Biomaterial Research Japan

7/28/2019 Biomaterial Research Japan

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EMBASSY OF FINLANDNational Technology Agency3-5-39, Minami-Azabu

Minato-ku, TOKYO 106-8561Tel:+81 3 5447 6035 Fax:+81 3 5447 6046

TEKES/TOKYO TECHNOLOGY SCANNING

Biomaterial Research in Japan

February 22, 2001

Tadaaki (Todd) Toyoshima

TEKES, Tokyo

Preface

This report lists who does what in the various fields of biomaterial research in Japan. Literature surveys

and interviews with researchers are recommended in order to more thoroughly research a specific

theme or topic. Tekes Tokyo will be pleased to help you in this respect.

Additional information: [email protected]


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Content

1 Strength and Weakness of Biomaterials in Japan.............................................................................................................42 Market ..................................................... ........................................................... .......................................................... ....4

3 Research on Biomaterials.................................................................................................................................................93-1 Academic Societies.................................................................................................................................................9

3-2 Major Biomaterial Research Groups ....................................................... ................................................................. ....10

4 Ceramics.........................................................................................................................................................................11

4-1 Introduction ..........................................................................................................................................................11

4-2 Artificial Bone Market..........................................................................................................................................12

4-3 Calcium Phosphate Paste......................................................................................................................................14

4-4 Advanced bone materials......................................................................................................................................14

4-5 Bone generation---tissue engineering ...................................................................................................................16

4-6 Drug delivery system using ceramics ...................................................................................................................18

4-7 Ceramics for cancer treatment ..............................................................................................................................18

5 Metal ..............................................................................................................................................................................18

5-1 Artificial Joints Market.........................................................................................................................................18

5-2 Titanium alloy on the market................................................................................................................................20

5-3 Titanium alloy under research ..............................................................................................................................21

5-4 Metals for dental application ................................................................................................................................21

5-5 Surface modifications ...........................................................................................................................................22

5-6 Other Metals .........................................................................................................................................................23

6 Polymers and Lipids.......................................................................................................................................................23

6-1 MS Coat................................................................................................................................................................24

6-2 MPC......................................................................................................................................................................24

6-3 Photoreactive polymers ........................................................................................................................................24

6-4 PIPAAm................................................................................................................................................................25

6-5 Polymeric Micelles ...............................................................................................................................................26

6-6 Nanoparticles and Microcapsules .........................................................................................................................26

6-7 Liposomes.............................................................................................................................................................27

6-8 Polysilamine .........................................................................................................................................................27

6-9 Perflourocarbonic acid..........................................................................................................................................28

6-10 Shape Memory Gels .............................................................................................................................................286-11 Hollow fibres ........................................................................................................................................................28

7 Drug Delivery and Targeting .........................................................................................................................................29

7-1 Overall situation ...................................................................................................................................................30

7-2 Active targeting ....................................................................................................................................................32

7-3 Passive Targeting..................................................................................................................................................32

7-4 Stimulus Responsive Materials.............................................................................................................................327-5 Electromagnetic Responsive Double Targeting....................................................................................................33

7-6 Biomolecular Design for Biotargeting..................................................................................................................337-7 Transdermal & Transmucosal Delivery................................................................................................................34

7-8 Company activities ...............................................................................................................................................35

8 Gene Delivery ................................................................................................................................................................36

8-1 Gene Therapy Protocols .......................................................................................................................................368-2 Research on non-virus vectors..............................................................................................................................37

8-3 Polycations and Liposomes ..................................................................................................................................37

8-4 Polymer Micelles ..................................................................................................................................................38

8-5 Fullerene and Cyclodextrin...................................................................................................................................38

9 Tissue Engineering.........................................................................................................................................................399-1 Regenerative Medical Engineering Programme by the Ministry of Education ....................................................39

9-2 The Yoshizato Group at Hiroshima University ....................................................................................................42

9-3 3D Tissue Module Engineering conducted by NAIR...........................................................................................43

9-4 Millennium Project...............................................................................................................................................43


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1 Strength and Weakness of Biomaterials in Japan

Japan is a world leader in dialysis membranes and artificial kidneys, with more than 50% of the world

market, owing in part to large fibre manufacturers such as Toray and Asahi Chemical. Furthermore,

Japanese companies excel at disposable materials. In contrast, most implant materials are imported,

with one exception, ceramic bone. Japan has been historically strong in ceramics, not to mention

electronic condensers and fine ceramics in the semiconductor industry. Developments in metal for

artificial joints are also catching up with the West.

There are many biomaterial researchers in Japan. But, Japanese companies are reluctant to take the risk

to be a pioneer in implant applications or prostheses. Japan's strength lies in external rather than

implanted components and devices. Most implanted materials have been developed in the West.

Components used in interface areas, such as dental materials and contact lenses, are both made in Japanand imported.

In Japan, researchers, in close collaboration with clinical doctors, have the advantage in discovering

clinical needs and conducting clinical trials. In this respect, bio-material institutes in medical

universities have many actual products to bring to market.

Most material researchers consider the future of biomaterials to be in tissue engineering or, more

narrowly, human cell engineering.

With regard to drug delivery systems (DDS), Japan has put successful DDS products on the market,

such asLeuprin, polymer nanosphere DDS; and SMANCS, a drug-polymer conjugate suspended in

lipiodole. Adhesive transdermal delivery systems, such as pressure sensitive adhesives containing

isosorbide dinitrate, are also advanced. Many controlled release type DDS have been developed and the

future seems to be in organ, cell, and molecule targeting.

In tissue engineering, there are at least three government-supported programmes under way.

The government has selected tissue engineering as one of the Millennium Projects, starting two

research institutes in the Kobe area: one for basic study and another for application.

2 Market

Japan imports most of its implant materials or devices. Valves for artificial hearts are 100% imported;

artificial blood vessels, 95%; bone bonding materials, 80%; artificial joints, more than 80%; and pace-

makers, 90%.


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Medical equipment and materials market in Japan

Production

(million

Yen)

Import

(million

Yen)

Expor

(millio

Yen)

Grand Total 1,487,903 834,509 365,042

Image diagnostics

systems

298,356 82,872 127,215

Treatment

equipment

194,395 196,049 46,408

Measuring and

monitoring systems

172,116 46,393 95,011

Total 163,445 267,879 28,232

Heart Valves 7 11,551

Pacemakers 55 44,042

Blood Tubes 109 5,941

Stents 384 14,905 16

Joints and Bones

total

20,187 84,534 10

Hip joints 8,195 20,167 10

Knee joints 1,234 13,969

Shoulder joints 139 358

Elbow joints 97 917

Metal bone 63 2,995

Resin bone 250 274

Ceramic bone 2,937 19Bone bonding

materials

5,442 41,061

Bone cement 44 1,789

Devices and

Equipment for

Function Support or

Replacement

Joints and Bones

Other related 1,781 2,979


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Intraocular Lens 3,936 12,268

Monitors & liquid

supply for dialysis

13,842 421 1,213

Dialysis equipment 73,912 2,499 22,094

Pumps &accessories for

Artificial Lungs

1,429 3,582 372

Artificial Lungs 2,261 2,922 678

Blood Cleaning

equipment

14,834 4,906 1,856

Blood circuits 11,962 15,101 1,072

Home Use Medical

equipment

151,691 8,048

X ray equipment 103,074 43,511 16,618

Total 90,752 17,026 2,142

Metal 50,378 4,005 27

Crowns (4/5 are resin) 6,022 907 232

Dentures (Almost all are

resin)

2,825 165 211

Bonding, Filling

materials

(1/2 are resin) 14,970 3,653 1,600

Impression

materials

5,278 1,180 17

Dental materials*

Others 5,071 547 33

Eye-related

equipment

87,647 49,129

Biopsy inspection

equipment

85,066 22,329 19,796

Healing & surgery

equipment

61,843 50,755 7,126


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Dental equipment 35,767 7,616

Equipment for

facility

29,095 2,979

Steel equipment 8,080 21,966

Hygiene materials 6,575Source: Ministry of Health and Welfare Medical Industry Statistics 1999.

*Issued a report, in Japanese, on The Market for Dental Equipment, Materialsand Chemicals published

Research Institute, is available (cost - Yen 100,000).

Medical equipment and materials market in JapanGrand total growth rate

Production Import Export

1995 1.4 % 17.5 %

1996 8.9 20.5 11.3 %

1997 4.0 5.8 21.8

1998 -0.4 11.2 21.7

1999 -1.3 0.0 35.8

The shipment and growth rates of implant materials

Shipment in 1999

(100 million Yen)

Growth rate in

1998/1997

(%)

Growth rate

Forecast in

1999/1998(%)

Cardiovascular equipment and

materials

710 7.4 15

Artificial bones and joints 513 -17.8 10


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Blood cleaning and separation

equipment

372 43 No data

Dialysis equipment 748 10 10

Total 2,524 7.7

Source:Market for Medical Equipment and Materials, Yano Research Institute.


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3 Research on Biomaterials

3-1 Academic Societies

According to Dr. Tsuruta, Emeritus Professor, Tokyo University, Japan has a long history in

biomaterials. Research on polymers for medical use started in 1958. The Higuchi model for a drug

delivery system was reported in 1961. The Toray PMMA hollow fibres for artificial dialysis were put

on the market in 1973 and the Kuraray EVA in 1978. The Ministry of Education funded biomedical

polymer research programmes from 1972 through 1989 under various programme names.

Prof. Tsuruta emphasised, in his keynote speech at the 2000 Biomaterial Conference, that all the

development work should have clinical applications as the target. He encouraged researchers to protectthemselves against unreasonable lawsuits. Companies should develop their risk management strategies

to deal with the biomaterial products and business. He predicted that future biomaterial research would

shift to tissue engineering.

The following academic societies are focused or have groups focused on biomaterials:

Japanese Society for Biomaterials was founded in 1978. It organises an annual conference and

publishes a bimonthly journal in Japanese. In addition to individual members, there are about 50

company members. The next annual conference will be held in Kyoto on October 22 and 23, 2001.

The Ceramics Societyof Japan formed a Bio-related Materials Working Group in October 1998. Thegroup has about 120 members and holds a conference and a Beginners Seminar once a year in

November or December. The society publishes two Japanese monthly publications calledJournal of

Ceramic Society of Japan (for papers) and Ceramics Japan (for reviews).

Japan Metal Society has a temporary (March 1998February 2001) group called High Performance

Biomaterial Study Committee. The committees aim is to review the appropriate data for the

development of biomaterials, and to decide the current status of evaluation methods and standards.

There are no industrial standards for biomaterials in Japan.

The Society of Polymer Science, Japan has a Research Group on Biomedical Polymers, which was

founded in 1972. The research group, 150 members, holds conferences on special themes, and review-type seminars. The society publishes an English-language monthly calledPolymer Journal.

Japan Society of Medical Electronics and Biological Engineering was founded in 1962, and now

has 4,000 members. It publishes an English-language quarterly calledFrontiers of Medical and

Biological Engineeringas well as a monthly journal in Japanese.

Japan Society of Drug Delivery Systems was founded in 1988 based on the DDS Study Group that

began in 1984. It now organises annual conferencesthe next is on July 12, 2001 in Osaka with

around 500 researchers expected to attend. The Society publishes a bi-monthly journal in Japanese

calledDrug Delivery System. The number of members is over 1,000. The Japanese Society of Drug


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Delivery Systems jointly publishes theJournal of Controlled Release (JCR) with the international

Controlled Release Society.

Japan Society for Tissue Engineeringbegan in 1984 asthe Organ Formation Working Groupandadopted its current name in 1998. It organises an annual meeting but does not publish a journal. Rather

it encourages Japanese members to send papers to Tissue Engineeringin the USA.

3-2 Major Biomaterial Research Groups

Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental UniversityThe oldest biomaterials research institute in Japan has three divisions: the Functional Molecule

division; the Materials divisionmetal, inorganic and organic materials; and the Bio Systems division.

The institute started dental material research in 1938 and diversified in 1966 to cover other medical

fields. It has more than 50 permanent researchers.

Tissue Engineering Research Institute at the Institute of Frontier Medical Sciences, Kyoto

UniversityThis institute is one of the largest Japanese research groups in tissue engineering. The institute was

founded in 1978 as the Medical Polymer Research Group and developed into the Research Center for

Medical Polymers in 1980, changing its name to Research Center for Biomedical Engineering in 1990,

and adopting the current name in 1998. With approximately 80 researchers, it covers all fields relatedto tissue engineering: biological function, tissue engineering, regeneration control, medical system

engineering, and clinical applications.

Professor Ikada was former head research director of the institute.

Institute of Biomedical Engineering, Tokyo Womens Medical UniversityProf. Okano, head of the institute, is an expert in temperature responsive polymers and his group is

working on applications of the polymers for DDS and gene vectors. Cell Sheet Engineering

technology is one of the approaches, promoted by the Institute, to produce complicated organs by

accumulating cell sheets one by one using temperature responsive surfaces, similar to the building of

large-scale integrated circuits. They recently started collaboration with the Science & Engineering

Department of Waseda University.

Professor Yoshizato's group, at Hiroshima University. This group is unique in that they are targeting

technology development based on tissue engineering to promote local industry, getting support from

both the central and local government and from close collaboration with companies.

3D Tissue Module Engineering group in NAIR

The 3D Tissue Module Engineering groupin the National Institute for Advanced Interdisciplinary

Research, under the Ministry of Economy and Industry, was formed in 1998 with more than 40


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researchers, including 11 foreign researchers. This group will likely be a core member in the new

laboratory in Osaka called METI Tissue Engineering Center, to be founded in 2002.

The International Symposium on Advanced Biomaterials and Tissue Engineering held in 1999 wasorganised by NAIR to promote industrial applications.

National Research Institute for Metal and National Institute for Research in Inorganic Materials,

bothunder the Ministry of Education and Science, are merging in April 2001. Each institute has a

biomaterial research group. After the merger, they will have approximately 15-20 researchers.

4 Ceramics

4-1 Introduction1

According to Prof. Nakamura, Kyoto University, there have been three generations of artificial bone:

the first generation is sintered hydroxyapatite (HAP) and A-W glass ceramics, the second generation is

porous material and Ca-P cement, and the third is the material together with a complex of bone growth

factors and bone cells.

Three materials for bone replacement have been invented since the 1970s: Bioglass, Sinteredhydroxyapatite, and Crystallised glass A-W. Japanese researchers have made a large contribution to thelatter two.

Dr. H. Aoki, former professor at the Tokyo Medical and Dental University, succeeded in synthesising

HAP and manufacturing sintered HAP in 1972, simultaneously but independently from investigators in

the USA. He discovered the high affinity of HAP to real bone in 1975. His research theme for

developing the manufacturing technology of synthesised HAP for artificial bone and teeth was funded

by the Ministry of Education for five years beginning in 1977.Apaceram now sold by Asahi Pentax

Co., the top market shareholder of bioceramics in Japan, is based on his research.

Prof. Kokubo, from Material Chemistry at the Kyoto University, announced in 1982, and began usingin 1991, Crystallised Glass A-W (SiO2-CaO-MgO-P2O5), which is made by the deposit of fine crystals

of HAP and wollastonite: CaOSiO2. Nippon Electric Glass Co. has sold A-W as Cerabone since 1991,

with annual sales of approximately USD 7 million. It has been used on 45,000 patients. They decided,

however, in October 2000 to discontinue the business at the end of 2000, because of slow growth and

the poor prospect for future profits.


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Comparison of physical properties

Bioglass HAP A-W Dense

natural bone

Spongy

natural bone

Density gcm-3

2.68 3.16 3.07 1.6-2.1Hardness HV 458 600 680

Compressed strength

MPa

500-1000 1080 100-230 2-12

Bending strength

MPa

42 115-200 215 50-150

Young modulus

GPa

35 80-110 118 7-30 0.05-0.5

Breaking tenacity

MPam1/2

1.0 2.0 2-6

Bone invasion speed

into ceramics space

Highest Low high

Containing SiO42-

(may be toxic)

Yes No Yes

Source: L.L.Hench (1998)J.Am. Ceram. Soc., 81, 1705-28

Increasing the tenacity and decreasing the Young Modulus of artificial bone to the level of natural bone

is a major challenge. One method of increasing tenacity is to coat titanium alloy with HAP. Another

approach is to use collagen-apatite composites. Hybrid materials of organic and inorganic components,

bonded at the molecular level to produce material similar to spongy bone, are being investigated.

Ceramics that can be moulded in surgery rooms are also required. Mitsubishi Material Co. has

developed biodegradable calcium phosphate bioactive paste (major component: a-Tri-Calcium

Phosphate) independently from Norian. Mitsubishi started selling the product called BIOPEX in June

2000 through the Welfide Corporation and Taisho Pharmaceutical Co.

Ceramics are also used in cancer treatment and drug delivery systems.

The major direction of current research is to develop materials to promote bone generation by means of

tissue engineering.

4-2 Artificial Bone Market

Market2

In 1998, the market size was USD 35 million, with a 4-5% growth rate expected until the market for

osteoporosis treatment takes off. The markets for artificial bones in foreign countries are small because

human and animal bones can be easily used. In Japan, such reuse is disliked for social and religious

reasons.


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There are two types of artificial bones: ceramics and resin. Sales of artificial bone ceramics totalled

USD 30 million and are growing. Artificial bone resin sales were USD 2 million in 1997 and have

levelled off. A liquid type resin, which solidifies within the human body, has attracted attention due topotential use in osteoporosis treatment, such asBIOPEXby the Mitsubishi Material Co.

Manufacturers

Manufacturers sales figures

Makers Sales

representative

Sales in

1999

(million

Yen)

Brand names and product types

Asahi Optical Asahi Optical 1,410 APACERAM: HAPNippon Special

Ceramics

Chugai

Pharmaceutical

600 CERATITE: Complex of HAP and

tri-calcium phosphate (TCP)

Sumitomo Osaka

Cement

Sumitomo

Pharmaceutical

560 BONECERAM: HAP

Both porous and dense types are

available.

Nippon Electric

Glass

Discontinued

bioglass business

Dec. 2000

415 BIOGLASS A-W

Mitsubishi

Materials

Welfide

Corporation

331 BONEFILL and BONETIGHT: HAP

BIOPEX: Paste ofa-TCP and 4CP

Kyocera Welfide 240 BIOCERAM: TCP

(Kyocera is an artificial joints

manufacturer)

Olympus Olympus 100 Osferion: b-TCP

Both porous particles and blocks are

available.

Source: The Market for Dental Equipment, Materialsand Chemicals, October 1999, Yano Research.

Direction of technology

Hard bone:

Increase of bone-forming ability with marrow transplanting and bioactive substances

Mitsubishi calcium phosphate paste will be followed by competitors

Composites

Soft bone or cartilage:

Replacement of cartilage with artificial bone has not yet succeeded clinically; therefore, joints need

to be completely replaced.


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The abrasion of joint sliding surfaces is still a challenge to be solved. Alumina-alumina sliding

surfaces and the composites of zirconia and alumina are under study.

4-3 Calcium Phosphate Paste

BIOPEX

Mitsubishi Materials Co. launched a calcium phosphate paste in June 2000. The National Health

Insurance system listed the material in 1999.

At the Japan Biomaterial Conference held in November 2000, Dr Shigeo Miwa from the Aichi Medical

University, reported the clinical trial results of Mitsubishi MaterialBIOPEX.

BIOPEX components arePowder:

Ca3(PO4)2: 75 % of the weight

Ca4(PO4)2O: 18

CaHPO42H2O: 5

Ca10H(PO4)6(OH)2 2

Liquid:

5 % chondroitin sulphate

12 % sodium succinate

83 % distilled water

BIOPEXcompletely changes to apatite in the body in three weeks. Its degree of crystallisation is low

and is similar to human bone. Unlike PMMA cement, it does not generate polymerisation heat nor

leach non-polymerised monomers to the cardiovascular system. The material is gradually absorbed by

the body: 50 % in 3 weeks. Its shortcoming is the weak compressive strength: 6090 M Pa. Therefore,

it is not suitable for artificial bone fixing.3

At the Biomaterial Symposium 2000, S. Yamamoto, of the Department of Oral Functional Science, at

the Hokkaido University, in co-operation with Asahi Optical Co., presented an application of Calcium

Phosphate Cement containing CM-chitin and succinic acid for bone augmentation. In rabbits, he

observed new bone formation in a crack in the skull.

4-4 Advanced bone materials4

Many researchers are still trying to develop next generation apatite using composites and surface

modification.

Nitta Gelatin Inc., together with Dr. Junzo Tanaka and Dr. Masanori Kikuchi, of the National Institute

for Research in Inorganic Materials, has developed artificial bone that has a flexural strength of about


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50 MPa, nearly comparable to that of the human bone (40200 MPa). It is a composite consisting of

protein-I type collagen and hydroxyapatite. A co-precipitate of a calcium hydroxide solution and an

aqueous phosphate solution containing collagen is formed via a cold isostatic pressure process. When

the new composite was transplanted onto a rat cranial bone, the composite started to be replaced bynatural bone in two weeks.

Nitta Gelatin Inc. is also manufacturing collagen for artificial skin in medical treatment.

National Industrial Research Institute of Nagoya has found that chitin and chitosan derivatives

containing phosphoric acid are effective in forming bone. The surface of chitin or chitosan is

modified by phosphoesterification and the derivative put into phosphoric acid and calcium-containing

liquid. After 10 days, the same volume of calcium phosphate as the derivative was obtained.

Electrically polarised hydroxyapatite is being studied by K. Yamashita, T. Kobayashi, andcolleagues, at the Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University.

In an oral presentation at the Biomaterial Symposium 2000, they reported that they found that apatite

can be electrically polarised and that the stored polarised energy can be measured. They also implanted

the polarised apatite in a dog, to evaluate bone formation, and found that new bone started to form on

the negative surface in less than a week. This phenomenon can find application in orthopaedic and

dental fields. The polarisation can stay in vivo for at least six months.

Surface modification of HAP with carboxyl group was reported by Dr. T. Matsumoto, and

colleagues, of the Dental School, at Osaka University, together with Dr. M. Okazaki, of Hiroshima

University, and Dr. Taira, of the Iwate Medical University. Their idea is based on the fact that bonegrowth factors such as b-FGF and BMP have affinity to negatively-charged surfaces. They succeeded

in producing HAP modified with a carboxyl group and evaluated the material using rats. They found

that the weight of the adhered protein and the cell adhesion ability depend on the volume of the

carboxyl group on the surface.

Zinc-releasing calcium phosphate on collagen is being studied for the promotion of bone generation

by Dr. Atsuo Ito, and colleagues, at the National Institute for Advanced Interdisciplinary Research

(NAIR), together with T. Hikita, and colleagues, at the Waseda University. They made a NaCl,

CaCl22H2O, KCl, ZnCl2 solution, introduced CO2, NaHCO3, Na2HPO412H2O, and put collagen sheet

in it. Low crystal apatite containing Zinc was obtained.

Dr. H. Kawamura, Kennan Hospital, together with Dr. Ito, at NIAIR, and S. Miyakawa, at the Tsukuba

University, conducted a long-term implantation test ofZinc-releasing calcium phosphate ceramics in

rabbits. At an oral presentation at the Biomaterial Symposium 2000, they reported that

0.316%ZnTCP/HAP significantly increased the bone-binding ratio but also increased bone absorption.

They are currently optimising the zinc content.

At a poster presentation at the Biomaterial Symposium 2000, results with magnesium-containing CO3

apatite-collagen composite were reported by Dr. Yasuhiko Yamasaki, of the Dental School of


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Hiroshima University, together with Y. Hamada, and colleagues, of the Dental School, at Osaka

University. They tried to synthesise crystals of magnesium-containing CO3 apatite starting from a

Ca(CH3COO)2H2O and Mg(CH3COO)24H2O solution. The powder crystal obtained is mixed with

0.5w% collagen to produce cylindrical pellets. The pellets are put in serum containing mouseosteoblasts. Adhesion assays revealed that the Mg composite showed higher adhesion than non-Mg

CO3 apatite.

Dr. Tanaka, at the Research in Inorganic Materials, and Prof. Miyairi, at the Tokyo Medical and Dental

University, are collaborating on creating a composite ofpolylactic acid and TCP for bone deficiency

20mm and over. They produced the composite with 20-50% polylactic acid and 80-50% TCP to make

0.3mm membranes. With conventional materials, only a 5mm bone deficiency can be rebuilt.

Carbonate apatite is being studied by Dr. Masahiro Hasegawa, and colleagues, in OrthopedicSurgery, at the Asahi Memorial Hospital, because human bone contains carbonate ions. They reported

at a poster presentation at the Biomaterial Symposium 2000, that they implanted porous apatite with 6

wt% carbonated ions in rabbits. Increased bone forming and the absorption of the material were

observed four weeks later.

Prof. Masanori Oka, Kyoto University, is developing artificial joint cartilage with titanium fibre

mesh injected with PVA hydrogel under high pressure. The properties of lubrication, shock

absorption, and the bio-affinity and adhesion to the bone bed are acceptable, except for the anti-

abrasion. Therefore, the material is suitable for partial hemiarthroplasty for fractures of head of the

thigh bone, osteochondritis dissecans, and chondromalacia. This is joint research with China andclinical applications are expected to be appear in a couple of years.

4-5 Bone generation---tissue engineering5

Professor Kunio Takaoka,inorthopaedic surgery, at the Shinshu University, received competitive

funding, of 21 million Yen a year, from Ministry of Education for his research on basic theory and

operation technique of Bone Morphogenetic Protein (BMP). He used a copolymer of polylactic acid

and polyethyleneglycol for the delivery of BMP, as well as for the scaffold. He succeeded in the

regeneration of the upper arm bone of rabbits in eight weeks. But the mechanical strength and

absorption of the scaffold was unsatisfactory.

Prof. K. Shinomiya,of the Tokyo Medical and Dental University, made a block ofHAP and collagen

composite attached with rhBMP and implanted it in a beagle. Bone generation was observed.


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Prof. Takafumi Yoshikawa, of the Department of Pathology at the Nara Medical University, and

colleagues, in co-operation with T. Uemura, and colleagues, at NAIR, have cultured rat marrow cells

to make bone. Immediate bone generation right after the transplantation of the cultured bone was

observed and osteoblast activity was maintained for a long period. The technology was confirmed withosteoblasts in 29 patients with osteoarthritis and rheumatoid arthritis and injuries.

At a presentation at the Biomaterial Symposium 2000, Dr. Ohgushi, in orthopedic surgery, at the Nara

Medical University, reported that he is going to culture marrow cells of patients who undergo bone

shaving surgery, taking marrow liquid and culturing the cells for three weeks to transplant them to the

patients.

Professor Kinoshita, of the Kanagawa Dental University, has machined PLLA (poly-L-lactic acid) to

meet the shape of the jaw and put sponge marrow taken from waste bone, regenerating jaw bone in six

months to one year. He has performed the procedure, clinically, 50 times. The success rate is 80%.

The National Institute for Research in Inorganic Materials and Institute of Biomaterials and

Bioengineering, Tokyo Medical and Dental Universityhave succeeded in developing a complex of

tricalcium phosphate (TCP) 70% and polylactic acid 30% to make 0.3mm thin membranes to

regenerate lost bone as much as 20mm wide. TCP accelerates bone generation and absorption rate of

the materials can be controlled by changing the ratio of the two materials. Moreover, the membranes

thermoplasticity makes it easy to form various shapes during surgery.

Guoping Chen, and colleagues, of the 3D Tissue Engineering Group, NAIR and T. Sato, at the

Graduate School of Medicine, Tsukuba University, have generated cartilage using biodegradablehybrid sponge. At the Biomaterial Symposium 2000, they reported that they introduced collagen

sponge into the pores of a copolymer sponge of lactic acid and glycolic acid. They successfully

cultured, both in vitro and in mouse, the cartilage cells of the knee joints of cows.

Slow release of BMP with gelatin hydrogel was successfully achieved by Dr. Masaya Yamamoto, at

the Institute of Frontier Sciences, Kyoto University. Release control was accomplished by changing the

water content of the biodegradable hydrogel. The hydrogel was a chemically bridged caustic gelatin

with glutalaldehide.

BMP easily dissipates in the human body even if it is injected. Dr. N. Saito, and colleagues, at the

Shinshu University, are developing an injectable DDS for BMP. They synthesised several blocks of

PLA-PEG copolymers, differing in molecular weight, and injected rhBMP-2 and the copolymers at the

temperature of 60 degree C into the front of a mouse femur and evaluated the bone generation.

It was reported in the Nikkei news that Takeda Chemical Industries Ltd. has developed a new

compound contained in slow release polymer capsules, 30 mm in diameter, that promotes the fusion of

fractured bone by activating a natural protein in the body for a month. Takeda is now moving to a

clinical trial after confirming its effect using mice. The compound, with the development name of


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TAK778, was originally licensed from a Hungarian company and improved by Takeda. It speeds up the

healing process of fractured bone by a third.

4-6 Drug delivery system using ceramics

Professor Makoto Otsuka, from the Kobe Pharmaceutical University, is one the most active researchers

of drug delivery systems using bone cement. He has studied apatite cement, bioactive glass and

complex of bioactive glass, each containing indomethacin. He found that apatite cement suppresses

drug release after a week, while bioactive glass allows drug release, even after three weeks. To

overcome the brittleness of the bioactive glass, he also tried bisphenol-a-glycidyl methacrylate and

triethylene-glycoledimethacrylate, resulting in a decrease in drug release after 100 hours.

During tests, he found drug release speed decreases when the calcium concentration in the liquid or the

blood is high. He is trying to apply his finding to a drug release system for osteoporosis patients usingestradiol. Calcium concentration in blood in osteoporosis rats is lower than normal rats, thus estradiol

release in bioactive glass is increased.6

4-7 Ceramics for cancer treatment

It is preferable to irradiate only locally to cancerous tissue. Y2O3-Al2O3-SiO2 glass was reported to be

suitable for local irradiation in 1987 by G.J. Ehrhardt and D.E.Day. It was achieved, with neutron

radiation of 20-30 mm sphere glass, resulting in conversion of89Y to 90Y, which radiates b rays. Dr.

Kawashita, Kyoto University, has shown that only Y2O3 microspheres with a diameter of 20-30 mm can

be easily obtained by high frequency induction heat plasma. Y2O3 shows excellent chemical durability,

better than Y2O3-Al2O3-SiO2 glass. Animal trials using the spheres are about to begin.

Normal cells can survive up to 48C but cancer cells die at around 43 C. Therefore, it could be

effective to heat up cancer tissue in deep parts of bodies, but methods of local heating are not available.

Dr. Kawashita has developed fine crystals of magnetite, iron, or iron zinc ferrite deposited in CaO-SiO2glass matrix and Li ferrite deposited in SiO2-P2O5 glass matrix. The heat generation of these

ferromagnetic spheres has been tested using rabbits. Other ferromagnetic spheres with higher heat

generating efficiency are under study.

5 Metal7

5-1 Artificial Joints Market


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Market

Units of artificial joints

1998(pieces) Growth rate(%) 1999(pieces) Growth rate(%)

Hip joints 55,742 2.2 58,932 5.7

Knee Joints 27,527 6.9 30,287 10.0

Shoulders 835 0.6 865 3.6

Knees 362 16 470 29.8

Others 508 1.8 705 38.8

Total 84,974 3.7 91,259 7.4

Source: Yano Group

Sales of artificial joints

1998(million Yen)

Growth rate(%)

1999(million Yen)

Growth rate(%)

Hip joints 40,071 2.6 42,488 6.0

Knee Joints 18,028 -0.1 19,951 10.7

Shoulders 418 12.9 418 0.0

Knees 160 25 217 36.6

Others 184 -1.4 260 41.0

Total 84,974 1.9 63,335 7.6

Source: Yano Group

Titanium alloys are increasingly used instead of SUS316 Co-Cr alloys (approximately 80 % of themarket) and have been receiving attention from researchers.

Major manufacturers

Companies Market share

(%)

Notes

Stryker Japan 26

BMS Zimmer 25

Debu Japan 12Kyocera 11 Kyocera has made 8,700 zirconia-based joints

since 1995.

Kobelco 2 The top titanium maker in Japan

K-MAX hip joints

Mizuho Medical

Nemoto Trading

Nakshima Propella

The market share of domestic companies is about 14% including Mizuho Medical, Nemoto Trading,

and Nakashima Propeller, in addition to Kyocera and Kobelco.


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Nakashima Propeller Co. Ltd. is expanding their artificial joints business to USD 20 million in five

years, three times the current amount. They are competitive in titanium alloys rather than ceramics.

Research Direction

Metals for orthopedic applications

(1)Vanadium-free Titanium alloys to replace the most popular Ti alloy: Ti-6Al-4V, that contains

cytotoxic Vanadium.

(2)Alloys with low modulusclose to that of real bone.

(3)Nickel-free shape memory and ultra-elastic alloysNickel may have adverse physiological effects.

Soft bone and cartilage:

Replacement of only a part of cartilage with artificial bone has not been clinically successful, therefore,

joints are completely replaced. Furthermore, the abrasion of joint sliding surfaces is an issue.

(1)Cross-linking of ultra high molecular weight polyethylene

(2)Alumina-alumina sliding surface and composites of zirconia and alumina

Dental applications:

(1)Alloys with low melting point are desired.

(2)Alloys easy to machine are desired

5-2 Titanium alloy on the market

A Vanadium-free alloy, Ti-6Al-7Nb, has been developed and put into practical use in Europe. In Japan,

a few Vanadium-free alloys are on the market or under research.

The Medical Material Division at Kobelco Co. developed Ti-15Mo-5Zr-3Al for cement-type hip

joints and Ti-6Al-2Nb-1Ta for cement-less hip joints. The objectives of Kobelcos development were

to avoid Vanadium and to achieve high fatigue strength. Ti-15Mo-5Zr-3Al was put into clinical trials in

1991 and has been used since 1995. Because its fatigue strength is 20% higher than that of Ti-6Al-4V,

the stem neck out-diameter can be made as small as 9mm.Therefore, the moving area of hip joints is

larger, which results in longer life for the joints.

Ti-6Al-2Nb-1Ta was put into a clinical trial on 1992 and onto the market in 1996. This alloy is less

susceptible to strength deterioration after heat treatment.


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Kobelco so far has used Ti-15Mo-5Zr-3Al with 3,500 patients and Ti-6Al-2Nb-1Ta with 2,000

patients.

5-3 Titanium alloy under research

Yoshimitsu Okazaki, at the Mechanical Engineering Laboratory under MITI, has developed, in co-

operation with Kobelco, Ti-15Zr-4Nb-4Ta-0.2Pd. Zr was added for higher strength; Nb and Ta were

added for improved anti-corrosion; Nb and Ta content was kept low for a lower melting point. The

composition of the alloy was optimised for a good balance of bio-compatibility, improved anti-

corrosion, and a low melting point. The alloy is currently being tested on rats.

Dr. D. Kuroda and Professor Mitsuo Niinomi, at the Toyohashi University of Technology; Dr. Fukui,

and colleagues, of the Dental School, Aichi Gakuin University; T. Kasuga, of the Nagoya Institute of

Technology; and Daido Special Steel Co. are developing Ti-29Nb-13Ta-4.6Zr alloy. The goal is to

design a new b-type titanium alloy that has a similar Young modulus as bone, but without using

cytotoxic materials. Biocompatibility of the alloy, evaluated via cell survival ratio, is the same as

titanium and better than Ti-6Al-4V.

They have also coated the alloy with calcium phosphate glass: 60CaO30P2O53TiO2 to improve bio-

compatibility, and found that the adhesion of the glass to the alloy is extremely strong.

Professor Hideki Hosoda, of Tsukuba University, an expert in shape memory alloys, is developing Ti-

based, Ni-free, shape memory. He has so far achieved good mechanical properties and high modulus

by adding Nb/Mo forb-phase stability and Al/Sn/Ga fora-phase stability. It seems he can develop

shape memory alloys with the specified elements, for medical use. He is looking for commercial

collaboration with researchers in the medical field, to study corrosion and bio-compatibility.

5-4 Metals for dental application

Dr Yoneyama, and colleagues, of the Institute of Biomaterials and Bioengineering, Tokyo Medical and

Dental University, are studying Ti-6Al-7Nb alloy produced by Daido Special Steel Co., for dental

casting application. They concluded that the alloy is promising because it is much easier to machine

than titanium and gives a clear smooth surface. Ti-Ni alloy wire produced by the Furukawa Electric Co.

showed ultra elasticity, which is a good for a material from which to develop a new functional device.

The titanium had an apatite coating


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A Ti alloy that is easy to machine is under study by Dr. Takada, and colleagues, at the Tohoku

University Dental School. They found that the addition of Ag or Cu improves two or three fold the

ability to machine the alloys.

Shu, and colleagues, at the Nagoya Institute of Industrial Technology under MITI, have shown that the

addition of Si, Ca and B in casting machines for dentists improves mechanical properties, for

example, by increasing tensile strength two fold.

Prof. Michio Ohta, at Kyushu University, has found from theoretical calculations that the optimum

composition for age-hardening alloys at mouth temperature is Au 50%, Cu 44% and Ga 6%.

Actually, an alloy with the above composition, plus Ir for finer crystal grains, has been commercialised

under the brand name Sofard.

Surface treatment of titanium using super critical water is under study by Dr. Ban, and colleagues,

at the Aichi Gakuin University Dental School, with the aim of increasing adhesion. The sheer bonding

strength of veneer resins to titanium was increased approximately 25% after super critical water

treatment.

5-5 Surface modifications

The National Industrial Research Institute of Nagoyahas succeeded in coating titanium alloy with

hydroxyapatite. They covered the alloy with titanium metal and then added hydroxyapatite particles

with a 30 mm diameter and heated up to 800-900C under pressure. They are going to proceed with

animal tests.

Professor Kokubo, of Kyoto University, in co-operation with Kobelco Co., is developing the formation

ofamorphous of graded sodium titanate layer on the surface of titanium in order to facilitate apatite

formation. It is a three-year study beginning in 1999 with 200 million Yen funding from the Japan

Science and Technology Foundation under the Science and Technology Agency.

The National Research Institute for Metals has a biometal research group called Center of Cell

Engineering headed by Dr. Hanawa. They focus on basic research on the interaction of cell and

metal:

-Changes in metal surfaces in vivo,

-Cell adhesion to an evaporated film of pure metal,

-Corrosion behaviour of amorphous alloy in a pseudo-bio environment,

-Electrochemical analysis of pure Titanium and SUS316L with culturing cells.


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Dr. Masao Yoshinari, and colleagues, of the Department of Dental Materials Science, Oral Health

Science Center, Tokyo Dental College, is studying surface modifications of titanium to suppress the

attachment of oral bacteria. They found that fluorine implantation on titanium polished surfaces

decreases the proliferation of bacteria.

Dr. Kawabe, and colleagues, of the Okayama University Biomaterial Lab., Faculty of Engineering, are

studying the control of apatite-forming ability on the titanium surface using an electrochemical

technique.

Dr. Shunsuke Fujibayashi, at the Graduate School of Medicine, Kyoto University, is studying the effect

ofsodium removal treatment on the bone bonding ability of bioactive titanium prepared with alkali

and heat treatment.

5-6 Other Metals

(1) Prof. Nakajima, at Osaka University, started the project called Development of advanced

medical equipment using porous metal supported by the government Millennium Project. It is a three

year project, from 20002002, at a total cost of USD 1.5 million. The project involves Yokohama

Medical University, Asahi Intec Co. as well as other science and technical universities. The Lotus-type

porous metal so far developed by the professor has uniform straight pores of 1-10 mm and a pore ratio

of 80%. They are aiming at making the porous metal with stainless steel, titanium, and shape memory

alloy to make guide wires, stents, artificial bone, and dental implants. According to Business &

Technology News, the market for guide wire and stents is USD 360 million with an annual growth rateof 23%, and the number of cardiovascular patients is 3.3 million. The market for artificial bone, joints,

and dental implants is USD 200 million with 14% annual growth with 3.2 million osteoporosis and

arthritis patients.

(2) Prof. Shinichi Nittas group, from Tohoku University, and Tokin Co. have jointly developed

artificial cardiac muscle with shape memory alloy to be attached on the outside of hearts. The

muscle is a Peltier effect element sandwiched by shape memory alloy plates. The size is 1cm*5cm with

0.5 cm of the thickness due to the Ni-Ti alloy. The power is supplied from outside the body by

electromagnetic induction. The group tested the effect using a goat.

6 Polymers and Lipids8

There were 40 presentations in the July 2000 Medical Polymer Symposiums. Many researchers are

working in this field for various applications such as drug delivery systems, gene delivery systems, and

scaffolds. Targeting and gene delivery is one of the most exciting challenges.


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6-1 MS Coat

MS Coat is the brand name of a desensitiser sold by the SUN MEDICAL CO. to relieve patients with

hypersensitive teeth. It was developed by the SUN MEDICAL, a subsidiary of Mitsui Chemical Co.,

and has been on the US market (Parkell Co. is the sales representative) since 1996 as Pain-free

desensitiser. It began sales in Japan in October 1999, through the Morita Co.

Its main component is a copolymer of methyl methacrylate and styrenesulfonic acid originally

developed by Prof. Nakabayashi, Institute of Biomaterials and Bioengineering Tokyo Medical and

Dental University.

A clinical trial done by the Tohoku University and the Tokyo Medical and Dental University showed

that 87% of patients with hypersensitive teeth achieved immediate relief after just one coating and 82%

remained pain-free even after one month. It consists of liquid-A: a MS Coat and liquid-B: calciumcompound.

6-2 MPC 9

A 2-metacryloyloxyethyl phosphorylcholine (MPC) polymer contains a phospholipid polar group

(phosphorylcholine group), the structural component of biomembrane; and a metacryloyl group, which

has polymerisation capability with many monomers. MPC polymer reacts extremely weakly with

biocomponents such as proteins and blood cells and shows a quite excellent thrombus resistance. Many

researchers, both domestic and abroad, are working on using MPC with blood cleaners, implant

sensors, blood pumps, blood circuits, artificial blood tubes, and drug carrier nanospheres.

MPC polymer was originally synthesised by Professor Nobuo Nakabayashi, at the Tokyo Medical and

Dental University, in 1978. Professor Kazuhiko Ishihara of the Department of Materials Engineering,

Faculty of Engineering, at the University of Tokyo has established volume production technology of

MPC funded by the government.

NOF Corporation (Nippon Yushi Co.) has completed a commercial plant for MPC, from monomer

through polymer, and anticipates various applications such as contact lens cleaning liquid, catheters,

and intraocular lens.

The combination of MPC with other materials such as polysulfone membranes and segmented

polyurethanes is one of the research themes of the above groups.

6-3 Photoreactive polymers

Dr. Yasuhide Nakayama, an expert in photoreactive polymers, at the National Cardiovascular Disease

Center Research Institute, has synthesised, in co-operation with Prof. Takehisa Matsuda, of Kyushu

University, photoreactive gelatin by introducing side chains to the benzophenon group, which generates


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6-5 Polymeric Micelles

Professor Kataoka, at Tokyo University, studies polymeric micelles and intelligent biomaterials. Block

copolymers that have both hydrophilic and hydrophobic polymer chains in a molecule form micelles, inwater, with hydrophilic chains outside and hydrophobic chains inside. The micelles are excellent drug

carriers, because they can hold drugs firmly in their inner cores and control the drug release by

changing the molecular structure of the inner cores. Since polyethyleneglycol (PEG) is used for the

hydrophilic chains, the immune system does not recognise the micelles as foreign materials. The

micelles also have an enhanced permeability and retention effect, useful for tumour targeting. Micelles

containing doxorubicin have been shown to be highly effective against tumours.13

Dr. Kataoka is also developing a glucose-responsive insulin release system using the equilibrium of

phenylboronic acid in aqueous solution. He has made a gel by introducing acrylamidephenylboronic

acid (AAPBA) to N-isopropylacrylamide (NIPAAm). The gel can release or retain insulin (on and off)

in accordance with the glucose concentration.14

He and Prof. T. Aida, from Tokyo University, are applying micelles to DDS for cancer, with a

porphyrin dye covered by dendrimer. Porphyrin purges active oxygen in response to light radiation and

attacks cancerous cells. The dendrimer is positively-charged, therefore, it tends to attach to the surface

of cancerous cells. Thus, the micelles have 28 times more capability of killing cancerous cells than

porphyrin alone. Kataoka is planning to cover the micelles with lipid particles, with the diameter of 50

nm, because he already showed that lipid particles gather efficiently around cancerous cells.

6-6 Nanoparticles and Microcapsules

(1) Mucosal peptide delivery systems are under development by Prof. Kawashima, and colleagues,

at the Gifu Pharmaceutical University. They have succeeded in developing lactide glycolide copolymer

(PLGA) nanospheres containing peptides. They have also succeeded in modifying the surface of the

nanosphere with chitosan, for mucosal adhesion. This is because mucosa has a negative charge and

chitosan is positively charged, thus the movement in the digestive tract is delayed. They have shown

the prolonged effect of calcitonin using rats.

They have also made powdered nanospheres for pulmonary delivery that coagulate when inhaled and

disperse in the lung and thus function as nanospheres. They have observed that the dextranconcentration in blood is extended by using the powdered nanospheres.

(2) Prof. Kikuchi, and colleagues, of the Nara Institute of Science and Technology, have succeeded

in developing unique spheres with silicon oxide on the surface of a liposome. The diameter of the

sphere is 200 nm. The silicon oxide layer is 5 nm. They named the spheres, Cerasome. There were

problems with adhesion or deformed shapes due to the soft surface of the liposomes. The siliconised

spheres will solve such shortcomings. The researchers are going to add bio-affinity to the surface.

(3) Prof. Takayama, and colleagues, of the Hoshi Pharmaceutical University, have developed a

neural network system to get the optimum mixing ratio of various materials in order to achieve a micro


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capsule of a certain diameter. For example, by mixing three materials, such as a component in

soybeans, cholesterol and oleic acid, to form a capsule suitable for drug delivery to a cancerous part of

the liver, 15 experiments will be enough to get a regression formula in order to determine the ratio for

the mixture. At present, as many as 1,000 experiments are needed to determine the optimum ratio.

(4) Dr. Ichijou, and colleagues, of the National Institute of Materials and Chemical Research, are

working on molecularly imprinted polymers. They have succeeded in synthesising polymer

microspheres that can discriminate for oxytocin.

6-7 Liposomes

Prof. Konno's group, at the Research Institute of Advanced Science and Technology, Osaka Prefecture

University, are studying functional liposomes modified with polymers. They have succeeded increating a temperature-sensitive liposome that releases drugs at a temperature slightly higher than body

temperature. The temperature response of the liposome copolymer depends on a kind of lipid, for

example, egg yellow phosphatidylcholine did not show remarkable drug release in temperatures higher

than the lower critical solution temperature (LCST). But adding dioleoilphosphatizylethanolamine

(DOPE) gave a very sharp temperature responsive liposome. They have synthesised copolymers, such

as NIPAM-AAM (poly-N isopropylacrylamide-Acrylamide) and NIPAM-N-APr (Acryloylpyrolidine),

that have LCST around 40 degrees C.

A cationic liposome modified with N-APr has also been synthesised. The affinity with negatively

charged membranes is controlled by heating.

They have also synthesised succinylated polyglycidol, which showed high membrane fusion ability. It

can be used for a delivery system into cells.

6-8 Polysilamine

Associate Professor Sachio Nagasaki, at the Science University of Tokyo, is studying polysilamine, anew heterotelechelic oligomer. It can be prepared through an anionic polyaddition reaction between

divinyldimethylsilane and 3,6-diazaoctane. It is a gel, with more flexibility than natural gum, but swells

and turns solid with a pH change from alkali to acid. Applications for artificial muscle, drug delivery

systems, and sensors are anticipated.


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6-9 Perfluorocarbonic acid

Dr. Azumi, and colleagues, of the Osaka National Research Institute, are studying the displacement

characteristics of gold plating on perfluorocarbonic acid membranes, which has been developed byAsahi Glass Corporation. The displacement occurs in response to an electric field. The degree of the

displacement depends on the type of counter ions. They have applied the technology to micro catheters

that bend in an electric field.

6-10 Shape Memory Gels

Dr. Takashi Miyazaki, and colleagues, of the Hokkaido University, are working with shape memory

polymer gels prepared from acrylic acid and stearyl acrylate. The shape memory effect takes place at

the transition between the amorphous state in the dry condition and the crystalline state, formed withthe addition of a certain amount of water. Their aim is to create a reliable valve.

6-11 Hollow fibres

Hollow fibres are used for dialysers, and artificial lungs.

Materials and the manufacturers of hollow fibres

Makers AsahiMedical Kuraray Nikkiso Teijin Terumo Toray Toyobo

Regenerate

Cellulose

* *

Cellulose

Acetate

* *

Polyacryl-

nitril

*

Polymethyl-

methacrylate

*

Ethylene

vinylalcohol

*

Polysulfone * * *

Polyester-

polymer alloy

*

Synthetic polymers are now more used in place of cellulose, with polysulfone as the most popular.


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7 Drug Delivery and Targeting

At the Controlled Release Society in Paris in 2000, Prof. Okano, from the Tokyo Women's Medical

University, was given the Founders Award for his research on temperature responsive polymers. Dr.

Okada, from the Takeda Pharmaceutical Co., and Dr. Toda, a former Takeda employee, now with

Ohtsuka Pharmaceutical Co., were also honoured with the Nagai Innovation Award for their invention

ofLeuprin and its success on the market. It is proof that Japan is world-class in both research and

commercialisation of DDS.


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7-1 Overall situation

Current status of fine particles used in DDSKinds Materials Size Characteristics Sustained

drugs

Exa

rese

Liposomes Lecithin, Cholesterol,

Charged substances

20nma

few mm

Suitable for localising,

targeting, slow release,

surface modification,

and activation of

microphage

water-soluble

& lipid-

soluble

Top

Mo

anti

Hem

Cat

Con

con

Lipid

microspheres andnanospheres

Lecithin, Soya oil,

Olive oil

0.2 mm

(ave.)

Lymph targeting,

Thermal sterilisation

lipid-soluble new

lipoamp

(50

Emulsion Synthetic surfactants,

Oily substances

Greater

than 0.1

mm

Used mainly for

embolisation and

absorption

enhancement

water-soluble

& lipid-

soluble

Art

cell

Ora

insu

PTH

Mixed micelles Synthetic surfactants,

Unsaturated fatty

acids

5 nm Enhancing absorption

in intestines

lipid-soluble

Polymer micelles Synthetic polymers 50200

nm

water-soluble

& lipid-

soluble

Adr

Cal

PA

cop


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7-2 Active targeting15

Prof. Kitajima, at the Medical School, Keio University, in co-operation with Associate Prof. Imoo, at

Okayama University, made ribonuclease combined with EGF by joining the genes ribonuclease and

EGF and introducing them into E. coli. Ribonuclease is an enzyme that destroys RNA and inhibits

protein synthesis, thus killing cancerous cells. Cancer cells have receptors that bind EGF, a cell growth

factor. The combination was effective in a mouse transplanted with cancer cells.

In addition to EGF, the transferrin receptors and galactose receptors of the liver cancer cells are under

study by Prof. Hashida, at Kyoto University and Prof. Kataoka, at Tokyo University.

Dr. Kataoka is also trying to bond recognition molecules to micelles, to provide both passive and active

functions.

7-3 Passive Targeting16

The EPR effect (enhanced permeability and retention effect) was discovered by Dr. H. Maeda. Through

this effect, polymers are selectively accumulated in cancer tissues compared with normal tissues.

SMANCS is a DDS drug utilising this effect.

Liposomes modified with polyethyleneglycol can reduce non-specific interactions and suppress uptake

into the reticuloendothelial system. Prof. Maruyama, at the Faculty of Pharmaceutical Science, Tokyo

University, reported a remarkable increase in residence time in the blood with the modified liposomes.

Polymer micelles that have been developed by Prof. Kataoka are now being tested in a DNA delivery

application.

7-4 Stimulus Responsive Materials

The following are not necessarily focusing on DDS but on intelligent materials.

(1) Prof. S. Aoshima, at Osaka University, is studying living cation polymerisation of vinylethers under

an additive base. For example, polyvinylether, with oxyethylene on side chains, is water-soluble at low

temperatures but shows phase-separation at a certain temperature (Tps). Tps is changeable by the kind

of side chains or random copolymerisation. When the polymer is mixed with polycarbonic acid, it

separated at below a certain pH (CpH) to form a complex.

(2) The New Energy and Industrial Technology Development Organisation (NEDO) started a five-year

research programme called Advanced Stimuli-Responsive Materials in 1996. The programme is

focused the intellectual environment responsive functions of biological systems used as industrial


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materials. The programme has the three targets: 1. Separation materials, 2. Control release materials, 3.

Actuating materials.

Research on control release materials is conducted at the Osaka National Research Institute in co-operation with Prof. Nagata, of Hokkaido University; Prof. Okano, of the Tokyo Womens Medical

University; Prof. Kataoka, of Tokyo University; Prof. Akashi, of Kagoshima University; Prof.

Kajiwara, of Kyoto Institute of Technology; and some companies.

Under the programme, Dr. Ueno, of the National Institute of Materials and Chemical Research, in co-

operation with Tokyo University and Chisso Corporation, is studying magnetic nano particles attached

with ligands or nucleonic acid. The particles can be easily recovered using magnets. They

copolymerised acryloylglycineamide and biotin monomer, and, just by mixing the copolymer with the

magnetic particles, they can get temperature responsive magnetic nano particles that aggregate at less

than 20 degrees C and disperse at a higher temperature. This technology has the potential to produce

molecular recognition and temperature responsive magnetic particles.

7-5 Electromagnetic Responsive Double Targeting17

Professor Takeshi Kobayashi, in Bioengineering at Nagoya University, is developing Electromagnetic

Responsive Double Targeting, in a combination of time targeting, controlled release by physical

stimulations from the outside, and space targeting, collecting medicines to a target area by use of

monoclonal antibodies and others.

Dr. Kobayashi is a developer of magnetic nano-spheres, which heat up in response to an alternating

magnetic field. He uses carboxy-methyl cellulose (CMC) and 25nm magnetite in a 4/1 weight ratio to

form extrusion needle-shaped magnetite 2-3 mm in diameter.

Dr. Wakabayashi of Nagoya University Medical School, has found a use for magnetic nano-spheres in

treatment of brain tumours. Liposomes containing magnetic nano-spheres were injected into rat

tumours and a 120Hz magnetic field was applied. The tumour disappeared through the heat generated

by the particles and the magnetic field. This was also successfully used in rabbits. A monoclonal

antibody that binds to tumours has been developed for brain tumours.

Magnetic nano-spheres are used for extracting target substances in blood. An antibody for insulin, for

example, is attached to the particles, then put into blood. Insulin bind to the particles. Insulin is thus

measured at a higher concentration. Glycosylated hemoglobin and fructosamine can also be measured.

This way diabetes can be diagnosed in 30-40 minutes with 50 ml of blood. The method is now being

marketed by a venture company called M-Bio.

7-6 Biomolecular Design for Biotargeting


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Biomolecular Design for Biotargeting was accepted by the Ministry of Education and Science from

1998 to 2000 as one of the Grant-in Aid Programmes for the Scientific Research on Priority Areas. The

goal of the programme is to review the targeting function of living cells or biomolecules and to

reconstruct it from chemical and engineering points of view. The programme consists of the followingfour research fields involving more than 70 researchers in universities:

1) Biomolecular recognition, represented by Professor Imanaka, Kyoto University

-Modifications of the molecular recognition capability of antibodies

-Signal transmitting function of antibodies

-Modifications of enzymes for molecule recognition analysis

2) Biomolecular architecture, represented by Professor Matusnaga, Tokyo Agriculture and

Engineering University

-Building a biomolecule architecture on the surface of magnetic nano-spheres and liposomessynthesis of a biomolecular architecture

Electromagnetic Responsive Double Targeting, described in the previous chapter, is one of the

outcomes of the programme.

3) Cell surface design, represented by Professor Tanaka, Kyoto University

-Culturing new functional micro organism cells

-Producing biovaccines with antigen presenting cells

Modifications of cell function by expressing chimera receptor molecules on the surface of animal

cells

Analysis of cell surface functions using artificial cells

4) Biotargeting, by Professor Kobayashi, Nagoya University

-Tissue and cell targeting of functional materials incorporated with biopolymers

Targeting of physiologically functional small molecules with stimulus-responsive polymers

Biomolecule targeting with an ultra-micro electrode device

7-7 Transdermal & Transmucosal Delivery

The Suguhara group has conducted a study of how drugs applied on the skin remain locally in dermis

or hypodermis or move to the entire body through blood vessels.18 The movement of a drug in skin is

rather complicated; therefore, the development of simple evaluation is indispensable.19

The direct transport of a drug from the nasal cavity to the cerebrospinal fluid was reported recently;

therefore, targeting the brain is possible.20


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How to obtain slow release of a drug and adhesion to mucosa is important, but on-off control of drug

release is also of interest. Polymer gels that change structure in response to temperature, pH, electric

fields, light, ultrasonic, and chemical substances are being studied by Prof. Okana's group.21

Studies on how to enhance the absorption of a drug in skin or mucosa have been carried out by the

Josai University group.22 Two Kyoto groups have studied absorption enhancing reagents.23

Electroporation is a relatively new method to enhance absorption.24 The Sugihara group reported that

pretreatment of a non-needle syringe increased the transdermal absorption of a drug.25 Phonophoresis

has also been used to enhance absorption.26

7-8 Company activities

(1) Leuprin, launched by the Takeda Pharmaceutical Co., is the most successful polymer nano-sphereDDS. It is sold in more than 10 countries, including the USA, with the annual sales of more than USD

1 billion. In 1989, Drs. Ogawa, Okada, Toda, and colleagues, at Takeda, succeeded in putting

leuprolerin acetate, an antagonist of the luteinising hormone-releasing hormone, in microspheres. The

microshperes, made of a copolymer of polylactic acid and glycol acid, to keep released leuprolerin for

more than a month after a one-time injection. It is currently used for the treatment of prostate cancer

and breast cancer.

(2) SMANCS, developed by Prof. Maeda, at the Medical School, Kumamoto University, and launched

by the Yamanouchi Pharmaceutical Co. in 1996, is the first and only polymer anticancer drug approved

by the Ministry of Health and Welfare. It is a styrene-maleic acid conjugate of neocarcinostatin. The

molecular weight of SMANCS itself is 16,000, but it bonds with albumin in the human body to behavelike a polymer with 80,000 MW. It gathers and accumulates around cancerous tissue, a phenomena

Prof. Maeda discovered in 1986 and called theEPR effect. SMANCS can be oil-philic by the butyl-

esterification of the carboxyl group of the styrene-maleic acid copolymer, thus, increasing the affinity

to lipiodol. Therefore, both the targeting and controlled release of lipiodol are achieved.

(3) A venture company called NanoCarrier Co., located in Chiba prefecture, has developed micelle-

type DDS that accumulate drugs near tumour tissues. The materials are molecules of hydrophilic

polyethyleneglycol (PEG) bonded with hydrophobic polyaminoacid. The particles have a diameter of

20200 nm. In an experiment using mice, the anticancer drug contained in the micelle accumulated

near the tumour tissue at a concentration seven times higher than the drug alone. They explain that

micelles normally cannot pass through blood vessels around healthy cells, but that passages in blood

vessels around cancerous cells are large enough for the micelles to go through.

(4) The DDS annual conference was held in July 2000 in Akita, Japan. The following information

related to biomaterial DDS was reported at the proceedings.

Nippon Shinyaku Co. is developing new drugs using lipid ultra fine particles (LNSTM) that contain

soybean oil and egg yellow lecithin and have a diameter of several dozen nano-metres. LNS does not

go to the liver or the spleen and has passive targeting capability.


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Welfide Co. is developing bromperidole, a drug for mental illness, contained in slow release polylactic

acid microspheres.

NOF Corporation (Nippon Yushi Co.) is ready to sell highly pure liposome for medical applications.

Ohtsuka Pharmaceutical Co. is studying liposome triggering cytokine production in human peripheral

blood cells.

8 Gene Delivery

8-1 Gene Therapy Protocols

Protocols for gene therapy that have been approved by the Advanced Medical TechnologyEvaluation Committee in the Ministry of Health and Welfare by the end of 2000.

No Date of

approval

Diseases Vectors/Gene Hospitals Number of

patients treated

1 1995 Feb. ADA

deficiency

Retro virus Hokkaido Univ. 1 (1995)

2 1997 May HIV infection Retro virus Kumamoto

Univ.

3 1998 Aug. Kidney cancer Retro virus Tokyo Univ.

Medical Institute

3

4 1998 Oct. Lung cancer Adenovirus Okayama Univ. 8

5 2000 May Esophagus

cancer

Adenovirus/

P53

Chiba Univ. 1

(19/12/2000)

6 2000 Feb. Breast cancer Retrovirus Japan

Foundation for

Cancer Research

7 With-drawn Liver cancer Adenovirus Tokyo Univ.

Medical Institute

8 2000 Jan. Malignant

gliomaCationic

Liposome

Nagoya Univ. 1

9 2000 Jan. Lung cancer Adenovirus Tokyo Jikei

Medical Univ.

1

10 2000 Jan. Lung cancer Adenovirus Tohoku Univ. 1

11 2000 Jun. Prostate cancer Adenovirus Okayama Univ.

12 2000 Jan Lung cancer Adenovirus Tokyo Univ.

Medical Institute

13 Under examin-

ation

Chronic

obstructive

arteriosclerosis

DNA alone Osaka Univ.


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As seen in the above table, Nagoya University is the only group that used non-virus vectors for clinicalstudies of gene therapy. Dr. Yoshida, and colleagues, have been developing gene therapy for malignant

glioma with the IFNb gene, using a cationic liposome as the vector. Liposomes are synthetic lipid

vesicles that can entrap drugs or genes within their aqueous compartment or lipid bilayer. Dr. Yoshida's

group conducted the first therapy in April 2000. They have also worked on entrapping recombinant

adenoviral vector in multi-layer liposomes to improve transduction efficiency and reduce virus

antigenicity. They have also succeeded, in co-operation with Aichi Medical University, in producing a

single chain antibody for type III mutant EGFR, which expressed in malignant glioma and not in

normal cells.

8-2 Research on non-virus vectors

Cationic liposomes and polycations are the two major materials used for gene vectors.

Prof. Akaike's group in the Department of Biomolecular Engineering in Tokyo Institute of Technology

and Prof. Hashida, in the Pharmaceutical Department in Kyoto University, are some of the major

research groups in the field.

Dr. Prof. Akaike's group, including Dr. Maruyama and Dr. Ishihara, has synthesised a graft

copolymer (comb type copolymer) with poly(L-lysine) as the backbone and hyaluronic acid side

chains. This polymer formed a complex with plasmid DNA. They showed with in-vivo experimentsthat the complex is specifically taken into the liver. They also made graft copolymer of poly(L-lysine)

and arabinogalactan, a kind of polysaccharide, and showed a similar targeting effect to the liver. They

are also studying the expression of graft copolymers in cells. Prof. Akaike is one of the leading

researchers of hybrid livers.

(1) Prof. Hashida, of Kyoto University, and Dr. Kawakami at Nagasaki University, have

synthesised a sugar-modified cationic liposome that created a complex with plasmid DNA. They used a

cholesterol derivative modified with galactose or mannose, both showing high gene expression in the

liver.

(2) Prof. Ogihara, Osaka University Hospital, has applied for permission to provide gene therapy

using catheters, in-stead of vectors, to bring hepatocyte growth factors to the target part of the inner

wall of the heart. Another group in Osaka University has already applied for permission to inject HGF

in cardiac muscle.

8-3 Polycations and Liposomes

(1) Prof. Mayumi and Prof. Nakagawa, of the Department of Pharmeutical Science at Osaka


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University, are studying membrane fusion liposomes, a kind of hybrid vector, with the membrane

fusion capability of a virus. They have also worked on the expression system in cytoplasm.

(2) Prof. Konno's group in Osaka Prefecture University, made a complex of succynilated polyglycidolbonded with transferrin and cationic liposome-plasmid conjugate. It was tested as a gene delivery

system for tumour cells and proved to have high expression efficiency, because plasmid DNA was

introduced into target cells through transferrin receptors.

(3) Increasing the gene-introducing efficiency of cationic liposomes to the level of viral vectors is

being studied by various research groups at the Tokyo University, Tokushima University, and Shizuoka

Prefecture University.

(4) Aomori Prefecture Green Bio Center has applied for permission to use a new gene vector,

positively charged polyornithine fused with plasmid, developed by Agriculture Research Center, to

create herbicide-resistant rice.

8-4 Polymer Micelles

Prof. Kataoka and his group are also applying micelles to non-viral vectors. Polyethylene glycol-poly

a, b-aspartic acid as anion block copolymer mixed with polyethylene glycol-poly L-lysine (PEG-PLL)

as cation block copolymer generates PIC with a diameter of dozens of nano-metres in a sharp diameter

distribution.

Micelles 80 nm in diameter were formed with plasmid DNA and PEG-PLL. Higher expression of

luciferase in cultured cells was obtained with the micelles, compared with conventional lipofectin. He

has also reported an environment-responsive micelle vector by reversible cross linking of the core

through disulfide bonds.

8-5 Fullerene and Cyclodextrin

(1) Prof. Nakamura, of the Science Faculty and Prof. Okayama, of the Medical Faculty in TokyoUniversity have been successful in applying for permission to use fullerene (C60) as a gene vector.

C60 is a cubic molecule with a 1 nano-metre diameter. Profs. Nakamura and Okayama bound DNA,

containing the luciferase gene on an expression plasmid, to positively charged C60, and added them to

a culture of monkeys kidney cells. Expression was achieved in 25% of the cells. Nakamura and

Okayama claim that C60 gradually enters into the nucleus. Thus, C60 can be a DDS. Together with a

pharmaceutical company, they have applied for a patent.


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(2) Prof. Kamigama, Pharmaceutical Faculty, Kumamoto University, is trying to use a conjugate of

cyclodextrin and dendrimer as a gene vector. The conjugate showed higher activity than conventional

LipofectinTM.

9 Tissue Engineering

Tissue engineering has been strongly supported by the government in the past several years and was

chosen as one of the focus fields of the Millennium Project supported by the Prime Minister. A national

centre of tissue engineering, from basic research to clinical development, is being built in Kobe.

Among the significant number of researchers in the field, Kyoto University is a pioneer in this field and

its research is considered world-class. Tissue engineering research in Japan is not lagging behind theUSA, but commercialisation or business applications are clearly behind. Only recently have some

venture companies been created.

According to J-TEC, a venture company for skin culture, the Japanese market potential for tissue

engineering will be USD 8 billion and the world market, USD 45 billion .

Three government-supported research programmes in the tissue engineering field are of note:

The Regenerative Medical Engineering Programme, supported by the Ministry of Education and

conducted by hundreds of university researchers---1996--2000 Hiroshima prefecture Tissue Engineering Project, organised by Professor Katsuoshi Yoshizato, of

Hiroshima University

3D Tissue Module Engineering, conducted by NAIR(National Institute for AdvancedInterdisciplinary Research), under the Ministry of Economics, Trade and Industry (A part of the

Millennium Project supported by the Prime Minister)

9-1 Regenerative Medical Engineering Programme by the Ministry of Education

The programme started in 1996 as a five-year programme with about USD 1 million annually. Theprogramme consists seven groups:

Tissue Engineering for Hard Tissues; Tissue Engineering for Soft Tissues; Biomaterials for Tissue

Engineering; Bioprocess Engineering of Functional Regeneration of Culture; Tissue Engineering for

Organ Regeneration, started in 1996; and, Function Regeneration, and Bio Tissue Engineering, started

in 1997 and 1998, respectively.


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readily adhere to them. Therefore, we transferred harvested cell sheets onto another cell type

monolayer and co-cultured them. The other method is a patterned co-culture utilising patterned graft of

PIPAAm by masked electron beam irradiation. One cell type is then cultured to confluency at 37'C.

Reducing the temperature to below 32 'C., the cells are only detached from these grafted patterns.Another cell type is then seeded over the same surface at 37'C. These subsequently seeded cells only

adhere to the now-exposed polymer-grafted domains. Initially, seeded cells remaining adherent on non-

patterned surfaces and cells added in the second seeding are then co-cultured at 37'C together in well-

ordered patterns. These two methods enabled long-term co-culture of primary hepatocytes with other

cell types. Hepatocytes co-cultured with endothelial cells maintained the differentiated functions such

as albumin synthesis for more than two months. These findings might be related to the morphological

resemblance to liver tubule structure in terms of the adjacent lining of hepatocytes and endothelial

cells.

(3)Bioprocess engineering of functional regeneration of culture

Prof. Ohshima, at Tsukuba University Medical School, is the project leader and the objective is to

develop an artificial liver.

They have developed the volume culture of liver cells: a packed-bed reactor using polyvinylformal as a

scaffold. Based on various other studies, they are currently trying to scale up reactors, create model

animals, and a mixed culture of foe

Biomaterial Research Japan

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