more promising experimental vaccines and antitoxins, analysis

should be done to weigh the advantages and disadvantages of side effects of the

experimental treatment with results of ricin intoxication.

While the United States may not currently be prepared for a ricin attack, current

programs and research are ongoing to ensure that programs of the future will be adequate.
 Ricin 22

References
3Dchem.com. (2003). Castor beans (Ricinus communis). Retrieved April 11, 2009,

from http://www.3dchem.com/moremolecules.asp?ID=13&othername=Castor

%20beans%20(Ricinus%20communis)
 Ricin 23

Audi J., Belson, M., and Patel, M. Schier J., and Osterloh J. (2005). Ricin Poisoning: A

comprehensive review. Journal of the American Medical Association, 294(18),

2342-2351. Retrieved February 25, 2009, form http://jama.ama-

assn.org/cgi/content/full/294/18/2342

Bigalke, H., & Rummel, A. (2005). Medical aspects of toxin weapons. Toxicology, 214,

210- 220. Retrieved March 10, 2009, from

http://tychousa3.umuc.edu/BSBD640/0902/9040/class.nsf/7e8f43891d9b6492852

57543000d4802/8525751d00757bc8852571bf006ff96a/$FILE/medical

%20aspects%20of%20toxin%20weapons%20including%20mycotoxins.pdf

Carra, J. H., Wannemacher, R. W., Tammariello, R. F., Lindsey, C. Y., Dinterman,

R. E., Schokman, R. D., and Smith L. A. (2007). Improved formulation of a

recombinant ricin A-chain vaccine increases its stability and effective antigenictiy

Vaccine, 25, 4149-4158. Retrieved February 28, 2009, from

http://www.sciencedirect.com.ezproxy.umuc.edu/science?

_ob=MImg&_imagekey=B6TD4-4NBRRJX-2-

1&_cdi=5188&_user=961261&_orig=search&_coverDate=05%2F22%2F2007&

_sk=999749978&view=c&wchp=dGLzVtz-

zSkWA&md5=130667f602cea6fa9895dd199a9e91f3&ie=/sdarticle.pdf

Centers for Disease Control and Prevention (CDC). (2003). Recognition, management

and surveillance of ricin-associated illness. Retrieved March 17, 2009, from

http://www2.cdc.gov/phtn/webcast/ricin/RicinScript.rev.07-14-04.htm

Centers for Disease Control and Prevention (CDC). (2008). Epidemiological overview

for clinicians. Retrieved April 10, 2009, from

 Ricin 24

http://www.bt.cdc.gov/agent/ricin/clinicians/epidemiology.asp

DOR BioPharm, Inc. (2008, January). DOR BioPharma reports achievement of two-year

stability milestone for RiVaxTM, its vaccine against ricin toxin. News Center.

Retrieved March 18, 2009, from

http://www.dorbiopharma.com/press/2008/2008JAN29.htm

DHHS. (2006). Guidelines for federal, state, and local public health and medical

officials. Retrieved April 10, 2009, from

http://emergency.cdc.gov/agent/ricin/pdf/ricin_protocol.pdf

Fan, S., Wu, F., Martiniuk, F., Hale, M.L., Ellington, A.D., and Tchou-Wong, K-M.

(2008, November).Protective effects of anti-ricin A-chain RNA aptamer against

ricin toxicity. World J Gastroenterol, 14(41), 6360-6365. Retrieved March 5,

2009, from http://www.wjgnet.com/1007-9327/14/6360.pdf

Food and Drug Administration (FDA) (n.d.). Pentacel: Diphtheri and Tentanus Toxoids

and Acellular pertussis adsorbed , inactivated poliovirus and hemophilus be

conjugate (Tetanus toxoid conjugate) vaccine in Sanofi Pasterur Limitted package

insert, (p 1- 34) http://www.fda.gov/cber/label/pentacellb.pdf.

Franz, D. R. (1997). Defense against toxin weapons. (pp. 1-60), U.S. Army Medical

Research Institute of Infectious Diseases, Fort Detrick, MD. Retrieved March 13,

2009, from http://www.usamriid.army.mil/education/defensetox/toxdefbook.pdf.

Franz, D. & Jaax, N. (1997). Ricin toxin. Textbook of military medicine: Medical

aspects of chemical and biological warfare. TMM publications

Griffiths, G. D., Bailey, S. C., Hambrook, J. L., Keyte, M., Jayasekera, P., Miles, J., &

Williamson, E. (1997). Liposomally-encapsulated ricin toxoid vaccine delivered

 Ricin 25

intratracheally elicits a good immune response and protects against a lethal

pulmonary dose of ricin toxin. Vaccine, 15(17-18), 1933-1939.

Griffiths, G. D., Lindsay, C. D., Allenby, A. C., Bailey, S. C., Scawin, J. W., Rice, P., &

Upshall, D. G. (1995). Protection against inhalation toxicity of ricin and abrin by

immunization. Human Experimental Toxicology, 14(2), 155-164.

Griffiths, G. D., Phillips, G. J., & Holley, J. (2007) Inhalation toxicology of ricin

preparations: Animal Models, prophylactic and therapeutic approaches to

protection. Inhalation Toxicology, 19, 873-887. Retrieved March 5, 2009, from

http://www.informaworld.com/smpp/title~content=t71365771

Hampton, T. (2004). Ricin vaccine devloped. Journal of the American Medical

Association, 292(12), 1419. Retrieved February 25, 2009, from http://jama.ama-

assn.org/cgi/content/full/292/12/1419-a

Hicks, P. P., Hartell, M.G., Nichols, D. A., Bhattacharjee, A.K., van Hamont J.E., and

Skillman, D.R., (2005). The medicinal chemistry of botulinum, ricin, and anthrax

toxins. Current Medicinal Chemistry, 12, 667-690. Retrieved March 10, 2009,

from

http://tychousa3.umuc.edu/BSBD640/0902/9040/class.nsf/7e8f43891d9b6492852

57543000d4802/8525751d00757bc8852571bf006ff96a/$FILE/med%20chem

%20of%20bot%20ricin%20and%20anthrax%20toxins.pdf

Kalb, S.R., and Barr, J.R., (2009). Mass spectrometric detection of ricin and its activity in

food and clinical samples. Analytical Chemistry, 81,(6), 2037-2042. Retrieved

February 25, 2009, from http://pubs.acs.org/doi/abs/10.1021/ac802769s.

Kende, M., Tan, X., Wlaxlowski, C., Williams, R., Lindsy, C., Giudice, D. G. (2007).
 Ricin 26

Enhancement of intranasal vaccination with recombinant chain A ricin vaccine

(rRV) in mice by the mucosal adjuvants LTK63 and LTR72. Vaccine, 25, 3219-

3227. Retrieved February 28, 2009, from

http://www.sciencedirect.com.ezproxy.umuc.edu/science?

_ob=MImg&_imagekey=B6TD4-4N08HD7-1-

K&_cdi=5188&_user=961261&_orig=search&_coverDate=04%2F20%2F2007&

_sk=999749983&view=c&wchp=dGLzVtz-

zSkWA&md5=96d17f6f0de78313abb740144c400dc5&ie=/sdarticle.pdf

Kirby, R., 2004. Ricin toxin: a military history. pp. 1-3. Retrieved March 19, 2009 from

http://www.wood.army.mil/chmdsd/pdfs/2004%20Apr/Kirby-Ricin-04-1.pdf

Maman, M.,and Yehezkelli, Y. (2005). Ricin: A possible, noninfectious biological

weapon in I.W. Fong and K. Alibek (Eds.), Bioterrorism and infectious agents: A

new dilemma for the 21st Century (pp. 205-216). New York: Springer.

Mayor, S., (2003). UK doctors warned after a ricin poison was found in police raid.

British Medical Journal, 326(126).

Olsnes, S. & Pihl, A., (1976). Abrin, ricin and their associated agglutinins. Chapman

and hall, London, pp. 129-173.

Pratt, T. S., Pincus, S. H., Hale, M.L., Moreira, A.L., Roy, C. J., Tchou-Wong, K. M.

(2007). Oropharyngeal aspiration of ricin as a lung challenge model for

evaluation of the therapeutic index of antibodies against ricin A-chain for post-

exposure treatment. Experimental Lung Research, 33, 459-481. Retrieved March

3, 2009, from http://www.informaworld.com/smpp/title~content=t713722902.

Reuters. (2007, August). DOR Biopharmacy announces initiation of second human

 Ricin 27

clinical trial for RiVax™, its ricin toxin vaccine. Retrieved March 17, 2009, from

http://www.reuters.com/article/pressRelease/idUS131156+07-Aug-

2008+MW20080807

Shea, D. & Gottron, F. (2004). Ricin: technical background and potential role in

terrorism. (Order Code RS21383). Washington, DC: CRS Report for Congress.

Shea, D. A., & Lister, S. A. (2003). The BioWatch program: Detection of bioterrorism.

(Order Code RL32152). Washington, DC: CRS Report for Congress.

Smallshaw, J. E., James A. Richardson, J.A., & Vitetta, E. S. (2007). RiVax, a

recombinant ricin subunit vaccine, protects mice against ricin delivered by gavage

or aerosol. Vaccine, 25(42), 7459 - 7469. Retrieved February 28, 2009, from

http://www.sciencedirect.com.ezproxy.umuc.edu/science?

_ob=MImg&_imagekey=B6TD4-4PJ69B7-1-

1&_cdi=5188&_user=961261&_orig=search&_coverDate=10%2F16%2F2007&

_sk=999749957&view=c&wchp=dGLzVtz-

zSkWA&md5=33ea71ca19fe0bf05e55d8f76e2221d0&ie=/sdarticle.pdf

U.S. Department of Health Human Service (DHHS). (2006, June). Response to a ricin

incident: Guidelines for federal, state and local public health and medical officials

Retrieved April 6, 2009, from

http://www.bt.cdc.gov/agent/ricin/pdf/ricin_protocol.pdf.

U. S. Department of Homeland Security (DHS), (2004) Biological Incident Annex in

National Response Plan, Retrieved April 6, 2009, from

http://www.iir.com/global/FusionCenter/NRPbaseplan.pdf.

Wellner, R. B., Heweston, J. F., and Poli, M. A. (1995). Ricin: Mechanism of action,
 Ricin 28

detection and intoxication. Journal of Toxicology-Toxin Review, 14, 483-522.

Woods, J B. (2005, April).USAMRID’s Medical management of biological casualties

handbook (6th ed.) U. S. Army Medical Research Institute of Infectious Disease

(USAMRIID). (pp. 93-96). Frederick, MD. Retrieved March 18, 2009, from

http://www.usamriid.army.mil/education/bluebookpdf/USAMRIID%20BlueBook

%206th%20Edition%20-%20Sep%202006.pdf.

Appendix

Table 1. Attacks and incidents involving ricin.

Year Incident
1978 Attempt to assassinate Bulgarian exile, Vladimir Kostov.
1978 Assassination of Bulgarian radical Georgi Markov in
London.
1991 Minnesota patriots council arrested for possession of 0.7
grams of ricin.

1995 130 grams of powdered ricin were detained from Thomas

Lavy while crossing into Canada from Alaska.
 Ricin 29

1997 Home laboratory found in Thomas Leahy’s home

indicting the production of ricin.

1997 Group of extremists in Minnesota create a home made

mix of ricin and chemical to target a US marshal.

2002 Kenneth Olson arrested for producing ricin.

2003 An envelope in Greenville, South Carolina mail facility
was found contaminated with ricin. Source never
determined.
2003 Six North African men arrested in London for
manufacturing ricin within their apartment.
2003 Secret services seized a letter contaminated with ricin
going into the white house with demands.
2004 Toxin ricin detected in the Dirksen Senate Office
Building.

Figure 1. Structure of Ricin Toxin (3Dchem.com).

 Ricin 30

Figure 2. Mechanism of Ricin (Audi et al., 2005).

Figure 3. Castor beans (Audi et al., 2005).

 Ricin 31

Figure 4 Countermeasure communication diagram (DHHS, 2006)

Glossary

CDC-Center for Disease Control and Prevention.

 Ricin 32

DHS- Department of Homeland Security.

ELISA - Enzyme-Linked ImmunoSorbent Assay.
EMS-Emergency Medical Services.
EOC-Emergency Operations Center.
EPA-Environmental Protection Agency.
ER - endoplasmic reticulm.
FBI- Federal Bureau of Investigation.
FDA- Food and Drug Administration.
HD-Health Department.
HHS- Department of Health and Human Services.
HSOC- Homeland Security Operations Center.
JFO- Joint Forces Operations.
JOC-Joint Operations Center.
LC-MS/M - Liquid chromatography tandem mass spectrometry.
MALDI-MS - Matrix-assisted laser desorption-ionization mass spectrometry.
LD50 - Median lethal dosage.
NRC-National Response Center.
RIP-II - type 2 ribosome inactivating protein.
RRCC- Regional Response Coordination Center.
USCG-United States Coast Guard.