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Logo of nihpaAbout Author manuscriptsSubmit a manuscriptHHS Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
 
Transplantation. Author manuscript; available in PMC 2010 November 12.
Published in final edited form as:
Transplantation. 1992 February; 53(2): 496–498.
PMCID: PMC2979306
NIHMSID: NIHMS242332

INHIBITION OF LIVER, KIDNEY, AND INTESTINE REGENERATION BY RAPAMYCIN1,2

For a decade, liver transplant recipients have been treated with cyclosporine, a drug with modest hepatotoxicity (1). Concern that CsA might inhibit hepatic regeneration or the ability of the transplanted liver to adjust its size to that of the recipient prompted studies by Makowka et al. (2) and others (35), which showed that regeneration actually was enhanced. A newer unrelated immunosuppressive agent, FK506, has the same properties (5). In addition, these 2 drugs have other actions that are collectively called hepatotrophic. The increase in hepatocyte replication that is caused by portacaval shunt in dogs is more than doubled by intrahepatic infusion of either of these drugs via the tied-off central portal vein, and the expected atrophy and organelle disruption is prevented (6, 7). Direct experimentation in nude rats has ruled out immune modulation of lymphocytes and NK cells as an explanation (8).

These observations have contributed to a hypothesis that CsA and FK506 modify signal transduction in a variety of cells, not limited to those of the immune system (7, 9, 10), and including signaling pathways associated with growth control. If this were true, it would explain the remarkable spectrum of adverse as well as desired effects of these agents. Important elements in this hypothesis included the discovery that the cytosolic receptors for FK506 and rapamycin (FK506-binding protein, FKBP) and cyclosporine (cyclophilin) are distinct proteins that exhibit peptidyl-prolyl cis trans isomerase (rotamase) activity (11, 12) and that these small-molecular-weight cytoplasmic proteins are in virtually all cells in the body, as well as in the nuclei of some (13, 14).

How these 3 drugs (cyclosporine, FK506, and rapamycin) block or modify signaling pathways (15) is not understood. Rotamases have been shown to facilitate protein folding (16, 17) and catalyze the interconversion of rotamers of peptidylprolyl bonds in vitro (18, 19), but the inhibition of this enzymatic activity is not related to the action of these drugs on T lymphocytes (20). The fact that signal transduction in T-lymphocytes is blocked in different ways by FK506 and rapamycin (20, 21) is of great interest, particularly because these 2 drugs are chemically related and have the same binding site (FKBP). Consequently, the hepatotrophic qualities of rapamycin were determined with the same dog and rat test models as had been used previously to test CsA, FK506 (27), and recombinant FKBP (22). In addition, the influence of rapamycin on renal and intestinal regeneration was determined. The results with rapamycin were profoundly different from those observed previously with the other 2 drugs.

The dogs underwent a completely diverting portacaval shunt. Immediately afterwards, an infusion catheter was inserted into the tied-off left portal vein for pump-driven constant infusion of the rapamycin over the next 4 days in the doses shown in Table 1. At the end of 4 days, the animals were injected with 0.2 MCi/kg intravenous 3H thymidine (New England Nuclear, Boston), and killed 2 hr later. Specimens were obtained for comparison of the hepatocytes in the left (infused) and right (not infused) liver lobes using previously described morphometric and autoradiographic techniques (6, 7). There was no effect (Table 1) on the hepatocyte atrophy and the increased hepatocyte renewal that are characteristic after Eck fistula and that are prevented both by cyclosporine (6) and by FK506 (7).

Table 1
Hepatocyte size and mitosis in dogs after ECK fistula with continuous intraportal infusion of rapamycin

For the regeneration experiments, fasted adult male inbred Fisher 344 rats weighing 180–200 g (Hilltop Lab Animals Inc., Scottsdale, P A) were assigned to groups, given water and food for 4 days, and treated daily with vehicle or rapamycin. On the fourth day, between 0900 and 1030, groups 2–5 underwent 70% hepatectomy under light ether anesthesia (5). Rats of groups 7 and 8 had right nephrectomy (2), and those in group 10 and 11 had a 40% resection of small intestine excluding the jejunum. Twenty-four hr after the operations or control period, 50 MCi-3H-thymidine was administered by intraperitoneal injection. The rats were killed 2 hr later by guillotine. Hepatic DNA synthesis and liver mitosis, kidney DNA synthesis and small bowel DNA synthesis were performed as described previously in cyclosporine and FK506 experiments (5, 23).

Hepatic regeneration was significantly inhibited by RPM with a dose relationship so that at 1.0 mg/kg the normal response was virtually eliminated (Table 2). Regeneration of the kidney and intestine was significantly inhibited with the intermediate dose of 0.3 mg/kg/day (Table 3).

Table 2
Effect of preoperative intramuscular rapamycin for 4 days on rat liver regeneration
Table 3
[3H]-thymidine incorporation in kidney and small intestine from normal, nephrectomized, or small intestine–resected rats treated or not treated for 4 preoperative days with rapamycin (0.3 mg/kg/day i.m.)

These results add to the significance of the previous demonstration that rapamycin affects T-lymphocytes differently than FK506 (20, 21) and lend support to the view of rapamycin as an inhibitor of growth factor receptor-associated signaling pathways. Instead of promoting liver regeneration and having the other hepatotrophic qualities common to FK506 and cyclosporine, rapamycin was antihepatotrophic. In rats submitted to hepatectomy, it inhibited instead of augmenting liver regeneration and in dogs submitted to Eck fistula it did not prevent the hepatocyte atrophy and organelle disruption that are caused by this operation but prevented by cyclosporine(6) and FK506 (7). The observations in rats constitute the first physiologic evidence that the immunophilin network that is thought to be pleiotropic in its metabolic, immunologic, and growth control actions may be involved in countervailing (opposite effects) regulatory processes. Regeneration of the kidney and intestine was also retarded by RPM administration, indicating that the growth-inhibitory effect of RPM is not liver-specific.

This suggests that rapamycin, a profound immunosuppressant, also should be examined as an antitumor agent. The implications are obvious in general and in transplantation in particular. Efforts to treat primary hepatic malignancies by total hepatectomy and liver transplantation have been plagued by a high rate of tumor recurrence. There might be a possibility of treating these patients or others whose extirpative procedures have been done for the reason of malignancy (24) with a drug that could prevent both rejection and tumor cell replication. Striking antineoplastic properties of rapamycin against a variety of mouse tumors (25) and against human tumors transplanted into nude mice (26, 27) have been described.

Footnotes

1This work was supported by Research Grants from the Veterans Administration and by Project Grants Nos. DK-29961 and GM-38627 from the National Institutes of Health, Bethesda, Maryland.

2The rapamycin was a gift from Dr. Joseph Y. Chang of the WyethAyerst Research Company, Princeton, New Jersey.

References

1. Klintmalm GBG, Iwatsuki S, Starzl TE. Cyclosporin A hepatotoxicity in 66 renal allograft recipients. Transplantation. 1981;32:488. [PMC free article] [PubMed]
2. Makowka L, Svanas G, Esquivel C, et al. The effect of cyclosporine on hepatic regeneration. Surg Forum. 1986;37:352. [PMC free article] [PubMed]
3. Kahn D, Lai HS, Romovacek H, Makowka L, Van Thiel D, Starzl TE. Cyclosporin A augments the regeneration response after partial hepatectomy in the rat. Transplant Proc. 1988;20 (suppl 3):850. [PMC free article] [PubMed]
4. Kim YI, Caine RY, Nagasue N. Cyclosporine A stimulates proliferation of the liver cells after partial hepatectomy in rats. Surg Gynecol Obstet. 1988;166:317. [PubMed]
5. Francavilla A, Barone M, Todo S, Zeng Q, Porter KA, Starzl TE. Augmentation of rat liver regeneration by FK 506 compared with cyclosporine. Lancet. 1989;1:1248. [PMC free article] [PubMed]
6. Mazzaferro V, Porter KA, Scotti-Foglieni CL, et al. The hepatotrophic influence of cyclosporine. Surgery. 1990;107:533. [PMC free article] [PubMed]
7. Starzl TE, Porter KA, Mazzaferro V, Todo S, Fung J, Francavilla A. Hepatotrophic effects of FK506 in dogs. Transplantation. 1991;51:67. [PMC free article] [PubMed]
8. Francavilla A, Starzl TE, Barone M, et al. Studies on mechanisms of augmentation of liver regeneration by cyclosporine and FK 506. Hepatology. 1991;14:140. [PMC free article] [PubMed]
9. Starzl TE, Fung JJ, Todo S. Contempo 90: transplantation. JAMA. 1990;263:2686. [PMC free article] [PubMed]
10. Schreiber SL. Chemistry and biology of the immunophilins and their immunosuppressive ligands. Science. 1991;251:283. [PubMed]
11. Harding MW, Galat A, Uehling DE, Schreiber SL. A receptor for the immunosuppressant FK506 is a cis-trans peptidyl-prolyl isomerase. Nature. 1989;341:758. [PubMed]
12. Siekierka JJ, Hung SHY, Poe M, Lin CS, Sigal NH. A cytosolic binding protein for the immunosuppressant FK 506 has peptidyl-prolyl isomerase activity but is distinct from cyclophilin. Nature. 1989;341:755. [PubMed]
13. Marks WH, Harding MW, Handshumacher R, Marks C, Lorber MI. Cyclophilin, the cyclosporine receptor: immunochemical distribution in normal mammalian tissues. Transplantation. 1991;52:340. [PubMed]
14. Ryffel B, Woerly G, Greiner B, Haendler B, Mihatsch MJ, Foxwell BMJ. Distribution of the cyclosporine binding protein cyclophilin in human tissues. Immunology. 1991;72:399. [PubMed]
15. Hultsch T, Rodriguez JL, Kaliner MA, Hohman RJ. Cyclosporin A inhibits degranulation of rat basophilic leukemia cells and human basophils. J Immunol. 1990;144:2659. [PubMed]
16. Fischer G, Bang J. The refolding of urea-denatured ribonuclease A is catalyzed by peptidyl-prolyl cis-trans isomerase. Biochim Biophys Acta. 1984;828:39. [PubMed]
17. Tropschug M, Wachter E, Mayer S, Schonbrunner ER, Schmid FX. Isolation and sequence of an FK506-binding protein from N. crassa which catalyses protein folding. Nature. 1990;346:674. [PubMed]
18. Fischer G, Wittmann-Liebold B, Kiefhaber T, Schmid FX, Lang K. Cyclophilin and peptidyl-prolyl cis-trans isomerase are probably identical proteins. Nature. 1990;337:476. [PubMed]
19. Albers MW, Walsh CT, Schreiber SL. Substrate specificity for the human rotamase FKBP: a view of FK 506 and rapamycin as leucine-(twisted amide)-proline mimics. J Org Chem. 1990;55:4984.
20. Bierer BE, Mattila PS, Standaert RF, et al. Two distinct signal transmission pathways in T lymphocytes are inhibited by complexes formed between an immunophilin and either FK 506 or rapamycin. Proc Natl Acad Sci USA. 1990;87:9231. [PubMed]
21. Dumont FJ, Staruch MJ, Koprak SL, Melino MR, Sigal NH. Distinct mechanisms of suppression of murine T cell activation by the related macrolides FK 506 and rapamycin. J Immunol. 1990;144:251. [PubMed]
22. Starzl TE, Schreiber SL, Albers MW, Porter KA, Foglieni CS, Francavilla A. Hepatotrophic properties in dogs of human FKBP, the binding protein for FK506 and rapamycin. Transplantation. 1991;52:751. [PMC free article] [PubMed]
23. Francavilla A, Barone M, Starzl TE, et al. FK 506 as a growth control factor. Transplant Proc. 1990;23:90. [PMC free article] [PubMed]
24. Starzl TE, Todo S, Tzakis A, et al. Abdominal organ cluster transplantation for the treatment of upper abdominal malignancies. Ann Surg. 1989;210:374. [PubMed]
25. Eng CP, Sehgal SN, Veznia C. Activity of rapamycin (AY-22,989) against transplanted tumors. J Antibiot (Tokyo) 1984;37:1231. [PubMed]
26. Houchens DP, Ovejera A, Riblet SM, Slagel DE. Human brain tumor xenografts in nude mice as a chemotherapy model. Eur J Cancer Clin Oncol. 1983;19:799. [PubMed]
27. Fiebig H-H, Berger DP, Winterhalter BR, Plowman J. In vitro and in vivo evaluation of US-NCI compounds in human tumor xenografts. Cancer Treat Rev. 1990;17:109. [PubMed]