New Developments in Transplantation of
Allogeneic Hematopoietic Stem Cells (HSC)

Lawrence A Solberg, Jr., Ph.D., M.D.

 
Introduction

Allogeneic bone marrow transplantation, or more accurately, allogeneic hematopoietic stem cell (HSC) transplantation, is a clinical treatment in which HSC capable of reconstituting the immune system and blood production (hematopoiesis) are transferred from one individual to another. The donor and recipient may or may not be related.

Principles underlying HSC began to develop after World War II showed humanity the deadly effects of radiation. By the mid 1950's, investigators working with mice had discovered that lethally irradiated animals could be protected from death by the transfer of bone marrow cells and that some form of tolerance of the recipient for the donor was involved. Other critical pieces of scientific underpinning were learned subsequently from experiments with mice and dogs. The main principles that emerged were that animals could survive bone marrow being infused intravenously; that marrow cells could produce a state in which an immune attack was directed against the recipient; that genetic factors controlled the extent and severity of the graft-versus-host effect; that irradiation and cyclophosphamide could reduce the rejection by the recipient of the infused marrow cells; and that methotrexate could ameliorate graft-versus-host disease without causing loss of the graft.1

In this article, I will review how allogeneic HSC transplantation is done; the outcomes of this treatment; the main problems involved with the technique and new approaches addressing those problems; and finally, the post-transplant complications that might be encountered by almost any health-care provider in our community.

Allogeneic HSC Transplantation

The treatment starts with a careful study of the HLA system of the recipient and proposed related or unrelated donors. In a companion article, my colleague Trina Genco, Ph.D. describes how HLA typing is done in the setting of allogeneic HSC transplantation. Another article will describe the critical contributions of the National Marrow Donor Program (NMDP) which is celebrating the 10,000th unrelated allogeneic HSC transplant it has facilitated!

If a matched donor is available, and if the recipient is medically healthy enough to undergo transplantation, HSC are then collected from the donor. The recipient is treated with chemotherapy or a combination of radiation and chemotherapy before receiving the donor HSC (this is referred to as conditioning in the jargon of the field). The conditioning regimen has two objectives: to suppress the recipient's immune system enough so that the new donor allogeneic cells will not be rejected and to kill malignant cells.

After receiving by intravenous infusion the donor stem cells the patient is supported through an acute phase in which the blood counts initially drop and then recover. The patient usually experiences regimen related side effects such as infection, mucositis, and occasionally may develop severe hepatic, or pulmonary damage. After the transplant, about two-thirds of recipients experience some grade of acute or chronic graft versus host disease (AGVHD or CGVHD). The GVHD can be severe enough to cause death or disability in one-half of the patients who get GVHD.

Patients also can develop a variety of long-term complications of the transplant procedure.

Collection of HSC

Allogeneic HSC have been collected primarily from normal related or unrelated donors through bone marrow harvest. In this procedure, donors are anesthetized, placed in a supine position, and multiple small punctures are made bilaterally into the pelvis near the posterior iliac crests. Approximately one liter of marrow is aspirated. The main risks are those of anesthesia and post-donation pain. Interestingly, 98% of normal volunteers in the NMDP who have donated bone marrow in this way would volunteer again.

An increasingly popular way of collecting allogeneic HSC is from the peripheral blood2 using apheresis equipment. The use of umbilical cord blood as a source of HSC is also steadily increasing.3 Figure 1 shows the increase in the use of HSC from peripheral blood and from cord blood donors reported to the International Bone Marrow Transplantation Registry (IBMTR).

In the apheresis process, normal donors are given subcutaneous injections of a growth factor, granulocyte colony-stimulating factor (G-CSF) that causes normal HSC to move out of the marrow and into the peripheral blood. Through intravenous lines in each arm, small amounts of blood are continuously removed from the donor and processed by the apheresis system. To selectively collect peripheral blood HSC, the apheresis equipment harvests the mononuclear cell layer. All other components of the blood are returned continuously, e.g. the red cells, plasma, and platelets. The apheresis procedure used is similar to that used for normal volunteers who donate single-donor platelets- except the normal platelet donors are not given growth factor.

For normal donors, the short-term safety profile of G-CSF seems acceptable, given the extensive use of this agent in patients receiving cancer chemotherapy and in patients with chronic neutropenia. There are acute side-effects that can occur, however, including the potential risks of marked leukocytosis to more than 70,000 per cubic millimeter. G-CSF dose reduction is used to maintain leukocyte counts below this level. Transient post donation cytopenias involving granulocytes, lymphocytes, and platelets may occur and are at least partly related to the leukapheresis procedure itself. These are generally asymptomatic and self-limited; follow-up blood counts are not necessarily required.

The problem of graft failure from inadequate HSC numbers has essentially vanished due to the availability of a much more quantitative way of measuring the HSC in peripheral blood collections. Hematopoietic stem cells have an antigen called CD-34 expressed on their surfaces. The availability of monoclonal antibodies to CD-34 has enabled the use of flow cytometry to enumerate the numbers of CD-34 positive stem cells has become a reliable way to monitor the collection of adequate numbers of hematopoietic stem cells4. Before, attempts to quantitate stem cell numbers with engraftment or other end-points involved laborious assays of colony forming cells - which were difficult to do and typically not well reproduced from lab to lab.

Progress in Sources of HSC

The need for more donors other than just fully-matched related donors is exemplified by CML, one of the leukemias most curable by allogeneic transplantation. CML is the most frequent indication for allogeneic bone marrow transplantation worldwide. Of over 5,725 recipients of HLA-identical sibling transplants done between 1991 and 1997 and reported to the IBMTR, 3-year probabilities of survival were 67% ± 2% for 2,830 transplants performed within 1 year of diagnosis and 57% ± 3% for 1,595 patients transplanted more than 1 year after diagnosis. Unfortunately, only 30% of persons with CML have an HLA-identical sibling donor.

Unrelated donor transplants can cure CML but do have higher risks of GVHD and transplant-related mortality. The National Marrow Donor Program has been a critical organization to help develop this source of cells. It has provided unrelated donor HSC to over 2000 patients with CML. The use of umbilical cord blood stem cells is also a growing and important source of donor stem cells, particularly for pediatric populations.3

Another development that has increased the availability of related but partially matched donors has been the development of procedures that allow successful transplantation despite the presence of some degree of antigen mismatch.5 This approach relies on the use of very large number of donated cells.6 The availability of molecular genetic techniques for HLA typing has helped by increasing recognition of specific mismatches that are less dangerous than others.

So both the increasing availability of non-related donors, including those via the umbilical cord blood source, and the ability to use partially mismatched donors is increasing the number of patients who can receive allogeneic HSC.

 

Diseases and Conditions Treated

Allogeneic HSC is used to treat two large groups of diseases: malignancies and non-malignant inherited or acquired conditions (Table 1). One potent and desirable side effect of graft versus host disease is a graft versus tumor effect which can markedly reduce the risk of recurrent malignant disease7. The non-malignant group contains numerous inherited conditions of blood formation or metabolic disorders, such as Gaucher's disease, subacute combined deficiency disease, and beta-thalassemia major. For the inherited disorders of metabolism, the new graft provides enough normal macrophages and other hematopoietic and lymphoid cells to ameliorate the underlying genetic defect.

Typical Outcomes

Table 2 shows current 3 year survival rates from IBMTR data for many common conditions treated with allogeneic HSC. The International Bone Marrow Transplant Registry (IBMTR) and the Autologous Blood and Marrow Transplant Registry (ABMTR) are voluntary organizations of basic and clinical scientists. The IBMTR has collected data from over 350 institutions performing allogeneic bone marrow transplants worldwide. In 1991, the ABMTR began collecting data from centers in North and South America on transplants using autologous bone marrow and/or blood cells. More than 200 autotransplant centers now contribute data to the ABMTR.

 

New Developments

From 1970 into the 1990's, steady progress was made in extending allogeneic transplantation to patients with a variety of conditions and diseases. Progress also was made in developing prophylactic regimens to prevent graft versus host disease _ although this still remains a major problem. Between 1991 and 1998, 15-20% of all deaths after allogeneic transplantation was from GVHD. Significant improvements were made in developing drugs and treatment strategies for preventing and treating cytomegalovirus infections and for treating all the other typical infectious disease problems that can arise. Little progress has yet been made on other serious, idiosyncratic complications related to allogeneic transplantation such as veno-occlussive disease of the liver or severe lung damage.

Reducing Acute Toxicity

The major improvement I will address is the development of what has been called non-myeloablative conditioning regimens. The golden standard for conditioning has been cyclophosphamide and total body irradiation. This and similar conditioning regimens cause major acute regimen-related side-effects such as cytopenias and GI tract damage.

The idea of significantly reducing the intensity of treatment, so that patients do not have low blood counts and do not even need hospitalization, is a major breakthrough. Slavin8 first used less cytotoxic immunosuppressive therapy to suppress the recipient immune system and to establish host-versus-graft tolerance for engraftment of donor inununohematopoietic cells. The original non-myeloablative regimen included fludarabine, anti-T-lymphocyte globulin and low doses of busulfan and was well tolerated without severe procedure-related toxicity. This approach does not have the direct regimen related toxicity on the recipient, e.g. the marked reduction in normal granulocytes and platelets that typically occur.8,9 Instead, the infused donor cells first establish a mixed-chimerical state, in which donor lymphoid cells co-exist with the recipient immune system, and then with infusion of more donor stem cells, full engraftment with donor hematopoiesis and lymphopoiesis is achieved.

Drugs that produce immune suppression but which have not typically been used in bone marrow transplant conditioning regimens in the past are being used and tested in many new combinations to achieve the right amount of host immune suppression with the minimum amount of toxicity. Drugs being used include fludarabine, cladribine (2CdA), melphalan, busulphan and thiotepa. Other modalities such as anti - thymocyte globulin, total body irradiation and cyclophosphamide are being used at lower than usual dosages. This approach is being actively investigated at many centers.

The final value of this approach has yet to be established because the toxicities of AGVHD and CGVHD remain unaltered. Nevertheless, a worthwhile sustained reduction in acute toxicity and hospitalization would add greatly to the use of allogeneic HSC transplantation.

The reader may have seen a report in a recent New England Journal of Medicine10 issue in which investigators at the NIH used non-myeloablative allogeneic stem-cell transplantation for metastatic renal cell carcinoma. The authors based their study on the two well known observations that allogeneic transplantation can result in a graft versus tumor effect and that some patients with renal cell carcinoma have malignancies that seem to respond to immune suppression.

They treated nineteen patients with metastatic renal-cell carcinoma using a preparative regimen of cyclophosphamide and fludarabine, followed by an infusion of a peripheral-blood stem-cell allograft from an HLA identical sibling or a sibling with a mismatch of a single HLA antigen. Cyclosporine was used to prevent graft versus-host disease.

Of the nineteen patients, nine were alive 287 to 831 days after transplantation ( the median was 402 days). Two died of transplantation-related causes, and eight from progressive disease. Metastatic renal cell cancer responded in ten patients (53 percent) with three having a complete response, and seven a partial response. The patients with complete remissions had remained in that status for 16, 25, and 27 months after transplantation. Interestingly, reduction in the size of metastases was slow, occurring a median of 129 days after transplantation. The responses often were not seen until cyclosporine was stopped and complete donor-T-cell chimerism was established. In multivariate analysis, acute graft-versus host disease was the only factor predicting a response (relative likelihood of a response, 11.0; 95 percent confidence interval, 1.4 to 98.5).

The authors concluded that non-myeloablative allogeneic stem-cell transplantation can cause regression of metastatic renal-cell carcinoma in patients who have had no response to conventional immunotherapy.

The significance of this study is that the non-myeloablative approach offers a fairly straight forward way of offering patients with immunologically sensitive solid tumors the chance to be treated with powerful, allogeneic graft-versus-tumor immunotherapy.

Treating Relapsed Disease

The treatment of malignant disease relapsing after allogeneic transplantation has also improved in the past decade. In most instances, instead of a second allogeneic transplant with all its cost, morbidity and mortality, the transplant team can collect peripheral blood lymphocytes from the original donor, using outpatient apheresis, and simply infuse them into the patient _ all as a simple outpatient procedure. This process is called donor leukocyte, or donor lymphocyte, infusion (DLI).11,12,13 In selected diseases, DLI has permanently eliminated the relapsed disease and reinstituted normal hematopoiesis. Unfortunately, this powerful graft-versus-disease response is not seen uniformly in all diseases, but the DLI approach has been particularly for patients with CML.

For CML, DLI can establish durable cytogenetic and molecular remissions, and complete donor hematopoiesis in about 80% of patients treated in chronic phase. Risks of DLI include cytopenias, generally occurring one to three months after infusion and graft-versus-host disease. DLI is now the treatment of choice for persons relapsing with CML after an allogeneic transplant.

Challenges Posed by New Drug Development Technology

One of the most successful areas in which allogeneic transplantation has been offered is for patients with chronic myelogenous leukemia (CML). About 70% of these patients are cured provided they are transplanted within one year of their diagnosis. The first complexity in managing these patients came when it was discovered that alpha-interferon can produce complete molecular remission in a small proportion of patients. What does one do then- transplant these patients or watch them? There is no right answer yet that has emerged, so most transplant centers discuss fully the risks of allogeneic transplantation with related or unrelated donors versus the risks and benefit of continued interferon therapy.

Now hypothesis-driven drug research has created a new dilemma. There is a new drug STI571 designed to specifically inhibit the abnormal protein kinase, the bcr/abl oncogene product, that is abnormal in CML. STI571 is an oral agent producing complete remissions in a significant number of patients with CML. How long will these remissions endure? Should curative allogeneic transplantation be withheld when a patient has achieved a complete remission on this agent?

Overall, this progress is good for the patient, but it increases the need for clinical trials to clarify the best strategies for advising patients how to incorporate allogeneic transplantation into their management14.

Long-term Complications

Patients who have undergone allogeneic transplantation, either as children or adults, are now living for a long time. They can develop long-term complications of their transplants that may be recognized by physicians other than the original transplant physicians15,16. Table 3 lists the major delayed complications of allogeneic stem cell transplantation.

Secondary malignancies

Patients who have had allogeneic HSC transplantation need life-long surveillance for tumors. In a study16 of 19,229 patients, the transplanted patients had an overall increased risk of cancer, observed to expected , of 2.7. The cumulative rates were 2.2% at 10 years and 6.7% at 15 years. Melanoma, cancers of the buccal cavity, liver, CNS, thyroid, bone, and connective tissue were all increased. Risk was higher for patients transplanted at younger ages and from higher doses of total-body irradiation. CGVHD and male sex were linked to buccal cancers.

In a study of 4294 patients from Seattle17, 82 patients developed a secondary malignancy, an approximately 2% overall risk. Adverse risk factors for secondary malignancies included: the use of anti-thymocyte globulin for treating acute graft versus host disease and use of total body irradiation. The relative risk for developing a secondary malignancy was increased approximately 3 x if total body irradiation was used in conditioning. The most common post-transplant malignancies in this series are shown in Table 4.

 

 

Psychosocial Effects

Patients who experience few or no acute complications of transplantation or of chronic graft versus host disease do quite well and have essentially normal activites by two years after transplant. For patients with significant complications, chronic disability, depression, and disruptions in family life can be severe. Some families experience major financial stress not just because of the costs of the transplant, but also due to the interruption of work by the patient and care-givers within the family. It is important to provide counseling, rehabilitation, and long-term support to patients who have such chronic problems after transplantation.

Airway and Pulmonary Disease

Airway and pulmonary diseases are the most devastating of delayed post-transplantation complications. If one has a patient with this type of condition, the prompt evaluation and treatment of pulmonary dysfunction and the rapid institution of antibiotic therapy can be life-saving. Altogether, about 10-15% of transplant patients will have significant post-transplant pulmonary disease. Testing indicating restrictive lung disease is often positive just after transplant and pulmonary function tests have been reported to improve over 3-4 years. About 10-15% of patients will
develop clinically significant obstructive pulmonary disease. In this case, aspiration or chronic sinusitis must be detected and treated aggressively. Perhaps the most feared long-term complication arises in patients with chronic graft versus host disease- bronchiolitis obliterans. In this subset, disease is usually present within two years of transplantation. The downhill course can vary- but some cases are quite aggressive. Recurrent pulmonary infections are common. In some patients immunosuppression with corticosteroid has been helpful.

Aseptic Necrosis.

Because most patients have exposure to corticosteroids to prevent or manage graft versus host disease, avascular necrosis is altogether too familiar- usually affecting the femoral or humeral heads. Many such patients will need joint replacement therapy.

Ophthalmologic Problems

Radiation and graft-versus-host disease can significantly damage the eye. Posterior cataracts begin to develop about one year after total body irradiation _ eventually affecting 20-35% of patients. GVHD can cause also keratoconjunctivitis sicca and its complications. Infections can occur leaving sequelae such as synecheiae and ectropion.

Dental Complications

Patients with CGVHD frequently have dry mouths from an immune sicca syndrome and are intolerant of acidic or spicy foods. Therefore, preventive dental care is important to reduce the incidence of cavities, and periodontal disease that occurs with increased frequency in these subjects.

Genitourinary Tract Dysfunction

Hemorrhagic cystis due to cyclophosphamide is now a rare complication. Some patients are left with some chronic renal insufficiency due to multiple insults to the kidney including radiation, cyclosporine, and antibiotics such as vancomycin. The hemolytic- uremic syndrome is a particularly feared complication of immunosuppression with cyclosporine.

Endocrine Dysfunction

Conditioning regimens with chemotherapy only, for example busulfan and cyclophosphamide have not produced hypothyroidism. Total body irradiation does cause hypothyroidism. Within the first year post-transplant, asymptomatic compensatory hypothyroidism begins in 30-50% and progresses to overt hypothyroidism over the next several years. All patient with overt hypothyroidism should be treated but the benefit of treating compensated hypothyroidism is unclear. Thyroid function tests should be made annually or earlier if symptoms develop.

Fertility

Alkylating agents such as cyclophosphamide and busulfan impair female reproductive function. The patient's age is important is resisting this effect, e.g. 5.2 gram of cyclophosphamide can produce ovarian failure in 40 year-old women while those younger than 25 can tolerate up to 20 grams. Cyclophosphamide is toxic to the testicular germinal epithelium with a dose-response effect- age does not appear to play a role. Men who receive lower doses of cyclophosphamide can have a reversible oligospermia lasting one year or more. In one study, only 5 of 323 men who received cyclophosphamide and total body irradiation had normal spermatogenesis and had fathered 9 normal children.

REFERENCES

  1. Storb and Thomas in Bone Marrow Transplantation, Eds Forman, Blume, and Thomas. Boston.Blackwell Scientific Publications,Cambridge,MA. 1994. Ch 1 and 2:pp3-15.
  2. Korbling M, Chan KW, Anderlini P, et al. Allogeneic peripheral blood stem cell transplantation using normal patient related pediatric donors. Bone Marrow Transplantation, 1996: 18:885-90.
  3. Rocha V, Wagner JE Jr, Sobocinski, KA, et al. Graft-versus-host disease in children who have received a cord-blood or bone marrow transplant from an HLA identical sibling. New England Journal of Medicine. 2000. 342:1846-1854.
  4. Anderlini P, Przepiorka D, Seong C, et al. Factors affecting mobilization of CD34+ cells in normal donors treated with filgrastim. Transfusion, 1997: 37:507-12.
  5. Aversa, F, Martelli MM and Reisner Y. Use of stem cells from mismatched related donors. Current Opinion in Hematology, 1997: 4:419-422.
  6. Bornhauser M, Thiede C, Brendel C, et al. Stable engraftment after megadose blood stem cell transplantation across the HLA barrier: the case for natural killer cells as graft-facilitating cells. Transplantation.1999. 68:87-88
  7. Weiden PL, Flournoy N, Thomas ED, et al. Antileukemic effects of graft-versus-host disease in human recipients of allogeneic-marrow grafts. N Engl J Med .1979:300:1068.
  8. Slavin S, Nagler A, Naparsek E, et al.: Nonmyeloablative stem cell transplantation and cell therapy as an alternative to conventional bone marrow transplantation with lethal cytoreduction for the treatment of malignant and nonmalignant hematologic diseases. Blood 1998: 91:756-763.
  9. Sandmaier BM, McSweeney P, Yu C, et al. Nonmyeloablative transplants: preclinical and clinical results. Seminars in Oncology. 2000. 27:78-81.
  10. Childs R, Chernoff A, Contentin N, et al. Regression of Metastatic Renal-Cell Carcinoma after Nonmyeloablative allogeneic Peripheral-Blood Stem-Cell Transplantation. New Engl J Med. 2000; 343:750-758.
  11. Verdonck LF, Petersen EJ, Lokhorst HM et al. Donor leukocyte infusions for recurrent hematologic malignancies after allogeneic bone marrow transplantation: impact of infused and residual donor T cells. Bone Marrow Transplantation. 1998. 22:1057-1063.
  12. Bishop MR, Tarantolo SR, Pavletic ZS et al. Filgrastim as an alternative to donor leukocyte infusion for relapse after allogeneic stem-cell transplantation. Journal of Clinical Oncology. 2000: 18:2269-2272
  13. Keil F, Haas OA, Fritsch G et al. Donor leukocyte infusion for leukemia relapse after allogeneic marrow transplantation: lack of residual donor hematopoiesis predicts aplasia. Blood, 1997: 89:2113-3117.
  14. Lee SJ, Anasetti C, Horowitz MM and Antin JH. Initial therapy for chronic myelogenous leukemia: playing the odds. Journal of Clinical Oncology. 1998. 16:2897-2903.
  15. Socie G, Stone JV, Wingard, et al. Long-term survival and late deaths after allogeneic bone marrow transplantation. New England Journal of Medicine 1999:341:14-21.
  16. Curtis RE, Rowlings PA, Deeg HJ, et al. Solid cancers after bone marrow transplantation. New England Journal of Medicine. 1997:336:897-904.
  17. HJ Deeg in Bone Marrow Transplantation, Eds Forman, Blume, and Thomas. Boston. Blackwell Scientific Publications, Cambridge,MA. 1994. Ch 39:pp538-

November, 2000/ Jacksonville Medicine

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