The Therapeutic Effect of Total Body Irradiation Followed by Infusion of Stored Autologous Marrow in Humans

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Cam it tar on Roomrch. C:*.cinruti Canard Hospital The Therapeutic Effect of Total llody Irradiation Pollawed by Infurion of Stored Autologourr Marrow Purpose of Studyi The goal of thi8 study is to teat the hypothesi8 that total body irradiation followed by infusion of stored autologous marrow i s effective palliative therapy for metastic malignancy fn human beingr. Total body irradiation ha# peon ueed for palliation by othsrr (1,4,;,6) but the doas usad has been United to lees than 200 R because of the bone marrow depression which follows largo doses of radiation. A combination of total body radiation and infusion of of total body radie.tion. Effoctive t o t a l body radiatiori therapy connidcred awies precaution when new drugr .I treatment schedules are being explored1 it may than be por sib lo

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t o troat tho occanionrl unexpectcdly novere hemopofctic . * c patient that after 800 rad totvl body iryadiation, preceded by 4 '!. .' cluccessful take of homologous marrow occurrod. ' of bow marrow cells will repopulate the nrarrw space i n mice . * ( 2 2 ) , r a b b i t s ( 2 3 ) and dog8 (21) intravenous administration of . 4 stored marrow cells prevented death a f t e r an otherwise lethal ..1,? dose of radiation. It is not possible t o apply this information 4 5 directly to human beings because of the variable response of ;;,\,*.!' . different species. We, and others (24) have established fn vitro .''a;?I'd involve total body radiation or adequate doses of radiomimetic 1.- ;: chcmotherapy. Therefore, it i s necessary to test humans M a ). I... following total body radiation and t o proceed with developmantal 9 : .! ..viability of bone marrow cello stored a t -80 degrees C. for as long as one year. However, almost allin vivo studies i n human beings where marrow transplantation !;as been attempted did not effcctivcness of marrow i n repopulating the marrow apace ..? tachniq5ea and refinomants of marr w infusion. In the management of patient8with malignancy who are to receive myelotoxic .? of o to red autologous marrow may well be lifr. savinq. ::t

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Patients will receive 200 rad total body frradiatioq mu !.+( treatment for metastatic malignancy, (A control group incruder '.*;*:j:- Patiento who have already been irradiated atthe 200 rad done . 2de *. .. level and did notre wive bone marrow in fue ion.)A second group of patients will be treated with 200 rad total body irradiation, followed by infusion through filters' of Lheix previously tstored marrow i.mcd i at ely after irradiation. P.itients seLected will be tP,osc who are to receive total body irradiatioi: i n the treatment of metastatic malignancy from varicus primary sites, excluding neoplasms which are treated by internal radiation, e.g., carcinoma of the thyroid. They will have & t a b 1 3 hematologic values and may have received local The harrow will be infurred intravenously with filtration to UpcninrJs of 297, 149 and 74 micton.9). Filters and tubing will be st cri io and pyrogen free, Initially, all t-arrow will be infused through the filters. If tho filtered marrow does no+ rep pr lla to the marrow space, then unfiltered marrow will be infused to determine whether filtration :$as the reason the in ft qi ion was not effective.

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radiation previourlv. Patients will be has Cincinnati General Hospital and will be treated with 200 rda :i ;(midline absorbed tissue dose) of total body irradiation at an dose of radiation approximates the therapeutic and hematologic e ffcc tr of 0.4 - 0.6 mill iqr ma of nitroqen mustard per kilogram 'I Caplet e blood counts including hemoglobin, hematocrit, ?I count, diffarential smears and cell indices will be obtained and 1, 2,3,6,9,12, etc, days aftcr irradiation according to n. t . the standard test days of Thana and UaLd (7): Pre- and poat1 1 irradiation immunologic capabilitics will be evaluated as vell. 327 r * Additional counts will be obtained a s clinically indicated. Granulocyte reserves will be tested with etiocholanolone so a8 to ascertain the safety of giving 200 rads to a patient (8.0 1 t , Bone marrow specimens for cytoloaic and pathologic i& examination will be obtained from the stctunl and posterior ilium the week prior to ir radi at ton by the method nf Kurnick (8,9) 01

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bone marrow will be cultured for sterility prior to freezing. 3 vitro viability of the marrow will be estimated by try ran blue DNA synthesis(11). and acrid in e orange 112) are being developed. Peripheral blood data will be compared to those which w e - but tests utilizing tissue culture (10) have already obtained inpatient who have been irradiated a t the same dose level but have not received marrow infusion..I There will be a continuous review of the results as they are being obtained. If it becomes obvious that the results are .. ' auto logo us marrow transfusion) will be compared with data I: The data from these two groups of patients(with and without obtained from patients with similar neoplasms who have been , .u n treated and from patients who have received chenotherapy (25)..p according to age, sex, race, type,duration and extent of neoplasm. Asa result of this compariscn, it will be possible t o the palliative value of total body irradiation at thie4 dose level with and without bone marrow infusion.v.. with any irradiation therapy there is the danger of < t i hematologic values in the control atoup. Two hundred rads of G i radiation injury. We hsve obscrved consistent depression of *

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observable therapeutic effect be anticipated but a180 a t 200 I, the effect of marrow reinfuaion will be detectable. 1 In thie Etudy the prodromal morbidity ~mmediatsly after irradiation (anorexia, nausea, vomiting) would approximate 80% and raaponde well to phenothiazines, r.s to mortality fig use at the doragee t o be evaluated, in human beings t h e LD50 for bone marrow apl asia occurs at about 350 R and " intestinal death. occurs above 1000 R. With hospital care it is felt that a human LD50 may approximate SO0 R or Itighet. The doses proposed are below these levels. When the mean eurvival time of patients treated with less than I G O rad, 150 rad, and 200 rad are compared there i r no progresoi-*e decrease related to dose (Tables I, XI). If the patients with survival times of 9 and 10 day8 are excluded, and the patients still living at the present time are included, there is s t i l l no significant difference in survival time which can be , aaaociated with the irradiation as determined by several statistical tests. In f a c t the data suggest that (1 therapeutic effect from the radiation therapy has been obtained, The use of autologous and isogeneic marrow i n humans has been investigated by others (88 9, 13, 14). %'hen marrow has been infused f.35 rapidly, cerekral hemorrhage, hemoglobinemia and encountered .,!-...en the rate of infusion has been maintained lelw

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There is dFaawecr?ant as t o tha naed for filtration. Kurnick doc:^ not fecl it is necessary a t thc time of infusion ; Thomas, L'errcbcc, and Pillow (13,lU,lS) rccorunend its use to minfmizs ttic incidence of pulmonarv erAolism. A series Of filters will be used with the now opc ratio riol constant rate To datc, we have infused unfiltered inarrow intravenously in five pa tier its. One patient hac! hemoqlobinuria. Autopsy material and 8 . Ymayuchi. No evidence of Yulrr.onary amtoli, pulmonary in farc tL on or sther disease as a result c;f marrow infusion was :11 ! at i ents will be fol1oi:ed carefully clinically. ! I cm n to lw ics l data are accumulated at Crewent ic ter vals ana all ;Tcasur'?s to control infection will IJe csed. TliL"; may include p lao tic barricr " Life Ielands", reverse isolation, and laminar air flow units if needed. Prophylactic antibiotic therapy will Previous r!ork Done i n Tt.is Area: U u ring the past seven ycars an ongoinr; study of the oetaLolic cffccts of total body irradiation hag been i n progress. (See enclosed ~ r o t o c o l sA and 8 ) . TO date, cvcr 7 0 patients have bCeA treated with25to 200 rad total to dy irradiation. The conparabla p atl cnt n treated with other agents. nono marrow f:c?

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23 patients h3s been stored; it has been infused intravenously in . blarkad hematologic depressiot, occurred in all patients vho -*., received more than 125tad total body radiation. There waa a J platelet count depression 25-40 days after irradiation. The eeM, .Y minimum leukocyte count of previously untreated patients who r e c e i v e d 1 5 0 rad total body radiation only was 1 2 6 4 f 1 1 4 0 , and f .iwas 1 1 4 0 2 816 when t h e r e had been previous therapy. The nadir of the leukocyte counts of patients who received 200 rad was 983$ A corn-arlrion of the hematologic chancjes inpatients a: receiving partial body irradiation with those who received total.r body irradiation revealed a paucity of change in the total WBC, platelcts, and hematocrit even when the patient received more - \ sere given partial bcciy irradiation, lymphopenia did not occur ...- -.than 209 rad partial body radiation. 111 2 of the 1 0 patients who until48 or 7 2 hours after exposure. .rZ Infusion of stored autologous boil@ narrow has been completed cells/cuhic nun. will be necessary for successful auzologous In 4patients who ware studied in detail,end orr duplication has ' 7.

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chromosome preparotioao (16).This observation nay bo w soci at ad vlth malignancy but baa ', increased following to to1 body irradiation. As an OutgrGwth of the experfence with was the one which might be undergoing endoreduplication was considered. Therefore, a method.of cell separation utilizing a endoreduplication, tho qu seti on,a st to whether thc rnaliynant cell Pic011 gradient and was undertaken. Friedman, et al., separation of fol loving this cell (17). lymphocytes from granuiocytes. hromosool?ie culture tcchniquer ' tud is s of a Another facet of considerable in t crc st in patients receiving large doses of to body irradiation ?-..IS been the question of 02 serum obk;ained from patients boforc and aTtcr t o t a l body alteration in the of hman scrum gai:,~.a u los ul in 3s a result of ( 1 8 ) . Incncdiate end delayed hypersensitivity after radiation, using the

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Content forms, am attached, have !:con used since May 1, 1965. Those form:. are signed -nl:f alter the patient has been advised of the research and study asscct of the procedure8 to be used. The patients are told this is ii € o m of therapy but that benefits cannot b e suarantccd. They are iv:vi.nvl of the possible * 'I complications, including the possibility of pulnonary in fu ct 'ri following rnzrrow infusion. Since total body raciiation is another mode of treatment for patients with an ul tir a at sly fatal disease,i it is only correct t o indicate th cp ossi kei lit y of therapeutic a advantages. All patients are inf.Iv.eb that rl rick exists, but that all precautions to prevent untoward results will be taken. :?any studies t:ave been carried out in animals regarding the '.' j protective effect of hone marrow follo*.-:ing radiation. They are -. .,.< well sunnarized in these paragraphs fror! Pcgq's ~ ook. (1) all c conic taemopolctic cells was first cicnonstratcd 1-y Lorens an&

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1 9 5 3 1 Alpcn and LIdum, 1 9 j U i Puzil, o t al., 1961; Josdan, et Sl., 1961 1 but tlia Cr~o~\cr::t.owig w u p sing more closely relatod dog@ have rmortad more succcsstul all o qe nic protection (Farrebeo, et de,195U; ':hornas, et a l e , 1 3 5 8 1 Thomir: i!t a l . , 1 9 5 9 1 . Others hove iincc conLirmd these findings (?'tanJ;ull, etale, "The survival of ir radiata d micc in fu wd with win cap l q 18, hPi/c*-p<sr, k-ide agtccmcnt that rat bonc marrow will protcct I t :*: most important that methods of protcctinri against autolotious 3arrow is the s a € e s t protective tc;chni.,;ic to Le used This r.t'&!:. i n pro>o?cd a!3 a Continuation of tka work cited fat the 1. To c'ralunte the role of total bo;. rsdi.ition and

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2. To evaluate autolagouo bonm m(~rrout ru u fusion in tha treatroont of bone marrow deprasalon from To dotormine whether storad autologou3 bone marrow therapy may play a role i n treatment of bcme mur w deprereion following acute radiation espo8ure fu war fate or occupationally induced accidents. Thi8 work i a conriderad to be of vital fnportanca not only for ir!ptoving thee aurvival of patients with €ar advanced cancer but for the Gutvival ~f the citizens of this nation In the event of nuclear warfare or a nmjor peacetime radiation accident.

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Harrow Transplantation. Year 3 5 Eoet Wvckar Drivo, Chicago Coniidcration of t h e Use of Bone Marrow Transplantation, p. 21 i n Progrsrs In Hematology, Vol. XI, Tccantinr, L. H. ed., Grune and Stratton, 1959. and Clinical Effects Induced i n 2 6 3 Cancer Patitfits by WholoBody X-irradiation with Nominal A i r Doses of 15 to 200 R. School of Aviation Madicine, US-, Report N o . 57-92. t i v s Effects of Total Body Radiation, Nitrogen Muetard and Trie th v lene Melamine on the Hemtopoietic System of Terminal Science -1 1 8 : No; 3056, 1 6 1 - 1 6 3 , August 7 , Sfnglc Doses of Total Body Radiation. The Am, J. of Roentgenology, Radium Therapy and Nuclear Medicine 2: No. 3 , 7. Thoma, G.E. and Wald, N . The Diagnosis and Management of the Accidental Radiation Injury. Irradiation. He ma top oie tic Depression i n Man zythe Infu.iOa of Stored Autogmoue Bone Marrow. Ann. Int. cede zr 1200 cent of Severe fat ro qe nic Hye?.--C.~~.Jr.,ssion. Diagnosis and Treatment of Acute Radiation In jut y,i361, Ganeva, World Zioalth Organization, p B 309. parationo of Leukocyteo Cultured from IIurnan Peripheral

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Bono WTOW,Proc. Inmt. Mod. Chicago 741 227, 1963. dine Oranga Viability Tcrst Applied t o Bono Miirrw Cells. I. Corral ation with Trypan Blue and Eosin Dye Exclusion Failure by Isogenic Marrow Infusion. H. Eng. J. Of The ColLection, Storage and Preparation of VA able Cadaver Narrow for Intravenous Uue, Blood 14s 190-107, 1959, reduplfcation i n Leukocyte Chromosomes, Preliminary Feport of its Ral ation to Cancer and Whole-Body Irradiation. The Lancet, September 5 , 196U, 494-495. Friedman, 8 , 1. and Toler, S. J. Chromosome Studies of Peripheral Blood kuk o cyte a Separated by Cent ri fuga tion in aFi coll Gradient. Ewp s rh n tal Hematology, No. 9, Irradiated Humaru. …