CA1163681A - Radio frequency induced hyperthermia for tumor therapy - Google Patents

Abstract

Abstract of the Disclosure The instant invention is directed to a noninvasive tumor treatment modality which results in a reduction of tumor mass and may lead to complete eradication of a tumor.
The inventive method comprises localized magnetically-couple. RF-induced hyperthermia mediated by a material which is non-toxic to and, preferably, compatible with animal tissue and has incorporated therewithin iron-con-taining crystals of such size, amount, composition, and magnetic properties to impart a coercive force of at least 200 oersteds to the material, and wherein the RF magnetic field has a frequency not in excess of about 10 kilohertz.

Description

~orre~ uaerer-Panzarlno 18-5-12 -`

11 ~3 ~81 RADIO FREQUENCY INDUCED HYPE~THERMIA FOR TUMOR T~ERAPY

Bac~ground of the Invention It nas been recognized that, when heat is applied to areas of animal tissue containing both normal and malignant cells sufficient to raise the temperature of such areas to the range of 41-44C, a preerential destruction of the malignant cells occur~. (Normal animal tissue is destroyed at a temperature of about 48C.) Examinations of tumors subjected to such heat treatments utilizing light microscope and electron microscope techni~ues ha~e revealed that the tumors undergo specific destruction with no substantial damage to adjacent normal cells such as fibroblasts and endothelial cells. The initial result of hyperthermia applied to solid in ivo tumor~ is the rapid increase of lysosomal enzyme activity in the cytoplasm of malignant cells with concomitant inhibition of respiratory metabolism.
Significantly, a simultaneous depression of anaerobic glycolysis does not take place in tha malignant cells, thereby promoting the accumulation of lactic acid first in the intracellular spaces and subsequently in the extracellular spaces. Inasmuch as most solid tumors exhibit slow exchange between intracsllular fluid and blood, and this circumstance is particularly true in the central tumor regions, acidic conditions become predominant within the tumor during hyper-thermia. This increasa in acidity leads di;rectly to enhancad lysosomal enzymatic activity (p~ maxima 5-5.5). The non-malignant cells surrounding the tumor sustain only minor and reversibl~ damage. Both malignant and normal tissue demonstrate a rapid and marked inhibition of RNA synthesis which is subs~quently followed by the partial inhibition of DNA and ~, .

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protein synthesis. A transitory effect on cell prolifera-tion has also been obser~ed. Nevertheless, those adverse effects are customarily eclipsed by the highly desirable rapid and ~ronounced lysosomal destruction occurring pre-ferentially in the malignant ~ells. The biochemical lesion(s) affecting both RNA and DNA metabolism and the ce~ls' ~bility to divide appear to be transient and are not believed to be the primary cause of hyperthermia-induced destruction. It has been postulated that a signi~icant factor in normal cell survival resides in the anatomical location of those cells, i.e., the normal cells axe located near the periphery of the tumor and are closely related to the blood ~essels, thereby m;nimizing the buildup of acidity in their immediate environ-ment, Hyperthermia has been found to interact synergistically with ionizing radiation treatment and chemotherapy, a factor which augments its clinical utility as an a~ticancer treat-ment modality.
The major obstacle impeding the widespread clinical utilization of hyperthermia in treating carcinomata has been the inability to deliver localized hyperthermia. Thus, early experimentation involving exposing an entire body to diathermy at temperatures of about 41~C had evidenced a temporary regxession of tumors, but shortly after the treat-ment the tumors began to grow rapidly again, Attempts have been made to localize heating in the tu~or-containing area with a minimum deleterious effect upon adjacent normal tis3ue through the use of ~;uch means as electromagnetic fields, e.g., lasers and microwaves, and radio frequency (~ ~ induced magnetic fields. The latter has been the subject of several publications, for example:

"Selective Inductive ~eating of Lymph Nodes", Gilchrist et

-2-11 ~3~ 8~

al., Annals of Surgery, 146, No. 4, pages 596-606, September, 1957; "Controlled Radio-Fre~uency Generator for Production of Localized Heat in Intact Animal", l~edal et al., American Medical Association Archives of Surger~, 79, pages 83-87, September, 1959; and "Selective Heating and Cooling of Tissue in Cancer Chemotherapy", Shingleton et al., Annals of Surgery, 156, ~o. 3, pages 408-416.
Those publications described the implantation of powdered magnetic materials, specifica}ly a magnetic form of iron oxide statedly defined as Fe2O3, into tissue. These particles became heated as a result of the coupling to the magnetic field through their dielectric and hysteresis loss.
Those s.tudies. utilized magnetic fields at radio frequencies between about 0.12-2 megahertz..
~ lthough initial experiments demonstrated that this method for localizing induction heating was operable in destroying metastases, two factoxs militated against this method being accepted as a useful modality for treating carcinomata. First, the magnetic form of iron oxide is insoluble in body fluids and in substantial concentrations may be toxic to and/or re;ected by the body, and, second, the normal tissue surrounding the tumor became too hot during the heating operation and was subject to necrosis.
Thi- latter effect was due to dielectric heating, i.e., heating resulting from ionic conducti~ity of body tissue and f.luids.
It has been recognized that more disparate heating between the suscepted region and the surrounding region would oc.cur if the excitation field were of lower frequency.
The heating of normal tissue takes place through dielectric heating which is a ~unction of the field of frequency squared _3_ 1~ 63~

or even higher, depending upon the loss tangent, whereas magnetic hysteresis heating ~aries linearly with field fraquency. Up to the presen~ time, however, there ha~ been no magnetic material with the necessary chemical, mechanical, and magnetic properties to be useful in contact with animal tissue while permitting the required heating to occur at reduced field frequencies.

Ob ective of the Tnvention The principal objective of this invention is to provide means for loc~lly heating tumors in animal tissue to tempera-tures within the range of about 41-50C, depending upon the time of exposure, such that the malignant cells are prefer-entially destroyed with essentially no damage or toxic effect upon adjacent normal tissue.

Summary of the Invention We have discovered that this o~jective can be achieved through localized magnetically-coupled, RF-induced hyper-thermia mediated by a material which is non-toxic to and, preferably, inert to or compatible with animal tissue and which has incorporated therewithin magnetic field suscepting cry~tals of certain size, composition, concentration, and ma~netic properties. A magnetic field is utilized having a sufficiently low frequéncy that dielectric heating effects are reduced to a negligible level. The magnetic properties are such as to maximize the hysteresis loss, i.e., the material exhibits high magnetization, high coerci~e force, and hi~h loop squaseness, each of those characteristics contributing to hysteresis heating. Those magnetic properties are also consistent with the practical available field induced within simple induction coil configurations. The frequency of the magnetic field is maintained at a suffi-ciently low level that essentially only magnetic hysteresis heating can occur. Hence, the frequency of the magnetic field will generally be in the range of 10 kilohertz or below. The dielectric loss of animal tissue and fluids is small in this low frequency range thereby minimizing heating in the absence of a magnetic susceptor. The in~enti~e materials disclosed herein promote effective magnetic hysteresis heating with no noticeable body rejection. This enhanced heating effect is due to the use of high concentrations of matri~ material, this matrix material being essentially free from any toxic or inflammatory effect upon the animal body.
~ence, the magnetic component i5 encapsulated in a matrix which is inert to or biocompatible with the animal body.
Iron-co~taining crystals ha~e been determined to be the most desirable magnetic field susceptor material. Certain organic plas~ics, e.g., TEFLO~ FTP, ha~e the necessary inertness and non-toxicity for useful matrix materials.
Howe~er, ceramic compositions selected from the group of glasses, glass-ceramics, and sintered ceramics which are non-toxic to and, preferably, compatible with animal tissue, and including in their struc~ure magnetic crystals of iron-containing compounds, ha~e been determined to constitute the mos~ desira~le target materials. It will be recognized that th~ efectiveness and e~ficiency of the materials are dependent upon their ability to translate magnetic energy into thermal energy, this ability being related to such factors as crystal type and concentration, the presence of precrystalline or semi-amorphous regions, and the like.
Consequently, it will be apparent that bodies containing 1~63~

substantial quantities of ferrimagnetic crystals will be preferred. In general, the magnetic iron-containing crystals will consist of magnetite ~e304) or a solid solution ferrite.
~o~ever, any material, glass or crystalline, su~tably encapsu-lated, having the required magnetic hysteresis response in low frequency RF fields will be opera~le. Moreover, inasmuch as the coercive force exhibited by the crystal phase varies with the size of the crystals, laboratory experience has indicated that the crystals should have a diameter in excess of 500~ (to exceed superparamagnetic size~ and, preferably, at least 10,000~ ~1 micron) to produce domain w~ll motion.

Thus in a broad aspect the present invention provides a ceramic selected from the group consisting of glass, glass-ceramic, and sintered ceramic suitab~e for inducing localized heating in the presence of a radio frequency magnetic field, said field having a frequency sufficiently low that essentially only magnetic hysteresis heating can occur, which has incorpor-ated therewithin magnetic iron-containing crystals of such size, composition, concentration, and magnetic properties to impart a coercive force of at least 200 oersteds to said material.
In a preferred embodiment the ceramic is a glass having a base composition selected from the group of phos-phates, silicates, and borates. Also preferably the iron containing crystals consist essentially of magnetite or a ferrite, with most preferred ferrite b~ing selected from the group of lithium, cobalt, nickel, manganese, and barium ferrite.

~3~8i Customarily, the animal tissue will be heated to temperatures within the range of about 41-44C inasmuch as those temperatures will cause necrosis of tumor tissue. In contrast, normal animal tissue is not destroyed until temperatures of about 48C are reached. It has been found~
however, that brief exposures to temperatures up to about 50C can be tolerated with Yery little destruction of noxmal tissue. Such higher temperatures quickly destroy tumor cells so the time of the treatment can be signifi-cantly reduced. Hence, a series or pulses of RF magneticenergy can be utilized; the time of each pulse effective to induce necrosis of tumor tissue with virtually no effect upon normal tissue can be determined empirically. The concentration of ceramic necessary to induce the desired heating effect can also be determined empirically. Thus, the upper temperature of heatlng can be controllably limited by suitably selectlng the ceramic and regulating the quantity ~hereof administered.

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Glasses and glass-ceramics of di~erse base constituents, e.g. r silicates, aluminosilicates, borosilicatesJ borates, and phosphates, and containing iron oxide in significant amounts are ~nown to the art. When batch materials for such glasses are melted under oxidizing or neutral conditions, the resulting glasses ca~ exhibit magnetic beha~ior, the magnitude of the beha~ior being a function of glass com-position, annealing schedule, presence of a minor amount of magnetic crystallization, etc. When glass melts are appro-priately quenched or glass bodies subsequently exposed tothe proper heat treatment, minute crystals structurally similar to magnetite can be developed and/or caused to grow in size within the gla~sy matrix and the ferromagnetic behavior then evidenced by the bodies is substantially enhanced. Glass-ceramic bodies are generally highly crystal-line, i.e., greater than 50% by volume crystalline. The following patents are illustrative of such products.
United States Patent No. 3,193,503 discloses the pro-ducti~n of glass-ceramic articles consisting essentially, expressed in weight percent on the oxide basis, of 16-50~
~gO, 37-60~ Fe2O3, 20-45% SiO2, and 0-15~ o~ mineralizers or nucleants Quch as CaF2, CoO, NiO, V2O5, ~oO3, and ThO2. The resultant articles were termed "magnetic ceramic ferrites"
but no crystallization identiication data were supplied.
United States Patent ~o. 3,594,360 describes the manufacture of glas-s-ceramics demonstrating ferrimagnetic properties. The compositions operable therefor consisted essentially, expressed in parts by weight on the oxide basis, of 35-55 F~2O3, 5-15 Li2O, 10-50 SiO2, and 1-15 ZnO.
The predominant crystal phase was stated to comprise a lithium ferrite.

1~ 63~ ~1 United States Patent No. 3,492,237 discussed glass-ceramic bodies having compositions within the Li20-Na20-A~2O3-Fe2O3-SiO2 system wher~in lithium ferrite is a primary crystal phase. The operable formulations have a mole ratio of SiO2:Na2O:A12O3 of 11-13:3-4:4-1 with 1-10 moles each of Fe2O3 and Li2O per mole o~ A1~03.
United States Patent No. 4,140,645 reports glasses and glass-ceramics contalning crystals o~ Fe304 with, optionally, a transition metal ferrite., for example, cobalt ferrite and nickel ferrite. The operable glass compositions were categorized into two groups, expressed in terms of weight percent on the oxide basis:
(a) 2-1.0% Na2O and~or K2O, 5-2~% B2O3, 15-40% FeO, 0-32% A12O3, and 35-6~% SiO2; and (b) 1.5-6% 1i20, 10-40% FeO, 10-20% A12O3, 45-66% SiO2, 0-5% TiO2 and/or ZrO2, and 0-5% B2O3, at least 1~ B2O3 being requir.ed when the proportion of FeO is less than 15%.
Likewise, the operable glass-ceramic compositions were formulated from two groups, expressed in terms of weight percent on the oxide basis:
ta) 2-10~ Na2O and/or K2O, 5-20% B2O3, 15-40% FeO, 15-32% A12O3, and.35-50% SiO2; and (b) 1.5-6% Li2O, 10-q.0% FeO, 10-20% A12O3, 45-66%
SiO2, 0.-5% T.iO2 and~or ZrO2r and 0-5% B203, at least 1% B203 being re~uired when the proportion of FeO is less than 15~.
Both the base glass and glass-ceramic compositions spontaneously precipitate Fe3O4 when the molten batches are cooled to a glass body. Subsequent heat treatment of the glass bodies g.i~s rise to the in situ growth of silicate crystals, e.g., muliite, beta-quartz solid solution, and beta-spodumene solid solution, yielding a highly-crystalline ., .

~ 3~#1 glass-ceramic body. The Fe304 crystals can experience some grain growth during that heat treatment.
It will be appreciated that, as a matter of convenience, the above patents report the total iron oxide content of the cited materials, customarily present as a combination of FeO
and FezQ3, as either "FeO" or "~e2o3 n . ~ence, in the interest of simplicity and because analysis of the individual proportions of FeO and Fe2O3 is tedious and knowledge of the precise content of each is unnecessary to the operation of the respective inventions, the full amount of the iron oxide present was expressed as either "FeO" or "Fe2O3".
Each of the above patents was directed to base glasses in the silicate system. Such compositions are operable in the present invention. Ho~ever, whereas there have been publications of silicate-based glasses suitable for bonding to bone or other living tissue, e.g., United States Patent No. 4,159,358 describing glasses consisting essentially, expressed in terms of weight percent on the oxide basis, of 40-60% SiO2, 10-32~ Na2O, 10-32~ CaO, 0-18~ CaF2, 0-2a~ B2O3, and 0-12~ P2O5, it has long ~een recognized that, in general, phosphate-based glasses are more compatible with living tissue than ar~ silicat~-based compositions. Conse~uently, the pre~erred base glasses, glass-ceramics, and/or ~intered ceramics of the present invention ha~e compositions founded in the phosphate composition system.
The desired locali2ed hyperthermia can be achie~ed through numerous ways. For example, an aqueous dispersion of the target c~ramic material in ~ery fineIy-divided form can be injected directly into a tumor and~or into normal tissue immediately a~jacent to the tumor. Subsequent exposure 1~ ~3~ 8~

by the defined RF induced magnetic field causes the target material to be heated.
In another embodiment, an aqueous dispersion of the powd~red ceramic material is injected via intravenous-or arterial routes at a site near or distal to the tumor.
Blood flow acts to transport the ceramic to the site of the tumor. Assistance in localizing the injected magnetic material utilizing this method can be found by guiding the passage with a magnet.
In the case of surgical, exposure of a tumor, the target material can be injected into or applied to the outside of the tumor. Hyperthermia will be induced through magnetic field induction heating before and~or after the incision has been clos~ed. Thus, the treatment can consist of a series of exposures. The ceramic will desirably be inert or else harmlessly degraded by body fluids, the degra-dation occurring slowly enough to permit the ceramic to be at the tumor site for a substantial period of time. ~ence, a succession of individual treatments with RF fields to secure localized heating can be conducted with only one implacement of mediating material.
~ t is possible to deri~atize the target ceramic particles with tumor specific ions or with antlbodies and/or other similarly bioactive mol2cules directed against the tumor, there~y causing specific localization of the ceramic in and/or around the tumor. Thus, agents specific to a particular tumor can be attached directly tQ the ceramic or through the use of chelating or other coupling agents.
The physical properties of tumors can also be utilized in localizing the target ceramic in the areas thereof. For example, tumors generally exhibit pH values either in the ~. .

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range of about 3-4 or in excess o~ about 8.5. The p~ of normal body fl~ids is about 7.4. Accordingly, it is possible to design magnetic target ceramics which precipitate at the pH value demonstrated by a particular tumor. Thereupon, the precipitated ceramic can be heated with the RF magnetic field.
Yet another method contemplates presensitizing the ceramic material to have affinity for a tumor species and thereafter`deli~ering the ceramic to the tumor sitP by injection, cannulation, magnetic guidance, and the like.
Such presensitization can involve the surface of the ceramic or the bulk thereof. For exa,mple, the ceramic can be etched and the pores filled with the sensitizing agent. Illus-trations of s.u~h sensitizing agents include gallium for lung carcinomata and ~Mg for low pH tumors.

Description of the Pre~erred Em~odiments.

Table I records several glass compositions, analyzed in terms of weight percent on the oxide basis, operable in the instant Ln~ention. The ac.tual batch ingredients therefor may comprise any materials, either oxides or other compounds, which, when melted together with the other components, will be con~erted into the desired oxide in the proper proportions.
~ he batch ingr.edients wer~ compounded, ballmilled to.gether to assist in achieving a homogeneous melt, and the mixture charged into silica, porcelain, MgO, or platinum crucibles. The crucibles,were introduced into a furnace o.perating at 1300-1550C and the batch ingredients melted toget~er for about one hour. Therea~ter, the melts were poured ~nto a w,ater cool.ed s.teeI mold and the melt quenched by a steel platen being immediately placed into contact with `. .` ` ` ` `
.

3~81 the top surace thereof. (In Examples 5-11 all t~e iron is reported in ~e~ms of Fe203.~

T~B~E I

Fe2O3 55 0 31.4 36.1 .36.2 34,4 19.3 FeO 4.4 11.5 18.3 12.8 P2O5 23.7 21.9 22.9 18.8 37.0 47.4 Li2O 11.6 10.. 2 - 7.8 15.3 19.2 SiO2 3.4 13.3 6.2 11.9 10.1 A123 0 4 11.5 5.5 11.0 0.26 12.9 MnO - - 10.4 - - -B2O3 ~ ~ - 2.0 MgO - - - - 2.. 38 0.25 Fe2O3 31.0 35.0 40.. 5 35.4 30.Q
P2O5 30.5 26.6 18.4 26.3 26.6 Li2O 14.. 4 14.~ Q.08 14.4 12.0 SiO2 18.8 20.0 10.8 14.2 16.6 A12O3 0.. 66 1.23 ~.15 0.14 13.7 .

MnO - - 1~.1 - -23 ~ ~ 4.84 NgO 4.68 4.46 10.3 5.46 2.81 No A12O3 ~as present in the batch materials for Examples 1, 5, and 7-10, no ~i2O was batched in Example 9, and no MgO
:: ~ was includad in the batch of Example-6. Th~ analyzed values recor.ded of those components: constitute impurities which were most likely picked up from the crucibles during melting.

1163ti~1 The melts crystalliz.ed upon cooling and X-ray diffrac-tion analyses were conducted upon the resultant products of Examples 1-4. Example 1 appeared to be almost completely crystallized, the major phase being hematitP (Fe2O3~ with a minor amount of what is believed to ~e a lithium iron phosphate.
The pattern of the latter phase did not identically match any published standard. Example. 2 exhibited substantially .re vitr~ous phase and the crystals appeared to consist of abaut 60% lithium ferrite (beli.e~ed to have the stoichiometry of LiFe3O8) and 40~ of what has been conjectured to be a lithium-doped magnetite. Thus, a slight shift in the ~e304 peak was ohser.ved. Example.3 also contained a significant amount of glassy phase with the bulk of the crystals demon-strating a diffraction pattern ~ery close to that of magnetite.
The p.re~ence of manganese in the starting materials suggests the possibility of manganese-doped magnetite or a small amount of manganese ferrite.. Example 4 appeared to have a microstructure similar to that o~ Example 2, the crystal phase consisting primarily of lithium ferrite and the postu-lated lithium-doped magnetite.. It has been hypothesized that B2O3 may be substituted in part for Li2O in the crystal phase. When placed in a magnetic field, Example 2`demonstrated the greatest activity followed by Example 4, Example 3, and Exam~le 1, in that order.
Ta~le II reports qualitative measurements of magnetism demonstrated by the exemplary compositions of Table I along with a determination of the magnetization exhibited by those compositions in a field of 700. oer.steds (M70Q). A measure-ment of the heating ~alue ~c~lories~loop/gram), as determined from the area under the hys.teresis loop, is tabulated for Examples 5-11. Finally, a qualitati~e judgment of the ii63t:i8~
squareness of the hysteresis loop is provided for Examples 5-11 .

. TABLE II

Examp~le Magnetism ~Q Cal /Loop~Gram S~uareness 1 2.9

4 ` 21 Very slight a. 42 1.76 x 10 6 Poor 6 Slight 0.67 1.48 x 10 6 Good 7 Slight 2.. 2 3.2 x 10-6 Very good 8 Strong . 6.2 2.48 x 10 5 Fair 9 ~ery strong 8.7 4.36 x io ~5 ~air Strong 7.3 1.03 x L0 5 Excellent 11 Very strong 13.26 2.0 x 10 4 Fair-good In general the preferred compositions useful in the Ln~entive method will consist essentially, by weight as analyzed on the oxide basis, of about 10-7Q% Fe2O3, lQ-60~
P2O5, Fe2O3 + P2O5 >50~ but ~9~%, 0-25% Li2O, 0-25% SiO2, Q-20~ A1203, 0-60%: B203, and 0-25% MgO. Where a ferrite crysta} phase ls desired, up to 25% of such metal oxides as CoO, NiO, and MnO may be included.
In th~ followin~ studies: illustrating the e~fectiveness of the instant in _ntion, particles of Example 1 comprised :

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~he magnetic ceramic mat~rial and murine adenocarcinoma of the breast BW10232 was utilized as the tumor. Example 1 was milled to particles having an average size of -2 microns or less and exhaustiveIy washed in isotonic ~ulbeccos phosphate buffered saline (DPBS) pH 7.2 to remove possible toxic by-products of the production and milling processes. Murine adenocarcinoma arose spontaneo~sly in the mammary gland of a C57~L/6~ ~B6) inbred mouse at the Jackson Laboratories, Bar Harbor, Maine in 1958. Gross. examination of the tumor reveals a soft, whitè, encapsulated mass with frequent hemorrhagic.zones. The tumo~ is palpable approximately 7-10 days post inoculation of tumor brei W nced, finely-divided ti~sue with variable proportions of ~ingle cell and clumped cellular masses).
Cryopreser~ed tumor having a vo.lume of about 1 cm3 was obtained from the National Cancer Institute Division of the Cancer. T.reatment Contract Production Facility, ~ason Research Institute, Worces.ter, Massachusetts. ~he tumor was stored at -2Q9C in a cryopreserv~tive medium and shipped to the applicants i~ dry ice (-7~C). The tumor was briefly held at -89C in a mechanical fre~zer until passed.
On the day of passage,. the cryopreserved tumor was qu~ck-thawed by immersing th~ frozen ampule into a beaker of di~tilled wa~er at.37 C. The tumor brei was asceptically transfexred to s.terile tissue culture petri dishes and minced with.~terile surgical 3calpel blades. All manipula-tions were ~er.f~rmed within a laminar flow containment hood recommended by the National Cancer Institute for oncogenic agents. of: undefined hazard. The finely-minced tumor was brought to a ~ina~ volume of 1.5 ml with medium 19~ (Grand Island Biological Company, Grand Island, New York~, 15 1~63~81 C57BL/J6 normal serum and brought through a sexies of decreasing diameter syringe needles ~8~20~2~t27~ until the tumor could be injected through a 27 tubexculin syxinge.
The initial subcutaneous inoculum ~5-10% suspension~ was divided equally between six C578L/6J age matched male mice (Jackson Laboratory, Bar Harbor, Maine). In approximately 10 days a 1 cm3 mass was grossly apparent in two of the six recipients.
~ariable-sized tumoxs we~e asceptically dissected free from normal fascia, debrided of necrotic tissue, and washed in DPBS. The tumor was thereàfter minced with sterile sur-gical scalpel blades, the brei diluted to an appxoximate 10%
suspension with DPBS, and passaged through decreasing diameter syringes as previously described. Analyses of the single cell population employing a 140 micron orifice and a -Coulter ZBI-H-4 channeIyzer ~Coulter Electronics, Hialeah, Florida) calibrated with lO micron spheres indicated a skewed population range from 20-380 ~m3 with the majority of the ceIls (~55~ being greatex than 8Q ~m3. Zero point 2 ml 2~ of the 10% tumor suspensions ~lx107 of 80 ~m3 or larger cells) were inoculated subcutaneously in the inguinal region.
The animals were closeIy monitored for appearance of tumor foci ~ia palpation. Any tumor cells not used for passage were resuspended in cryopreservative medium (medium 199, 15% C57BL~6~ normal serum, 10% DMSO) and refrozen to -80 C.
~umors so frozen were fully capable of-in sit.u growth when passaged ~f~er thawing.
The following is a general discussion of the experi-mental protocol performed in these studies. C57BL/6J male age matched mice were inoculatea subcutaneously with 0.2 cc of a 10% tumor preparation in the left and right inguinal * Trade mark. -16-11i3~
region. In most instances, a single left and right tumor focus developed and the inventive treatment was initiated when the tumors approached 4-5 mm diame~er (34-65 mm3 volume~.
The let inguinal region immediately adjacent to the base of the tumor was subcutane~usly injectPd with the ceramic suspension. Injection of the ceramic was conducted very slowly to avoid excessive pressure buildup within the tissue.
One in j ection site only was utili2ed in most cases because multiple injections tended to displace previously-injected ceramic out through the initial injection site. Multiple injections may be used if initial injection volumes are small. The right tumor received no ceramic but was exposed to a RF magnetic field. This action permitted the tumor-bearing animal to serve as both experlmental and control.
Prior to exposure to a RP magnetic field, surface temperatures of the inguinal tumor receiving ceramic and the axillary region were measured utilizing a calibrated micro-thermistor. The mice were then placed inside a plexiglass restrainer and po~itioned within a ten-turn, water-cooled solenoidal induction coil having a diameter of 3.5" and a height of 8". A 30 KW GCCQ motor generator was utilized to drive the coil at 10 kilohertz. The unit was capable of supplying up to 1400 oersteds within the coil. The mice were treated for five minutes at 10 ~ilohartz and 700 oersteds and t~ereafter removed ~rom the restrainer. The sur~ace temperatures o~ the treated tumor and the axilla or untreated (right) tumor were measured. In most instances, the mice received only one ceramlc injection and one exposure to the magnetic field. T~e animals were monitored daily with tumor diameters b~ing ascertained by means of a vernier caliper.
In a few experiments, only one inguinal tumor was carried :1163~81 and the mice were randomly divided into ceramic treatment alone, ceramic plus RF magnetic exposure, or no treatment.
Table III illustrates the localized heating effect caused by the R~ magnetically coupled induction of the subcutaneously implanted par~i~les of'Example 1. Hence, the average tumor surface temperature before treatment was 36.3C, whereas immediately after treatment the a~er~ge temperature was measured as 4Q.3C. In contrast, when the axillary temperatures before and af~er treatment were .measured, no significant increase in temperature was observed.
Accordingly, these data clearly attes.t to the capability of magnetic ceramic materials to induce localized hyperthermia.
Although'the recorded surface temperature meas~lred did not attain the preferred clinical hyperthermia range of 42.5-43C, the temperature of tissue closer to the ceramic particles was. un~uestionably hi'gher than 40.5C because of the .tumor regression obser.ved and described below.
Five C57BL/6J mice evidencing a single left inguinal tumor averaging about 8.2 mm in diameter were injected with 0.28 grams of Example 1 particles at t~e base of the tumor following the abave protocol. Four animals, viz.., a, b, c, and d, rec~ived a total body exposure to a RF magnetic field of 700 oerst~ds at'a frequency of 10 kilohertz for five minute.s w.hereas the ~ifth mo1~se, e, ser~ed as a ceramic-only control. Animals a and b ~containing p-articles of Example 1 and being exposed to a K~ ma~netic field~ e.xhibited g~owth af~er. 13 days in excess of that demonstrated by e, the control sample not treated. How~er, animals c and d (con-taining pa~ticles of'Example 1 and being subjected to a ~F
masnetic f.ieId~ d'emonstrated ~b~ut a twofold reduction in tumor volume on days 7 and 8 relative to animal e. To ascertain whether the obser~ed diminution of ~rowt~ rate was a reproducible result of thè ceramic ~ RF ma~netic field therapy, the double tumor bearing experimental protocol described above was preformed.
Five C5.7BL/6J mice, viz., f,.g, h, i, and j, bearing sin~.le left and right inguinal tumors ha~ing an 3verage diameter of about 5 mm were subcutaneously injected with 0.3 grams of Example 1 particles at the base of the left tumor only. After this injection the entire bodies of f, g, h, and i were exposed for five minutes to a 700 oersted RF
magnetic ~.ield at a frequency of 10 kilohertz. Thus, animal j, containing ceramic particles but not treated, ser~ed as the control. In an-imals f, g, and h, the left tumor mass grew at a diminished rate reIative to the non-treated right tumor. Animal h represented particularly interesting data s.ince no. detectable tumor was present on the right side at the time o the RF. ~xposure, but after sa~eral days a focus appeared which rapidly outqrew the treated left side.
Animal i responded poorly to treatment although gross exami-nation after seven days revealed a reduced le~t ~treated~
mass when compared to the right Cuntreated~ mass. Examination o tha tumors in control animal ; manifested essentially equivalent-growth on both ~he left and right sides.
~ i~ht days after. treatment the animals were sacrificed and careful gross. dissection of the mice was undertaken. An appreciabLe accumulation of ceramic was noted in the treated tumor a~ea with little or no o~ser~able spreading. This latte~ findin~ was of especial imyortance since it indicated that the RF. exposure can be effecti~ely reapplied on subsequent occasions af.te~ the initial injection of ceramic particles.

1163~8~

Th~ final weights o th~ tumors dissected cleax o~
normal tissue and ceramic were measured. ~he data rep~rted in Ta~le IV clearly illus~rat~ a tumor mass reduction factor (left side vs. right side~ ranging from 1.81 to 4.81.
Control animal j demonstrated slmilar left and right tumor masses wi~ a ratio of 0.91.
Examination of the ~ormal tis~sue immediately adjacent to the ceramic particles manifested essentially no evidence of necrosis resulting from th~ ceramic-mediated, magnetic field treatment.

TABLE III

Surface Temperature Surface Temperatu-e Before Treatment (C) A~ter Treatment (~~) Animal Ceramic Dose Tumor Axilla Tumor illa a 0.28g 36 37 42.5. 37 b 0.28g - - 40.5 f 0.3~`g 36 3~.5 40 37 g 0.30g 37 37 38.5 38.5 h 0.30g 37 37 43 38 i 0.30g 35.5 36.5 37.5 36.5 ` TABLE IV

Let Tumor Right Tumor Ratio knimal Mass (~) ass ~8) Right Tumor Mass:Left Tumor ~ass f 1.~5 3.35 2.03 g 2.20 4.0 1.82 h 0.80 3.85 4.81 1 lost* lost*
3.5 3.2 0.91 *Animal died on seve~th day. Gros~ observation on se~e~th day indicated an approximate two-old difference between left and right tumor masses.

The above studies, illustrating the absence of`mlgxa-tion o the ceramic particles ~rom the site of the subcutaneous injection, coupled with the excellent tumor mass reductions observed after a single five-minute RF exposure, clearly suggested that total tumor eradication could be possible where exposure times are increased and/or multiple treatments are applied. Moreover, a more highly m~gnetic ceramic ~ould require less time to heat the tumor tissue to the hyperthermia range. The effectiveness of the ceramic can also be improved by increasing the coercive force t~ereof via the addition of such doping ions, as cobalt into the ferritic structure.
In a ~ingle experiment with a C57BL/6J mouse utilizing the above protocol, 0.3 grams of Example 1 were injected intratumor and subcutaneously,at the margins of an inguinal tum~r. Th~ body of the animal was then subjected for one hour to a 700 oersted ~ield having a fre~uency of 10 kilo-hertz. Subs~uent examination indicated essentially total eradication of the tumor mass, thereby substantiating the above hypothesis that total tumor extirpation is feasible utilizing the inventive method. Further experimental work has demonstrated that administering pure unencapsulated magnetic susceptor materials, e.g., magnetite, at one-half the dosage le~eI utilized with the in~entive materials producad gross inflammation and ob~ious rejection.

Claims (11)

The embodiments of the invention in which an exclusive property or privilege is claimed are defined as follows:

1. A ceramic selected from the group consisting of glass, glass-ceramic, and sintered ceramic suitable for inducing localized heating in the presence of a radio frequency magnetic field, said field having a frequency sufficiently low that essentially only magnetic hysteresis heating can occur, which has incorporated therewithin magnetic iron-containing crystals of such size, compo-sition, concentration, and magnetic properties to impart a coercive force of at least 200 oersteds to said material.

2. The ceramic according to claim 1 wherein the ceramic is a glass having a base composition selected from the group of phosphates, silicates, and borates.

3. The ceramic according to claim 1 wherein the iron-containing crystals consist essentially of magnetite or a ferrite.

4. The ceramic according to claim 3 wherein the ferrite is selected from the group of lithium, cobalt, nickel, manganese, and barium ferrite.

5. The ceramic according to claim 1 wherein the frequency of the radio frequency magnetic field will not exceed about 10 kilohertz.

6. The ceramic of claim 1, characterized by the use for generating hyperthermia in a tumor mass surrounded by organic tissue.

7. The ceramic according to claim 6 wherein the ceramic material is derivatized with tumor specific ions or with antibodies and/or other similarly bioactive molecules directed against the tumor.

8. The ceramic according to claim 6 wherein the ceramic material is designed to precipitate at the pH value demonstrated by the tumor.

9. The ceramic according to claim 6 wherein the ceramic material is presensitized to have an affinity for the tumor species.

10. The ceramic of claim 2, 3 or 5, characterized by the use for generating hyperthermia in a tumor mass surrounded by organic tissue.

11. A ceramic as in claim 1, 2 or 3 wherein the coercive force is about 700 oersteds.