Document krwD04Rj9RX5JR98bvz2Eja0

FILE NAME: RT Vanderbilt (RTV) DATE: 0000 DOC#: RTV122 DOCUMENT DESCRIPTION: Final Report on Biologic Tests on Sample from RTV FINAL REPORT ON BIOLOGIC O N SAMPLE C P S - 2 7 5 - I * TESTS Report submitted to R T Vanderbilt C o ., Inc. Report prepared by: William E. Smith, M .D . Director, Health Research Institute Fairleigh Dickinson University *CPS-275-l is a sample of the tremolite portion o f a typical New York State talc ore normally containing 40 to 60 percent trem olite. This material was ground to a nominally respirable size (less than 10 micro meters) For test purposes. The sample is approximately 90 to 95 percent tremolite, the remainder being antigorite. AAABGESQT inljxt .WO.'- 4 WES0010.48*1*? Page 1 Previous work ?r> this laboratory has shown that intrapleural injection of hamsters provides an animal model for estimation of biologic effects of fibrous minerals. Fibrous pleural adhesions and tumors with characteristics of mesotheliomas have developed in male Golden Syrian hamsters of the LVGrLAK strain after single intrapleural injection of a variety of preparations of asbestos^1' 2' 3' . In the present work the hamster-intrapleural model was used to test for biologic effects, with special reference to fibrogenieity and carcinogenicity, of a milled sample of trem olite ore prepored by R. T. Vanderbilt Company, Inc. and identified as Sample CPS-275-1. This sample was received a t Health Research Institute as a white powder. For injection into animals, it was suspended in 0.9% sodium chloride solution in water at concentrations of 25 mg and 10 mg per 0 .5 ml. These suspensions were sterilized by autoclaving.250F for 15 minutes. Each test hamster was given a single injection via a 20 gauge needle into the right pleura! cavity of 0.5' ml of one or the other o f these suspensions. Control hamsters were given similar injection of 0 .5 ml of the autoclaved saline solution. > * ` < The experiment compared pathologic findings in 3 groups of male LVG:LAK hamsters injected a t the age of about 2 months. Each animal in Group 1 was treated with 25 mg CP5-275-1 . Each animal in Group 2 was treated with 10 mg CPS-275-1. Each animal in Group 3 received 0 .5 ml saline solution. Group 3 thus served as a control on the suspending medium and also on any tissue response to injury by the in jection procedure. The design of the experiment was to place 70 hamsters in each group, to kill 10 from each group a t 3 months for pathologic examination for any early lesions, and to maintain all others For long-term tests for carcinogenicity. In the first 3 months, an outbreak of ocute enteritis (a common spontaneous disease of hamsters) resulted in deaths in all groups (17 in Group 1, 29 in Group 2, 22 in Group 3), Additional hamsters were treated and entered into the 3 groups to replace losses from these early deaths. At necropsies of animals that died or were killed for specimens between 49 and 93 days, hamsters treated with control saline showed no remarkable changes in lungs or pleura, whereas slight pleural adhesions were found in all of 10 hamsters that had been treated with 25 mg CPS-275-1 and in 14 of 16 hamsters that had been treated with 10 mg CPS-275-1. Extent of these adhesions was rated on a 1 to 4 scale, with 1 representing minimal and 4 representing extensive. As shown in Table 1, adhesions rated minimal (I) were found in 7 animals and slightly more extensive adhesions, rated 2, were found in 4 animals on the 25 mg dose. In animals on the 10 mg dose, no adhesions were found In 2 animals, adhesions rated minimal (1) were found in 13 and slightly more extensive adhesions, rated 2 , were found in a single anim al. In previous work, we hove seen more extensive adhesions (rated 3 or 4) in hamsters within 3 months after intropleurai injection of 25 mg or 10 mg doses of chrysotile asbestos, Histological sections Were prepared for microscopic examination of lungs and pleuro from each of the 5 hamsters killed 92 days after treatment with the 25 mg dose and from each of 5 hamsters that died between 80 and 93 days after treatment with the 10 mg dose. These sections wore stained with a modified Gomori's One Step Trichrome Method using W ES001049.. Page 2 aniirne blue(s). Examination of these sections shows areas of fibrous thickening of the pleura in all of the 5 hamsters on the 25 mg dose. These oreos stain blue indicative o f collagen. Sections on the 5 hamsters on the 10 mg dose show a small a re a of blue- staining fibrous thickening of the pleura in only a single anim al. ' Accordingly, the findings show that, under the conditions of these experiments, Sample CPS-275-1 is slightly fbrogenc for hamsters a t the high (25 mg) dose, but only questionably fbrogenc at the 10 mg dose. In previous work in this laboratory, the earliest tumor attributed to intrapleural injection of asbestos was found in a hamster examined 151 days after injection of chrysotilei3) . In the present experiment, the number of hamsters surviving 135 doys or longer was 53 in Group 1, 49 in Group 2 , and 59 in Group 3. These animals were followed as long as they lived and then examined by necropsy. As shown in Table 2, most of these animals died of natural causes, a Few were killed because moribund, and all survivors were killed for examination between 600 and 610 days. Average survivorship was 392 days in Group 1, 428 days in Group 2, 421 days in Group 3 . Differences in'survivorship could not be correlated with pathologic findings. As the,animals grew older, lesions in the pleural space were not progressive and resembled lesions described in animals examined between 49 and 93 days. Corrtmon findings at necropsy were enteritis (22 cases in Group 1, 17 in Group 2 , 23 in Group 3) thrombi in cardiac atria (7 cases in Group 1, 4 in Group 2 , II. in Group 3), renal disease (5 cases in Group 1 , 8 in Group 2 , 7 in Group 3). Enteritis, cardiac thrombi and renal disease are common causes of death in untreated control hamstersi8). As a measure of general health, average body weights of hamsters in the 3 groups were recorded at intervals during the experiment and are shown in Table 3 . From Table 3, . ft can be seen that average weights of the 3 groups were closely comparable throughout the experiment. The weight records thus reveal no evidence for toxicity of any of the treatments and support comparability of findings in respect t yields of tumors. Except for 2 missing or eaten animals in each group, all animals were examined by necropsy. At necropsy, gross findings were recorded. Histopathologic diagnoses of all tumors were mode by our consultant in pathology, Harold J . Sobel, M .D . No mesotheliomas were found. Malignorit tumors were a liposarcoma in group 2, a melanoma and 3 adrenal adenocarcinomas in Group 3 . Benign adrenal tumors (adenomas) were found in 3 animals in Group ! , 1 animal in Group 2 , and 3 animals In Group 3 . In addition, a hepatoma and a benign tumor of the spleen were found in Group 3 . We do not relate any of these tumors to treatment. AH tumors found in this experiment have been reported as spontaneous tumors in hams tersi7' 8' . After single Intrapleural Injection of male LVGsLAK hamsters in groups of about 50 hamsters each, we have previously reported mesotheliomas in 8 to 10 animals on a 25 mg dose of chrysotfle, in 4 animals on a 10 mg dose of chrysotile(3). O f 27 mesotheliomas in hamsters after intrapleural injection of 25 mg chrysotile, th earliest was found at 151 days after injection, 6 within 372 days and !5 within 500days(3). In the present experiment, no mesotheliomas and no other tumors related to treatment were found in any animals at either the 25 mg or 10 mg do*se of CPS-275-1. We conclude that, under the conditions 1 of this experiment, CPS-275-1 pfoved non-carcinogenic. These findings have been reported in part in a publication now in p r e s s i . . ...WES001050 TABLE I Rating O f Lungs For Pleural Adhesions. 0 = no adhesions Sample CPS-275-1 Dose? 25 mg 1 = minimal adhesions Sample CPS-275-l Dose: 10 mg - 4 = extensive adhesions 0.9% NaCl (Control) Hamster No., N o. days after injection Rating of adhesions 3 92 ` 15 92 59 92 56 92 57 92 29 // 30 76 49 55 46 55 13 49 Average 2 2 2 1 1 ' 1 2 1 1 1.4 1-iamster o. N o. days after injection Rating ' of adhesions 17 ' 93 50 84 45 83 48 82 9 82 6 81 22 81 40 81 47 80 38 80 36 76 27 74 28 71 26 71 31 69 20 53 Average 1 1 1 1 . 1 1 1 1 1 0 0 1 * 2 1 1 - ] 0.9 Hamster No. N o . days after injection Rating of adhesions 49 37 35 13 . 11 20 15 16 18 46 89 ~0 84 0 75 0 75 . 0 74 0 71 0 71 0 71 0 71 0 70 0 Average 0 WES001051 TABLE 2 .TESTS FOR CARCINOGENICITY OF CPS-275-1 Eac h n u mb e r r e f e r s to an i n d i v i d u a l ha ms t e r and is t he n u mb e r of days b e t w e e n I n j e c t i o n and d e a t h . Group 1 25 mg CPS- us : S2 K 183 154 rz li'- KNM 3 * 223 723 22? ,, 2U UNM 239 2?S 2?S 356 32S 320 323 327 333 K 323 354 243 349 375 374 357 391 427 435 443 471 454 489 496 500 Adrenol 505 5W 518 518 527 Adenoma 530 K 550 K AdreneJ Aeenomo 533 578 K Adrenal Adenoma 578 ' 597 500 Kl 6C0 KE 6COX6 604 K6 Group 2 10 mg CPS-275-1 Group 3 Saline Control 174 175 187 1SS 194 222 2*8 NNE 254 N N f 25? 277 301 3 1 ? LlpdiGfCwiTl 321 334 234 344 385 333 415 422 432 436 444 454 441 .473 <82 485 ' 43? 459 K ' 493 493 495 4 95 A d ra r,a! A c e t 'o nys 5C , 510 514 541 550 577 55? 6 0 3 KE 6C0KE 6C 0 KE 603 KE 6 0 0 KE 400 K . 4 0 4 KE 4 0 4 KE * . NNc no necropsy becog* cte NNM * r o necropsy becous* missfng K killed becouse moribund KS * killed io ferminole cxperlmani 135 135 N N E 145 145 153 154 156 165 U>7 175 182 X I &3 20A M alig n an t M elonom o 223 226 259 ' 291 306 315 372 3&5 387 393 14 A erenot Adencn-o 427 45* *60 *77 477 430 <32 4S 492 496 N N 497 500 517 524 5<7 553 554 542 574 551 AAdd renal Adenocarcinoma 585 355 . 4 M KE 401 601 KE H ^oiorjd, Adrenol Adenome 601 KE A. drenal Aderocorcinomc 401 KE 4 0 2 K E .Adrenal Adenosofcincma 6 02 KE Benign Turner Spleen 402 KE PulmonoryAdancma^otis 6 02 KE .Adrenal Adenoma 4 02 KE 4 0 2 KE 4 1 0 KE 6 1 0 K WES001052 f TABLE 3 Average Body W eighs In Grams Of Hamsters In Teste For Carcinogenicity At Intervals In Days (14) After Infection. . Group 1 2 3 Start 94 gms 95 " ' 92 " 100 Dgys 158 gms 154 * 147 " 200 Days 159 gms 166 * 149 " 300 400 500 560 Da>* Days Days Days 169 gms 168 gms 158 gms 157 gms 161 .167 " 159 " 157 * 159 159 " 152 154 WES001053 REFERENCES 'Sm ith, W. E ., L. M iller, R.E. Elssser and D .D . Hubert. Tests for carcinogenicity of asbestos. Ann New York Acad. S ei. 132 (Art 1);456-488, 1965. 2 Smith, W .E. and D .D . Hubert. The intrapleural route as a means for estimating carcinogenicity. Pp. 93--101 in Experimental Lung Cancer. Carcinogenesis and Bioassays. Ed. by E. Karbe & .J.F. Park. Springer-Verlag, Berlin/New York 1974. 3 Smith, W .E. Experimental studies on biological effects of tremolite talc on hamsters,* Pp. 43-48 in Proceedings of the Symposium on Talc. Washington, D .C. Information Circular 8639. A . Goodwin, comp. U.S. Bureau of Mines. 1974. 'Smith, W .E. , D .D . Hubert, H .J. Sobet and E. M arquet. Biologic tests of tremolite in hamsters. In Conference on Occupational Exposures to Fibrous and Particulate Dust and Their Extension into the Environment. Washington, D .C ., Dec. 5-7, 1977. Society for O ccup. and Environ. Health. In press. "'Sobel, H .J ., E. Marquet, W .E. Smith and D .D . Hubert. Asbestos-induced mesotheliomas in hamsters: similarities to human mesotheliomas and presence of type C virus particles. Federation Proc. 37:A100, 1978. KChurg, J . and A . Prado. A rapid Mallory trichrome stoin (chromotrope-aniline blue). A .M .A . Arch. Path. 62:505-506, 1956, 'Sfchuk, G ., J . G . Fortner, B.K. D erand A.B. Russfield. Spontaneous tumorgenesis in hypophysectomized Syrian (Golden) hamsters. Cancer Res 26:2154-2164, 1966. * Pour, P. et a l. Spontaneous tumors and common diseases in two colonies of Syrian hamsters. J . Nat*! Cancer Inst. 56:931-935, 937-948 , 949-961, 963-974, 1976. WES001054-