induced pancreatic malignant tumours

Transcription

1 Targeted radotherapy wth 177 Lu-DOTA-TATE n athymc mce wth nduced pancreatc malgnant tumours J. RODRIGUEZ-CORTES, C. ARTEAGA DE MURPHY *, G. FERRO-FLORES, M. PEDRAZA-LÓPEZ, E. MURPHY-STACK #. Department of Nuclear Medcne, Insttuto Naconal de Cencas Médcas y Nutrcón Salvador Zubrán, Méxco. Faculty of Medcne, Unversdad Autónoma del Estado de Méxco, Department of Radoactve Materals, Insttuto Naconal de Investgacones Nucleares, Méxco. #Department of Pathology, Hosptal Santelena Méxco. Correspondng Author: *Consuelo Arteaga de Murphy, PhD. Insttuto Naconal de Cencas Médcas y Nutrcón Salvador Zubrán Vasco de Quroga #15. Delegacón Tlalpan. Códgo Postal 14. Méxco D.F. Méxco. Tel. 5 (55) ext. 4 e-mal: consuelo_murphy@yahoo.com.mx 141

2 Abstract Malgnant pancreas tumours nduced n athymc mce are a good model for peptde receptor targeted radotherapy. The objectve of ths research was to estmate pancreatc tumour absorbed radaton doses after admnstraton of 177 Lu-DOTA-TATE n mce as a therapeutc radopharmaceutcal that could be used n humans. AR4J murne pancreas cancer cells expressng somatostatn receptors, were mplanted n athymc mce (n=18) to obtan the 177 Lu-DOTA-TATE boknetcs and dosmetry. To estmate ts therapeutc effcacy 87 MBq were njected n a tal ven of 3 mce and 19 days p.. there were a partal relapse. There was an epthelal and sarcoma mxed tumour n the kdneys of mouse III. The absorbed dose to tumour, kdney and pancreas was 5.5 ± 7. Gy, 17.5 ±.5 Gy and 1.6 ±.3 Gy respectvely. These studes justfy further therapeutc and dosmetry estmatons to ensure that 177 Lu-DOTA-TATE wll act as expected n man consderng ts kdney radotoxcty. Key words: tumour dosmetry, 177 Lu-DOTA-TATE, tumour uptake n athymc mce 1. Introducton Neuroendocrne tumours over-express somatostatn receptors (SSTRs). These receptors are the targets of radolabelled somatostatn analogues whch are used for the detecton and therapy of wde spread dseases, hard to treat wth radotherapy or whch are refractory to chemotherapy [1]. Lutetum-177 s a radonuclde wth a half lfe of 6.71 d, β max.497 MeV (78 %) and.8 MeV (11%) γ radaton. Octreotate (TATE), a cyclc octapeptde (DPhe-Cys-Tyr-DTrp-Lys-Thr-Cys-Thr-COOH), s a somatostatn analogue that targets somatostatn receptors over-expressed n cancer cells []. The TATE-DOTA conjugate (1,4,7,1-tetraaza-cyclododecane-N,N,N,N -tetracetc acd) can be labelled wth 177 Lu for radopeptde therapy of neuroendocrne tumours, 14

3 manly carcnods. The cell lne AR4J, from murne pancreas cancer expresses somatostatn receptors, especally SSTr. Induced malgnant pancreas tumours wth AR4J cells n severe mmunodefcent mce are a good model for targeted radotherapy. There are not reports about the behavour of a pancreas cancer tumour nduced wth over expressng SSTRs AR4J cell lne, nor concernng the 177 Lu-DOTA- TATE absorbed radaton dose n mce [3-6]. The objectve of ths research was to estmate nduced pancreatc tumour absorbed radaton doses and to evaluate 177 Lu- DOTA-TATE as a therapeutc radopharmaceutcal that could be used n humans.. Expermental Preparaton of 177 Lu-DOTA-TATE. The DOTA-TATE peptde conjugate was syntheszed wth a purty > 98 % as analyzed by hgh performance lqud chromatography (HPLC) and mass spectroscopy (PChem, Austra). A 5 µl alquot (1mg/mL 1 % ethanol) was dluted wth 5 µl of 1M acetate buffer ph 5.5, and 5 µl of the 177 LuCl 3 n.5m HCl (185 MBq, >1.85 TBq/mg) was added. The mxture was ncubated at 9ºC n a block heater for 3 mn. Qualty control. Instant thn-layer chromatography on slca gel (ITLC-SG), sold phase extracton (Sep-Pak C-18) and reverse phase HPLC analyses were used to determne the radochemcal purty and stablty. The stablty n human serum was carred out at 37 o C takng samples for analyss at dfferent tmes from 3 mn to 4 h. Inducng cancer tumour n mce. Anmal studes were carred out n athymc mce n accord wth the Mexcan Offcal rules and regulatons for handlng laboratory anmals. Male, g (n=) were mplanted 1 x 1 6 (.5 ml) AR4J murne pancreatc cancer cells (ATCC, Rockvlle, MD, USA) n the rght dorsal muscle. Tumour growth was frequently montored and the sze was measured wth calpers n cm. 143

4 Boknetc studes. When the tumours were vsble a mean of 18.5 MBq 177 Lu-DOTA- TATE was njected n a tal ven of mce (n= 3 for each tme nterval). After, 4, 4, 48, 7 and 168 h mce were sacrfced n a CO chamber. The organs or tssues (blood, heart, lung, lver, spleen, kdneys, pancreas, femur muscle and femur bone) were excsed, blotted dry and put nto plastc tests tubes, prevously weghed. The weght of the organ was recorded. The actvty of the organs was detected n a crystal scntllaton well-type detector. A dluted alquot of the njected actvty (I.A.) was taken as 1% and the % uptake of each organ or tssue was calculated. Therapy studes. To 3 mce ~ 87 MBq of 177 Lu-DOTA-TATE was njected n a tal ven. One mouse, wthout radopeptde admnstraton, was used as control. Dosmetry estmatons. The number of dsntegratons that occurred n the man source organs was calculated from the boknetc data. Actvty s the number of dsntegratons per unt tme and when ntegrated over tme gves the total number of dsntegratons (N) expressed per unt of ntal actvty n the source organ (MBq-h/MBq) (equaton 1). t= N source = A dt (1) The absorbed dose to organs was evaluated accordng to the general equaton : D t= source target source = Nsource DFtarget source sources () where D s the mean absorbed dose to a target organ from a source organ and target source DFtarget source s a dose factor (equaton 3). Φ DFtarget source = (target source) (3) The terms are the mean energy emtted per dsntegraton for the varous -type radatons (-type emssons, n E ). The Φ terms are the absorbed fractons that depend on the propertes of the -type emsson and the sze and shape, and separaton 144

5 of the source and target organs. DF values were calculated as reported Mller et al. [7] usng the beta-absorbed fractons n a mouse model calculated by two Monte Carlo radaton transport codes, MCNP4C and PEREGRINE (voxel-based). As kdney and pancreas are overlappng surface areas, the average dose was calculated accordng to equaton 4 and 5. D pancreas = N pancreas Φ (pancreas pancreas) + Nkdney Φ (pancreas kdney) (4) D kdney = Nkdney Φ (kdney kdney) + N pancreas Φ (kdney pancreas) (5) Pathology studes. Pathology routne studes of the tumours and kdneys were undertaken to determne f the growth caused by the AR4J cells was of cancerous orgn or from a foregn body reacton. 3. Results and dscusson Radopharmaceutcal purty. The labellng effcency or radochemcal purty was >98 %. The labelled peptde was stable n human serum after 4 h. Boknetcs. The radopharmaceutcal had a rapd blood clearance and renal elmnaton. There was uptake n the tumour, pancreas and n the kdneys and mnmal uptake n the other organs. Fgure 1 shows boknetc data for the man source organs. Tumour, kdney and pancreas uptake at h post njecton were % I.A./g, 7.7 % I.A./g (1.71 %/organ) and 4. % I.A./g (.4 %/organ) respectvely. The tumour/blood rato was and the tumour/muscle rato was 8. Therapy. The sze of the tumours decreased after the therapeutc dose. For the tumour of mouse number I there was a 9 fold decrease and.3 and 1.5 fold for mce II and III (see fgure ). There was a partal remsson and the tumours started growng agan untl the anmals were sacrfced. The tumours mght have been completely reduced wth a 145

6 second therapy dose. The control mouse had a 6. cm tumour 8 d after cell mplantaton. Dosmetry. Calculated average radaton absorbed doses to tumour, kdney and pancreas were 5.5 ± 7. Gy, 17.5 ±.5 Gy and 1.6 ±.3 Gy respectvely (see table 1). The maxmum tolerated dose to the kdneys s 5-4 Gy and n our study t was 19.8 Gy for mouse number I. A second therapy dose could produce severe kdney radotoxcty. The pancreas receved a lesser dose. Lews rats bearng small cell lung tumours had a 1% cure rate after repeated njectons of 77 MBq of 177 Lu-DOTA-TATE [8]. Targeted radopeptde therapy delvers a cytotoxc dose to all the cells wthn a tumour whle sparng normal tssues. Saturaton of the SSTRs, heterogenety, blood flow, vascular densty permeablty and bologcal dfferences may have nfluenced the uptake n tumour and SSTR-postve normal tssues [1]. Pathology studes. Pathology studes demonstrated that the sphercal (but nvadng adjacent tssues) tumours were of hghly undfferentated type of cancer compatble wth pancreatc cancer. There was an epthelal and sarcoma mxed tumour n the kdneys of mouse III. Athymc mce have dfferent grades of mmunologc defenses n spte of beng called nude (wthout mmunologcal defenses) or athymc. A mouse wth a small tumour shows a great mmunologcal reserve and the anmal wth the large tumour s a true athymc anmal wth no defenses aganst mplanted cells. Mouse III was not completely athymc because t had a small dorsal tumour and a kdney tumour. In humans t has been reported that D-lysne or a gelatne based plasma expander reduce kdney uptake and toxcty and ncrease the effectve tumour dose [8, 9]. 146

7 4. Conclusons Tumour growth s stll subject to unknown bologcal factors among whch we have mentoned varous possbltes. We could not fnd reports about the behavour of nduced cancer tumour nor concerned radaton dose. The am of ths prelmnary research s to encourage addtonal nvestgatons on the role of 177 Lu-DOTA TATE n the treatment of malgnant tumours. Acknowledgement Ths study was supported by the CONACyT-Mexco (SALUD-4-1-3) and the Internatonal Atomc Energy Agency (IAEA). References [1] R.E. Wener and M.L.Thakur, Bo Drugs (5). [] G.A. Spyroulas, A.S. Galans, C. Petrou, et al., Med. Chem (5). [3] A. Nkolopoulou, T. Mana, P. Sotrou, et al., J. Pept. Sc (6). [4] C. Arteaga de Murphy, M. Pedraza- López, G. Ferro-Flores, et al., Nucl. Med. Bol (1). [5] W.A. Breeman, A. Mearadj, A. Capello, et al., Int. J. Cancer (3). [6] M. de Jong, W.A. Breeman, B.F. Bernard, et al., Int. J. Cancer 9 68 (1). [7] W.H. Mller, C. Hartmann-Santar, D. Fsher, et al., Cancer Bother. Radopharm. 436 (5). [8] A. Schmtt, P. Bernhardt, O. Nlsson, et al., Cancer Bother. Radopharm. 31 (5). [9] J.E. van Eerd, E. Vegt, J.F. Wetzels, et. al., J. Nucl. Med (6). 147

8 Table 1. Radaton absorbed dose n source organs after admnstraton of 177 Lu-DOTA- TATE n athymc mce wth nduced pancreatc malgnant tumours.* ATHYMIC MICE Mouse I Mouse II Mouse III Injected actvty (MBq) Tumour radaton absorbed dose (Gy) Kdney radaton absorbed dose (Gy) Pancreas radaton absorbed dose (Gy) * boknetc studes were used for the dose estmates. 148

9 Fgure captons Fgure 1. Boknetcs of 177 Lu-DOTA-TATE n the man source organs of athymc mce wth nduced pancreatc malgnant tumours. Fgure. Tumor sze after njecton of 177 Lu-DOTA-TATE n athymc mce wth nduced pancreatc malgnant tumours. 149

10 TUMOUR PANCREAS A(t) =.8 exp( 9.7t) +.9 exp(.1t) +.8 exp( 1.3E t) N = A(t)dt = 6.9 MBq h/mbq R = A(t) =.9 exp( 14.9t) +.5 exp( 1.1 t) +.1exp( 9.8E 3t) N = A(t)dt =. MBq h/mbq R = Tme (h) Tme (h) N = R =.97 KIDNEY A(t) =.9 exp( 1t) +.3 exp( 5.3E t) +.3 exp( 5.5E 3t) A(t)dt =.7 MBq h/mbq Tme (h) 15

11 Mouse I Sze of the AR4J cell tumour (cm ) Tme (d) Mouse II Mouse III Sze of the AR4J cell tumour (cm ) Tme (d) Sze of the AR4J cell tumour (cm ) Tme (d) *NOTE: The control mouse (wthout treatment) had a 6. cm mplantaton. tumour 8 d after cell 151