Artemisia BioMedical, Inc
The Discovery of Artemisinin
Better by Natural Design™
March 2014
PNWBIO, Seattle
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(Re-)Discovery of Artemisinin
 Being considered for the Nobel Prize in Medicine
 The Nobel rules specify no more than three winners
and no posthumous prizes, either.
 Tu Youyou awarded 2011 Lasker~DeBakey Clinical
Medical Research Award for discovering artemisinin
and its use in treating malaria
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Science in China 1950-1960
 Stopped interacting with the West
 Younger scientists did not have access training
comparable to that found in Europe and the US
 Major project was the development of atomic and
hydrogen weapons and later artificial satellites
 Medical research projects were few
 Facilities were crude when compared to Western labs
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Cultural Revolution
 The Great Proletarian Cultural Revolution started in
1966
 Intellectuals, including scientists, were publicly
humiliated and forced to labor on collective farms.
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Cultural Revolution
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Scientific publications within China were stopped
Books and journals were recycled for their paper
Science was no longer taught at middle schools
Western training was cause for imprisonment or
banishment to the countryside
 However, the military protected scientists working on
projects of national importance
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1960’s Vietnam
 Late 1950’s program by WHO to eradicate malaria in
Southeast Asia had failed
 There was Plasmodium falciparum resistance to all
known anti-malarials including chloroquine
 Soldiers on both sides were infected with malaria
(roughly ½ at any given time)
 In 1966, experts were sent by the USA and China to
study the problem
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Malaria Transmission Cycle
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Vietnam Was an Ideal Malaria
Breeding Ground
 Vast tunnel network containing ants, poisonous
centipedes, snakes, and mosquitos
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China Offers to Help Ally
 Ho Chi Min asked Chairman Mao Zedong for help.
 Malaria was also a problem in southern China
 Hundreds of thousands of Chinese troops were in
Vietnam
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Prophylaxis First
 In 1966, scientists for the Chinese military began
reviewing existing antimalarial drugs for the
prevention of malaria.
 Three were selected and pairs were used in
combination tablets to use prophylactically against
malaria.
 A new cure was still needed.
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Project 523
 Leaders from the Chinese Military, the Ministry of
Health and the National Commission on Science and
Technology met on May 23, 1967
 Coordinating office located in the Military Academy
of Medical Sciences
 Participating institutions were spread across China,
involving hundreds of individuals, from Beijing,
Shanghai, Yunnan, Shangdong to Guangdong and
Guangxi
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Project 523
Composed of:
 The General Logistics Department of the Chinese People’s
Liberation Army
 The State Science and Technology Commission of China
 Ministry of Public Health of PRC
 Ministry of Chemical Industry of PRC
 Commission of Science and Technology for National Defense
 Chinese Academy of Sciences and
 The General Pharmaceutical Industry Corporation
 ~Five hundred scientific workers from sixty+ institutions
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Project 523 Aims
 New drugs for drug-resistant falciparum malaria
 Combine traditional Chinese and modern Western
medicine
 Emphasized discovery of new drugs from traditional
Chinese medicine
 Drugs should be safe with few adverse effects
 Drugs should be highly effective with rapid onset of
action
 Administered infrequently for prevention
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Two Approaches Used
 Copy Western drugs and synthesize new derivatives
• P. falciparum was resistant to existing drugs
• Modifying drugs would make them effective
• Combination therapy with existing and new drugs
 Investigate anti-malarial drugs frequently used in
traditional Chinese medicine and folklore medicine
• Malaria had been endemic to China for 3,000
years
• Different regions may have unique treatments
• Back to the future approach
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Synthesis of New Drugs
 Take existing drugs and make new family members
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Artemisia annua L
 Qinghao grows in south and north China
 168 BC - "Fifty-two Prescriptions”
from Han Dynasty Tombs
 340 AD - antimalarial application was
first described in Zhou Hou Be Ji Fang
("Handy Therapies for Emergencies")
 Crushed plants and crude plant extracts
were used by Chinese farmers in the
1950s and 1960s to treat malaria
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Beijing Institute of Materia Medica
 1970 – scientists reviewed ancient and current records
and identified 808 anti-malaria candidates
 ~100 candidates tested at the Military Academy of
Sciences in a mouse model of malaria
 Mouse model transferred to Beijing
 Soak Qinghao in water, compress the plant and drink
the juice to treat malaria
 Replaced ethanol extraction with ether extraction
(lower boiling point)
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Cooler Extraction
 Success rate with ether extraction material was 100%
cure rate in mice with P berghei
 The neutral portion of the extract was identified as
containing the active element
 Tested in animals and a few healthy volunteers and
deemed safe
 1972 – tested in 9 P vivax patients and 11 P
falciparium patients, failed in 2 P falciparium patients
 Project head office requested purified active element
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Clinical Failure
 Purified extract from Beijing Institute showed cardiac
toxicity in animals
 Tested on 3 researchers and did not show toxicity
 Field tested in 8 patients, 6 had normalized
temperature, but 2 patients had cardiac toxicity, and 2
showed no signs of cure
 Results not shared with project head office
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Relay Race
 1972 – Work at Beijing Institute duplicated in
Shandong and Yunnan provinces using their local
artemisia plants
 Both groups were able to make purified artemisinin
 Monomer isolated at Shandong Institute of Chinese
Materia Medica by Wei Zhenxing did not show
cardiac toxicity
 Luo Zeyuan from the Yunnan Institute of Materia
Medica got active isomers from ether extract
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Progress in Yunnan
 Yunnan group found their extract cleared P berghei
rapidly from mice and did not effect heart, liver, or
kidneys
 They identified a variant of Artemisia annua that had
the highest amounts of artemisinin from local plants
 They identified the Youyang area in the Sichuan
Province as having plants with 10-15 the
concentration of artemisinin
 Material was provided to other research institutions
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1974 – Clinical Success
 Group at Beijing could not produce purified material
 Shandong group compared crude with purified
artemisinin extracts in 26 patients with P vivax
 All patients were cleared of parasites and no evidence
of cardiac toxicity was seen
 Most patients had recurrence of parasites within 28
days
 Tried combining artemisinin with prophylactic tablet
to prevent recurrence
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1974 – Clinical Success Pt 2
 Material from the Yunnan group was used to treat P
falciparum
 The first 3 patients responded very quickly to
artemisinin; faster than existing therapies
 14 P falciparum patients including 3 with malignant
malaria were treated and 12 showed clearance of the
parasites by 20 hours
 Parasite development was arrested at the tiny-ring
form stage
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Findings Made Public
 1972 Artemisia annua (sweet wormwood) extract
shown to cure malaria
 Several groups of scientists at different institutions
needed to determine structure of active ingredient
 1977 first Chinese paper describing Qinghaosu but
omitting antimalarial action
 1982 first paper in English and available outside of
China
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Artemisinin and Derivatives
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Results of Project 523
 Chemical compounds were developed for the acute
and emergency treatment of malaria, prevention of
malaria, and eradication of malaria
 More than 10,000 chemicals synthesized, 40,000
chemical samples were screened, 1,000 possible
active substances identified, 38 underwent preclinical
screening, 29 approved for clinical trials
 14 drugs passed all testing and were used
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1979 Discovery of Qinghaosu Award
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Ministry of Health nominated 6 institutions
Academy of Chinese Traditional Medicine; Ministry of Health
Shandong Institute of Chinese Traditional Medicine
Yunnan Institute of Materia Medica
Institute of Biophysics of the Chinese Academy of Sciences
Shanghai Institute of Organic Chemistry of the Chinese
Academy of Sciences
 Guangzhou College of Traditional Chinese Medicine
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Were the Chinese First?
Milutin Stefanovic
1972, first publication
describing artemisinin
No mention of malaria
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Structure and Mechanism
 1993 - malaria parasite survives in its host by
consuming approximately 25% of the hemoglobin in
the host's red blood cells and stores iron
 Artemisinin comes in contact with the iron converting
it into a toxic chemical, releasing a free radical that
destroys the parasite
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Artemisinin Actions
 Kills malaria parasites
- Heme-derived iron reacts with artemisinin in food vacuole
to produce lethal amounts of ROS
 Kills cancer cells
- Free iron reacts with artemisinin in lysosomes to produce
ROS
- Death by activated programmed cell death pathways (PCD)
 Inhibits angiogenesis
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Cancer Cells and Iron
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All cells need iron for growth
Normal cells control iron uptake by neg feedback
Cancer cells accumulate iron beyond needs
Iron becomes a target for anticancer therapies
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Block iron uptake
Scavenge iron from blood
Use iron to trigger free radical reactions
Cancer cells deficient in free radical scavengers
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Iron and Cancer
 Cancer cell lysosomes contain higher levels of redox
active iron
- Nat Rev Cancer. 2005 Nov;5(11):886-97
 Breast cancer cells have higher transferrin receptor
expression levels and reduced ferroportin levels
- Sci Transl Med. 2010 Aug 4;2(43):43ra56. doi:
10.1126/scisignal.3001127
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Lysosomes
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Free Radical Reactions
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Creation of ROS is Iron Dependent
+iron
-iron
Cells treated with 0 (purple), 10 (green), 25 (red), 50 (blue), and 100 mM (orange)
DHA. Cells on right treated with iron chelator desferroxamine.
H Kenji, http://hdl.handle.net/2115/53216
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Programmed Cell Death Pathways
 Lysosomes accumulate free iron and artemisinin
 ROS from artemisinin causes membrane
permeabilization
 Hydrolytic enzymes are released from the lysosome
lumen into the cytosol
 Cathepsins activate pro-apoptotic proteins
 BH3 interacting-domain death agonist and caspase 8
activate mitochondrial outer membrane
permeabilization and apoptosis
 J Biol Chem. 2011 Feb 25;286(8):6587-601
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Artemisinin Multiple Anticancer Effects
Multi-targeted,
Selectively Promiscuous™
Firestone GL, Sundar SN. Anticancer activities of artemisinin and its bioactive derivatives. Expert
Rev Mol Med. 2009 Oct 30;11:e32.
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NCI In Vitro Cancer Screening
 Trioxane Dimers exhibit cancer cell cytotoxicity superior to
gold standard chemotherapy in NCI screening panel
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Effect of Art Derivative in vivo
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Questions?
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