Silica Xerogel as an Implantable Carrier for Controlled Drug

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*Corresponding author. Tel.: #358-2-2727690; fax: #358-2-

2727273.

E-mail address: pirjo.kortesuo@orion." (P. Kortesuo)ଝPresented in part at the 23rd Annual Meeting of the Society for

Biomaterials, New Orleans, 1997.

Biomaterials 21 (2000) 193}198

Silica xerogel as an implantable carrier for controlled drugdelivery* evaluation of drug distribution and tissue e! ects

after implantationଝ

Pirjo Kortesuo*, Manja Ahola, Stefan Karlsson, Ilkka Kangasniemi,Antti Yli-Urpo, Juha Kiesvaara

Orion Corporation, Orion Pharma, Pharmaceutical Development Department, P.O. Box 425, FIN-20101, Turku, Finland

Institute of Dentistry, University of Turku, Lemminka( isenkatu 2, FIN-20520, Turku, Finland

Received 30 September 1998; accepted 15 July 1999

Abstract

The purpose of the present study was to examine controlled delivery of toremifene citrate from subcutaneously implanted silica

xerogel carrier and to evaluate silica xerogel related tissue e! ects after implantation. Toremifene citrate was incorporated into

hydrolyzed silica sol in a room temperature process. Toremifene citrate treated silica xerogel implants were tested both in vitro and

in vivo using healthy mice. Silica xerogel with tritium-labelled toremifene was implanted subcutaneously in mice for 42 d. To

determine the amount of tritiated toremifene remaining in the silica discs at the implantation site, the discs were excised periodically

and radioactivity measured. The amount of tritiated toremifene in the implant after 42 d was still about 16% and the amount of silica

xerogel about 25%. In a histopathological study silica xerogel did not show any tissue irritation at the site of the implantation.

A "brotic capsule was formed around the implant. No silica xerogel related histological changes in liver, kidney, lymph nodes and

uterus were observed during the implantation period. The silica xerogel discs showed a sustained release of toremifene citrate over

42 d. Histologically, toremifene-related changes in the uterus were also detectable at all studied time points. These "ndings suggestthat silica xerogel is a promising carrier material for implantable controlled drug delivery system. 1999 Elsevier Science Ltd.

All rights reserved.

Keywords: Sol}gel processed silica xerogel; Implantation study; Drug delivery; Toremifene citrate; Tissue distribution

1. Introduction

Sol}gel processed porous silica glasses have a con-

siderable potential as carriers for controlled drug release

[1}4]. In recent years, processing factors controlling thedegradation rate of the matrix in vitro have been exten-

sively studied [1,3]. However, in vivo degradation and

drug delivery of low-temperature processed silica xerogel

are not well documented.

Parenterally implanted sintered sol}gel produced

glasses containing 85% silica and also pure silica glasses

are biocompatible [5,6]. Bioactive glasses in soft tissue or

bone enhance "broblast or osteoblast formation result-

ing in increased collagen formation, which in turn bonds

bioactive glass implants to bone or soft tissue [5,7].

However, crystalline silicon dioxide in the form of aero-

sol is known to cause a rapid in#ux of in#ammatory cellsand increased collagen deposition in lungs and histologi-

cal changes in the pulmonary lymph nodes [8].

Toremifene citrate, a non-steroidal antiestrogenic com-

pound, which has antitumor activity in breast and en-

dometrial cancer, was used as model drug in this study

[9]. Local therapy of antiestrogens after hormone-depen-

dent breast cancer surgery could give a targeted and

long-lasting disease control while minimizing systemic

distribution.

In this study toremifene citrate/ H-toremifene was in-

corporated into the silica gel matrix during a room-tem-

perature gelation process and silica gel was dried toa constant weight at 403C without sintering at a higher

temperature. The objectives of this work were: (i) to

0142-9612/00/$- see front matter 1999 Elsevier Science Ltd. All rights reserved.

PII: S 0 1 42 - 9 6 1 2 (9 9 ) 0 0 1 48 - 9



evaluate the capability of silica xerogel to deliver tor-

emifene in a sustained manner, (ii) to study tissue distri-

bution of toremifene and (iii) to evaluate histological

e! ects after implantation of the silica xerogel itself on the

uterus, liver, kidneys, lymph nodes and at the implanta-

tion site.

2. Materials and methods

2.1. Preparation of toremifene citrate incorporated

silica gels

Silica gel was prepared by the hydrolysis and polycon-

densation of tetraethoxysilane (TEOS) with polyethylene

glycol (PEG, M

4600), water and acetic acid in a

mole ratio of TEOS : PEG : H

O : CH

COOH"

1.0 : 0.0012 : 14.2 : 0.5 at room temperature. Toremifene

citrate (Orion Corporation, Turku, Finland) was dis-solved into the silica solution after 1 h hydrolysis at the

concentration of 33 mg/g. Tritiated toremifene was syn-

thetized as described earlier [10]. The speci"c activity of

the material was 16 Ci/mmol. It was added to the silica

sol to produce a "nal radioactivity of 16 Ci/g. Solution

was pipetted into round moulds (100 l/mould) for poly-

condensation and aging at 403C for 18 h. The poly-

merized silica gels were soaked in water to leach out

residual organic matter within the gel and dried at 403C

to a constant weight. The silica xerogel implants were

round with the diameter approximately 4.7 mm and

height 0.9 mm weighing about 15 mg.

2.2. In vitro dissolution test

The dissolution pro"les (each data point is the mean of

2 values) of H-toremifene and silica from test specimens

were studied using the USP XXIII dissolution apparatus

II (paddle method, Sotax AT6, Basel, Switzerland) at

a constant temperature (373C). Simulated body #uid

(SBF, pH 7.4) containing 0.5% (m/v) sodiumdodecylsul-

phate was used as dissolution medium. SBF was pre-

pared by dissolving reagent-grade NaCl (136.8 mM

),NaHCO

(4.2 mM), KCl (3.0 mM), K

HPO;3H

O

(1.0 mM), MgCl;6H

O (1.5 mM), CaCl

;2H

O

(2.5 mM) and Na

SO

(0.5 mM) in distilled water. The

solution was bu! ered at pH 7.4 with tris(hy-

droxymethyl)aminomethane (50 mM) and hydrochloric

acid.

The volume of dissolution medium was 250 ml and the

rotation speed 50 rpm. At each sample interval a 2 ml

sample was withdrawn and replaced with fresh medium.

To study the amount of tritiated toremifene released

from the matrix, samples of 0.5 ml were mixed with 10 ml

of scintillation liquid (Ready Safe2+, Beckmann, CA,USA) and measured by scintillation counting (Model

81 000, LKB-Wallac, Turku, Finland).

Degradation of silica xerogel matrix was determined

by measuring dissolved Si(OH)

spectrophotometrically

as a molybden blue complex at 820 nm [11].

2.3. Animal experiments

The animals were housed according to the recommen-

dations in DHEW pub. (NIH) 85-23 entitled &Guide for

the care and use of laboratory animals'. Thirty female

mice (C57Bl, Denmark) with an average weight of about

19.6 g (SD 1.2 g) received implants subcutaneously on the

back on either side of the backbone with tritium-labelled

toremifene loaded silica implants (a total of 2 implants

were placed in each mouse) and 30 mice with untreated

silica implants. The H-toremifene dose was about

80 Ci/kg (0.8 Ci/implant), toremifene citrate 350 mg/

kg (approx. 3.4 mg/implant) and silica xerogel about

1.53 g/kg body weight.

2.4. Histology

The animals were killed at 7, 14, 21, 28, 35 and 42 d

(5 mice per time period) after implantation. The silica

xerogel implants on the left side of the backbone were

removed together with the surrounding tissue and "xed

in 70% ethanol and embedded in PMMA (Technovit).

Sections of 20 m were stained with toluidine blue. Sam-

ples of liver, kidney, mesenterial lymph node and uterus

were "xed in bu! ered formaldehyde and embedded in

para$n. Sections of 6 m were stained with hema-toxylin}eosin. Samples were evaluated using light

microscopy. The samples of the implantation site were

studied from all interim kills but liver, kidney, lymph

node and uterus only from the animals treated 7, 28 and

42 d.

2.5. Drug release and silica xerogel erosion studies

The mice were killed, whereupon the silica xerogel

implants on the right side of the backbone were cut o!

from the surrounding "brous capsule and dried at room

temperature in a desiccator for 24 h. The percentage of implant remaining at each time point was calculated. To

determine the amount of toremifene remaining in the

implants, the dried implants were dissolved in 0.1 N

NaOH solution and the activity was measured in liquid

scintillation counter.

2.6. Organ radioactivity counting

After killing the mice, the tissue samples taken from

liver, kidney, uterus, lung, blood and application area

were burned in an oxidizer (Junitek, Kaarina, Finland)and the radioactivity was measured by scintillation

counting.

194 P. Kortesuo et al. / Biomaterials 21 (2000) 193}198



Fig. 1. In vitro release of H-toremifene and silica from silica xerogel

matrix.

Fig. 2. Percentage of remaining silica xerogel implant and H-tor-emifene radioactivity at di! erent time points in vivo.

Fig. 3. Biodistribution of H-toremifene radioactivity at di! erent time

points in some of the organs in mice.

3. Results

3.1. In vitro and in vivo release of tritium-labelled

toremifene and erosion of silica xerogel matrix

About 90% of the loaded tritium-labelled toremifene

was released after 99 h in vitro. The amount of silica

released in 99 h is equivalent to about 65% weight loss of silica xerogel matrix (Fig. 1).

Weight loss of silica gel matrix in vivo was about

75 wt% during 42 d. The erosion rate was fast during the

"rst 28 d and then decreased (Fig. 2). By an estimate from

the dissolution pro"le, the silica xerogel implant weigh-

ing about 15 mg is totally absorbed after 60}70 d.

The silica xerogel discs showed sustained release of

toremifene during the test period. About 40% of loaded

toremifene eluted during the "rst week. After that the

release of toremifene slowed down and 50% was released

in 3 weeks. The amount of

H-toremifene remaining inthe implant after 42 d was still about 16% (Fig. 2).

The radioactivity of tritiated toremifene was detected

upto 42 d in the liver, kidneys, and lung. The radioactiv-

ity reached a maximum value at 14 d and then remained

relatively constant until the end of the implantation test.

Radioactivity was not found in blood or uterus (Fig. 3).

Toremifene-induced uterine glandular hyperplasia, the

characteristic hormonal change seen in mice treated with

this drug, was observed at all studied time points.

3.2. E w ects after implantation

The local e! ect of the silica xerogel implant was re-

stricted to the close vicinity of the implant. A well-

organized "brous capsule was formed around the im-

plant from 14 d time point onward (Fig. 4A). The implant

did not cause necrosis and the in#ammatory response

observed was mainly due to a minor in"ltration of mac-

rophages at time points 14 d and later. Acute in#amma-

tion was observed only at the 7-d point. Phagocytic

macrophages were present close to the implant at 14 d or

later (Fig. 4B). No accumulation of dissolved material

occurred.

Toremifene released from the implant caused tissueirritation, necrosis and an unfavorable environment for

phagocytic macrophages causing an accumulation of

P. Kortesuo et al. / Biomaterials 21 (2000) 193}198 195



Fig. 4. (A) An untreated implant (S) in a mouse 35 d after implantation. The implant is surrounded by a "brotic capsule (arrow, original magni"cation

16;). (B) The surface (S) of an untreated implant 21 d after implantation. The "brous capsule (arrow) is well formed and phagocytic macrophages(open arrowheads) are abundant close to the surface (original magni"cation 100;).

dissolved silica inside the "brotic capsule. These "ndings

were present from the 14 d point onward.

No extensive silica xerogel related histological changes

could be observed in the liver, lymph nodes or kidney at

the 1.5 g/kg dose.

4. Discussion

The silica xerogel matrix gave sustained release of

toremifene citrate for more than six weeks. This was

con"rmed by erosion/release studies, activity measure-

ments of tritiated toremifene in tissue samples and by

histological studies. Histologically, toremifene-related

changes in the uterus were detectable at all studied time

points. About 40% of loaded toremifene was eluted from

silica xerogel matrix during the "rst week. After that the

release rate slowed down and about 16% of the loaded

amount was still remaining after 42 d. Therefore, radio-

activity in tissue samples reached a maximum valueat 14 d and remained relatively constant in the liver,

lung and kidneys throughout the test period. It was




demonstrated earlier that as a lipophilic agent, tritium-

labelled toremifene was distributed throughout the body

with highest radioactivity in the liver and lung [10].

Toremifene citrate is metabolized without "rst-pass

metabolism. After enterohepatic circulation, the metab-

olites are excreted mainly in feces. In tritiated toremifene

the radioactivity is not eliminated within the metabolic

pathways [10]. Compared to sintered silica xerogels(7003C) where a silica xerogel matrix was impregnated

with toremifene citrate, incorporation of drug molecules

into the non-sintered silica xerogel matrix retarded the

dissolution of toremifene citrate both in vitro and in vivo.

However, a non-sintered silica xerogel matrix degraded

faster in vivo than sintered silica gel in an earlier in vivo

study [12]. It is known from literature that sintering

decreases the dissolution rate of silica [13].

The in vitro toremifene citrate release was faster than

in tissue. About 90% of the loaded amount was released

after 99 h and the amount of remaining silica was about35% at the same time point. Weight loss of silica xerogel

matrix in vivo after 42 d was about 75%. The results

show that the in vitro release pro"le does not correlate to

the in vivo pro"le on a real time basis. The origin of the

di! erence of the behavior between in vitro and in vivo

data is probably due to the di! erence in the surface area

of the implant to the volume ratio of the solution at

subcutaneous implantation site and in dissolution study.

However, the observed release pro"les suggest that ero-

sion of silica xerogel controls the release of toremifene

citrate from the matrix.

The untreated silica xerogel implant did not causeirritation of the surrounding tissue. Local e! ects at the

implantation site are considered as a normal wound-

healing process. Acute in#ammation was observed only

at the 7-d point probably as a postoperative response.

Implantation of foreign material caused acute in#amma-

tion of relatively short duration. The "nal stage of heal-

ing response was the development of a "brous capsule at

the 14-d point around the implant, which typically has

been considered a sign of biocompatibility of the material

[14,15]. Sol}gel produced glasses also appeared non-

toxic in in vitro experiments showing normal growth of

"broblasts [6]. A "brous capsule may retard the release

of drug from implanted material and distribution to

other organs [14]. However, in this study toremifene-

related changes in the uterus were observed at all time

points.

Toremifene-treated implants caused irritation at the

implantation site throughout the study period. The dif-

ference in tissue reactions between untreated silica

xerogel and with H-toremifene treated implant may be

due to irritating e! ect of toremifene, which causes an

unfavorable environment for phagocytic macrophages.

In earlier studies toremifene has proved to have an ir-ritating e! ect following intraperitoneal administration

(unpublished result). As an antiestrogenic agent, tor-

emifene may also stimulate "broblast growth by TGF-

or radioactivity may produce tissue in#ammation [16].

Silica xerogel dose of 1.5 g/kg did not produce any

histological changes in liver, kidney, lymph nodes and

uterus during the test period (42 d). However, earlier

studies claim that silica has several biological e! ects,

which a! ect the immune system particularly [17]. Espe-

cially crystalline quartz (silicon oxide) is known to becytotoxic in macrophages and following chronic inhala-

tion, quartz dust causes increased granulation tissue in

the pulmonary or tracheobronchial lymph nodes of rats

[17}19]. The destructive e! ect of crystalline quartz is

probably due to the property of the crystalline form to

disrupt various biological membranes. However, these

"ndings were not observed with amorphous sol}gel pro-

duced silica xerogel.

In conclusion, silica xerogel gave sustained release of

tritium-labelled toremifene for six weeks. The release of

toremifene was controlled by the bioerosion of the silicaxerogel matrix. No silica xerogel related changes in liver,

kidney, uterus or lymph nodes were observed even

though the applied amount of silica xerogel was high

(1.5 g/kg body weight). Furthermore, silica xerogel was

biocompatible with the surrounding tissue. Thus silica

xerogel seems to be promising carrier material for im-

plantable controlled drug delivery system.

References

[1] Bo K ttcher H, Kallies K-H, Haufe H. Model investigations of con-trolled release of bioactive compounds from thin metal oxide

layers. J Sol}Gel Sci Technol 1997;8:651}4.

[2] Nicoll SB, Radin S, Santos EM, Tuan RS, Ducheyne P. In vitro

release kinetics of biologically active transforming growth factor-

1 from a novel porous glass carrier. Biomaterials 1997;18:853}9.

[3] Tan BH, Santos EM, Ducheyne P. Ultramicroscopic pore size

and porosity of xerogels for controlled release of biological mol-

ecules. Fifth World Biomaterials Congress, vol. 161, 1996.

[4] Unger K, Rupprecht H, Valentin B, Kircher W. The use of porous

and surface modi"ed silicas as drug delivery and stabilizing

agents. Drug Dev Ind Pharm 1983;9:69}91.

[5] Wilson J, Pigott GH, Schoen FJ, Hench LL. Toxicology and

biocompatibility of bioglasses. J Biomed Mater Res 1981;

15:805}17.

[6] Palumbo G, Avigliano L, Strukul G, Pinna F, del Principe D,

Angelo ID, Annicchiarico-Petruzzelli M, Locardi B, Rosato N.

Fibroblast growth and polymorphonuclear granulocyte activa-

tion in the presence of a new biologically active sol}gel glass.

J Mater Sci Mater Med 1997;8:417}21.

[7] Klein CPAT, Li P, de Blieck-Hogervorst JMA, de Groot K. E! ect

of sintering temperature on silica gels and their bone bonding

ability. Biomaterials 1995;16:715}9.

[8] Absher MP, Trombley L, Hemenway DR, Mickey RM, Leslie

KO. Biphasic cellular and tissue response of rat lungs after eight-

day aerosol exposure to the silicon dioxide cristobalite. Am

J Pathol 1989;134:1243}51.

[9] Kallio S, Kangas L, Blanco G, Johansson R, Karjalainen A,

Perila K M, Piippo I, Sundquist H, So Kdervall M, Toivola R. A new

triphenylethylene compound, Fc-1157a. I. Hormonal e! ects. Can-

cer Chemother Pharmacol 1986;17:103}8.

P. Kortesuo et al. / Biomaterials 21 (2000) 193}198 197



[10] Kangas L, Haaparanta M, Paul R, Dirk R, Sipila K H. Biodistribu-

tion and scintigraphy of C-toremifene in rats bearing DMBA-

induced mammary carcinoma. Pharmacol Toxicol 1989;64:373}7.

[11] Koch OG, Koch-Dedic GA, Handbuch der Spurenanalyse.

Berlin: Springer, 1974. p. 1105.

[12] Kortesuo P, Ahola M, Karlsson S, Kangasniemi I, Kiesvaara J,

Yli-Urpo A. Sol}gel-processed sintered silica xerogel as a carrier

in controlled drug delivery. J Biomed Mater Res 1999;44:162}7.

[13] Li P, Ohtsuki C, Kokubo T, Nakanishi K, Soga N. Apatiteformation induced by silica gel in a simulated body #uid. J Am

Ceram Soc 1992;75:2094}7.

[14] Anderson JM, Niven H, Pelagalli J, Olano! LS, Jones RD. The role

of the "brous capsule in the function of implanted drug-polymer

sustained release systems. J Biomed Mater Res 1981;15: 889}902.

[15] Anderson JM. In vivo biocompatibility of implantable deliv-

ery systems and biomaterials. Eur J Pharm Biopharm 1994;

40:1}8.

[16] Border WA, Noble NA. Targeting of TGF- for treatment of

disease. Nature Med 1995;1:1000}1.

[17] Burns CA, Zarkower A, Ferguson FG. Murine immunological

and histological changes in response to chronic silica exposure.

Environ Res 1980;21:298}307.

[18] Allison AC, Harrington JS, Bibeck M. An examination of thecytotoxic e! ects of silica on macrophages. J Exp Med 1966;124:

141}61.

[19] Gopinath C, Prentice DE, Lewis DJ. Atlas of experimental tox-

icological pathology. Norwell, MA, USA: MTP press, 1987.


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Silica Xerogel as an Implantable Carrier for Controlled Drug