Intestinal transplantation is reserved for the treatment of patients with complicated intestinal failure and is frequently their last resort for survival. This is always the case with the more complex liver-intestinal and multivisceral transplants, which are life-saving procedures for patients with extensive abdominal catastrophes. Acute rejection has been recognized as the principal obstacle to successful clinical intestinal transplantation from the outset. Successful outcome was impossible with azathioprine and rare with cyclosporine immunosuppression (1–4), but results have improved significantly with tacrolimus (Tac) (Prograf, Fujisawa Pharmaceuticals, Tokyo, Japan) immunosuppression (5).
Advances generally meant the addition of more powerful drugs, including induction with OKT3 and daclizumab (Zenapax, Roche Laboratories, Palo Alto, CA) and the addition of adjuvant therapy with cyclophosphamide, mycophenolate mofetil, and, more recently, sirolimus (Sir) (Rapamune, Wyeth-Ayerst, Pearl River, NY) (6–10). This powerful, continuous immunosuppressive therapy has led to an increased risk of overimmunosuppression and drug toxicity, both short and long-term.
Efforts have also been made to facilitate graft acceptance through immunologic manipulations. Excellent preliminary results have been reported when donor bone-marrow infusions have been combined with irradiation of the intestinal graft (11,12). Long-term and more short-term data are eagerly expected to confirm the early findings and dispel some uneasiness about the long-term effects of radiation on the intestinal graft.
Alemtuzumab (Campath-1H, Berlex Laboratories, Montville, NJ) is a humanized monoclonal antibody directed against the CD52 antigen, a pan T, B, and natural killer cell, and monocyte marker. It rapidly depletes lymphocytes, monocytes, and other cells without affecting neutrophils and hematopoietic stem cells. The depleted cells begin to reemerge gradually during a period of 6 months but do not return to baseline levels.
This activity is believed to prevent an aggressive lymphocytic immune response after transplantation and allow a more gradual engagement of the host immune system under low conventional immunosuppression.
This slow engagement is believed to encourage the development of “prope” (almost) tolerance (13,14). This term implies a compromise but may well be a step toward the ultimate goal of achieving true immune tolerance.
Encouraging results have been reported in 31 kidney recipients treated with alemtuzumab and half-dose cyclosporine (13). Similar conclusions were reached by a separate group of investigators who tried to induce tolerance with a carefully planned preoperative regimen of alemtuzumab in recipients of living-related kidneys. They observed that although tolerance was not achieved, the patients required small amounts of (Sir-based) immunosuppression (15). We hypothesized that alemtuzumab combined with low doses of Tac may reduce the number and intensity of acute cellular rejections and allow graft acceptance with lower maintenance immunosuppression in intestinal transplantation.
To our knowledge, this is the first study of the combination of alemtuzumab and Tac. We present our updated results in this report.
This was a prospective study that included all the adult recipients of isolated and composite intestinal grafts performed at our center since January 2001. The follow-up was through July 1, 2002. Informed consent was obtained before enrollment in the study, and the institutional review board of the University of Miami approved these studies.
Alemtuzumab was administered in four doses (0.3 mg/kg intravenously): preoperatively, at the completion of the transplant, and on posttransplant days 3 and 7. Tac levels were maintained between 10 and 15 ng/dL with enteral or intravenous administration. No baseline steroids were given. Tac was substituted with Sir (levels 5–15 ng/dL) for Tac-related complications. Peripheral blood lymphocytes and monocytes were monitored daily during the patients’ hospitalization and monthly after discharge. Surgical techniques, monitoring of the graft, and immunoprophylaxis were performed according to our existing protocols (6–8).
Peripheral blood leukocytes and their subsets were enumerated using a standard method with a Cell-Dyn instrument (Abbott, Laboratories, Abbott Park, IL).
Suspected or mild rejections were treated with steroid boluses (methylprednisolone [SoluMedrol; Pharmacia and Upjohn Co., Peapack, NJ] 0.5 g three times/day) or courses of low-dose steroids (methylprednisolone 20–40 mg/day) until reversal occurred. Moderate rejections were treated with steroids or OKT3. Severe rejections were treated with OKT3 (5 mg/day for 10–14 days).
Between January 9, 2001, and July 1, 2002, 21 consecutive adult patients received 24 grafts (Table 1): 14 intestinal (11 primary and three retransplants), one liver-intestinal, and nine multivisceral (five with and four without a liver graft). We analyzed the results of the patients who had undergone a posttransplantation follow-up of more than 2 months. Sixteen patients were followed up for 2.4 to 16 months (median 9.3 months); twelve of them are alive at the time of this report, and all 12 grafts are functioning normally.
The administration of alemtuzumab was well tolerated by all patients without any significant side effects.
Lymphocytes and monocytes in peripheral blood were swiftly depleted after the administration of alemtuzumab. Lymphopenia persisted beyond 4 months (Fig. 1), whereas monocytes were replenished approximately 2 months after induction with alemtuzumab (Fig. 2).
There were seven deaths in this study of patients receiving alemtuzumab (three in the immediate postoperative period and four during the follow-up); however, none of the deaths seemed directly related to a toxicity of the drug.
Two recipients of multivisceral grafts (patients 13 and 14), both intensive-care-unit bound, became unstable during transplantation. Patient 13 developed pancreatitis of the multivisceral graft and sepsis and died the first day posttransplantation, and patient 14 died of sepsis the seventh postoperative day. The cause of sepsis in patient 13 was found to be a fungus most likely related to a central venous line, but the cause could not be identified in patient 14. These two patients were significantly debilitated; this, combined with the rapidity of their demise and manner of death, lessens the likelihood of any toxic effect of the drug.
A third patient (patient 11) developed hemolytic uremic syndrome (HUS) and was converted to Sir on the sixth posttransplant day. He died as the result of pulmonary embolism on postoperative day 9. His isolated intestinal graft was normal at the time of death. We do not believe that systemic thrombi were formed as a result of this episode, although this remains unproven.
Two deaths were the result of severe rejection. Patient 19 experienced a sudden severe rejection on postoperative day 45 that was resistant to treatment with steroids and OKT3. The graft was explanted, and although the patient received a second small bowel graft, he developed severe rejection again and died 87 days after his initial transplant. Patient 10 developed severe rejection on postoperative day 80 and died 9 days later despite treatment. He could not undergo retransplantation because of the complete lack of venous access. Patient 2 developed recurrent episodes of ileus because of opiate abuse and later died of an opiate overdose 321 days posttransplant. Patient 1 died of complications of ileostomy closure 1.5 years after transplantation. This death was totally unexpected.
Conversion to Sirolimus
Three patients were converted to Sir: patients 9 and 11 for HUS and patient 10 for Tac nephrotoxicity. One of these patients (patient 9) is alive and well 8.5 months after a multivisceral transplant.
Frequency of Acute Rejection
As mentioned previously, patients 10 and 19 experienced severe rejection. Patients 1 and 15 developed moderate rejection 3 months posttransplantation, and patient 20 developed moderate rejection 1 month posttransplantation. Patient 1 was successfully treated with OKT3, and patients 15 and 20 were successfully treated with steroids. Patients 2, 3, 4, 6, 8, 9, and 12 were suspected or found to have mild rejection 5 to 294 days after transplantation and were successfully treated with steroids. Patients 9, 15, and 20 remain on maintenance steroids.
The intestinal component of the only liver-intestinal graft in this series (patient 5) developed severe diffuse arteritis and inflammation of the mesentery, which caused persistent intestinal obstruction. There was no rejection detectable in repeated endoscopies and mucosal biopsies. There was no rejection or vasculitis in the liver graft. No infectious source or technical imperfections could be found. The intestinal component of the first graft was replaced with another cadaveric graft 18 days after the original transplant. Immunosuppression given after retransplantation consisted of Tac and baseline steroids without induction. The patient is doing well 10 months after the transplant, and both grafts are functioning normally without any evidence of rejection.
Patient 1 developed cytomegalovirus enteritis after treatment of moderate rejection with OKT3 and responded to treatment with ganciclovir (Cytovene, Roche Laboratories) and cytomegalovirus immune globulin (CytoGam, MedImmune Inc., Gaithersburg, MD).
There were four fungal septicemias most likely related to central venous access lines; one of them occurred at the time of transplantation and was fatal (patient 13). The rate of fungal infections in the alemtuzumab-treated patients was not significantly different from the rate we have observed in our small bowel-transplant patients (A.G. Tzakis, unpublished data, 2002). Therefore, it is unlikely that the profound immunosuppression in the alemtuzumab-treated patients predisposed the recipient to any higher rate of opportunistic infections compared with our other immunosuppressive protocols.
Patients 9, 11, and 12 developed HUS. Patient 11 died of pulmonary embolism on the ninth postoperative day. The syndrome resolved in the other two patients: The immunosuppression was converted to Sir in patient 9, and the Prograf dose was lowered in patient 12.
Our preliminary results show that, as with low-dose cyclosporine and Sir in kidney transplants, alemtuzumab seems to ease the way of the intestinal grafts into the host when given with low-dose Tac.
Tac was used as maintenance monotherapy at levels that were half or less than usually used in intestinal transplantation. No maintenance steroids or other immunosuppression was given.
The combination of alemtuzumab with the low dose of Tac seemed to protect the grafts from acute rejection more effectively than has been reported with other commonly used regimens (7,9,10). Rejections were generally mild, requiring only temporary steroids. In the 16 patients that were followed up for an average of 9.3 months, only three presented with significant rejections requiring OKT3 treatment, resulting in graft loss in two of them. This is in sharp contrast with the existing experience in which as many as 20% of the grafts are lost to acute rejection (10,12).
Of note, all three serious rejections were seen between the second and third postoperative month, coinciding with recipient hematopoietic cell repopulation. These data indicate that monitoring should be intensified during this time period.
There is a profound depletion of leukocytes after alemtuzumab treatment, and this is reflected by a concomitant reduction in the “normal” levels of tissue leukocytes seen within the early posttransplant biopsies. Thus, there may be only subtle histologic changes that occur during mild acute rejection. The histologic diagnosis of rejection was unambiguous in the severe and moderate rejections. In this regard, it may eventually be necessary to further subdivide the milder histologic variants among the alemtuzumab-treated individuals. However, this is only a potential issue, and many more patients will have to be evaluated before this can be appropriately considered.
Local factors may also play a role in the loss of the intestinal graft; for example, patient 5 showed that the contemporaneously transplanted liver from the same donor was not affected by the acute rejection. Regardless, we considered the intestinal loss as an immunologic event, although the mechanism is not understood.
Four patients (5, 7, 16, and 17) of the 16 that were followed up for an average of 9.3 months had no rejection of any type, and three of them (patients 7, 16, and 17), recipients of isolated small bowel grafts, never received steroid treatment.
The incidence of opportunistic infections seen here was low, although longer follow-up is needed for a more accurate assessment. Central venous access-related infections tend to be technical in origin and are believed to have little association to the immunosuppression.
We are concerned with the three cases of HUS seen in this series. HUS has been described in intestinal grafts with Tac immunosuppression (16,17). It is interesting that paroxysmal nocturnal hemoglobinuria (PNH) has been described as a complication of alemtuzumab treatment in rheumatoid arthritis and non-Hodgkin’s lymphoma (18,19). It seems to be related to recipient-derived CD52(−) T and B cells possibly because of a mutation or an alteration in a regulatory factor. Because there are similar clinical features between HUS and PNH, there is the possibility that there may be related pathophysiologic mechanisms involved between these processes, and this is enhanced when alemtuzumab targets these particular antigenic specificities. In PNH, a somatic mutation of the PIG-A gene in a hematopoietic stem cell is responsible for the production of cells deficient in the surface antigens that are attached to the cell membrane by the glycosyl-phosphatidyl-inositol (GPI) link. Alemtuzumab is directed against the GPI-linked CD52 antigen. In a study involving patients with chronic lymphocytic leukemia who received alemtuzumab therapy, the author supports a dual theory for the development of these GPI-deficient T-cell clones, stating that the patient has to have a preexisting PIG-A mutation and a positive selection factor (alemtuzumab treatment) that enhances the survival of these GPI-deficient, CD52 antigen-negative cells (20).
Finally, our results indicate a common problem of intestinal and multivisceral transplantation: operating on patients who are too ill to survive. At this terminal stage, it is often difficult to distinguish between progressive deterioration versus an acute event (e.g., the fungemia in patient 13). A decision not to proceed with the transplant often means the death of the patient, because matching grafts are hard to find. It is equally difficult to recognize intractable drug addictions (patient 2) under these circumstances.
Three of the seven deaths (patients 11, 13, and 14) were related more to the underlying grave condition of the patient than to the transplant and immunosuppression.
Fortuitously, several patients have been able to recover and return to apparent good health after transplantation even after having been so ill.
Our results are preliminary. Longer follow-up and larger series are needed to confirm our observations.
We thank Allan D. Kirk, MD, Transplant Autoimmunity Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland, for reviewing the manuscript.
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