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Allogeneic Stem Cell Transplant with Non-Myeloablative Conditioning


Medical editor Lorentz Brinch MD
Hematologist
Oslo University Hospital

General

An allogeneic stem cell transplantation with non-myeloablative conditioning is currently an option which is still experimental. The method and indication is therefore under constant change.

Conditions:

  • Patient is not appropriate for a conventional allogeneic stem cell transplantation (age, complicated illness, < 70 years).
  • The disease is potentially curable with an allogeneic stem cell transplantation.
  • No CNS involvement from the disease. 
  • Left ventricle ejection fraction ≥ 40 %.
  • No serious disturbances in lung function (need for oxygen supply, DLCO < 30 %, FEV1 < 30%).
  • No serious disturbances of biochemical liver parameters, that is:
    • bilirubin more than 2 fold above the normal limit 
    • ASAT and ALAT more than 4 fold above the normal limit
  • Karnofsky score greater than or equal to 50%.
  • Blood pressure 150/90 with standard hypertension medication.
  • Renal function almost normal.

The principle for this type of treatment is to utilize the anti-tumor effect from the donor's T-lymphocytes (and NK cells). The mechanism for this effect is unclear and the frequency of response varies from disease to disease. The method consists of a conditioning regimen which is strongly immunosuppressive, but not myeloablative, to achieve a mixed and subsequent full T-cell chimerism. When the result is successful, the autologous tumor cells are destroyed by the allogeneic lymphocytes. 

Indications

  • Patients with an HLA-identical relative donor or 10/10 identical unrelated donor, who are not suitable for conventional allogeneic stem cell transplantation. Preferably within diagnosis-specific clinical protocols. Must be reported to EBMT.

Possible diagnoses: 

  • Mantel cell- and follicular lymphoma with recurrence after HMAS and chemosensitive disease
  • Chronic lymphocytic leukemia after unsuccessful chemotherapy, including fludarabine
  • Chronic myeloid leukemia with unsuccessful cytogenetic response to optimal drug therapy
  • Multiple myeloma after HMAS
  • AML in research protocol - patients with potential family donor

Goal

  • Cure the disease.

Background

Donor

A blood related donor is preferred and currently, a blood related donor with up to one HLA (human leukocyte antigen) mismatch may be used. 

If the patient does not have a blood related donor, it may in some cases be considered whether an unrelated donor with HLA, A, B, C, DR and DQ identity can be used. Despite serological techniques, unrelated donors may still have small differences in their HLA molecules which are of significance for GVHD and rejection reactions, therefore genomic typing is performed.  

Stem cell harvesting

Hematopoietic stem cells are harvested by extracting bone marrow from the donor's hip bone or by mobilizing hematopoietic stem cells from bone marrow to blood. Mobilization can be done with the help of hematopoietic growth factors, most often granulocyte colony stimulating factor (G-CSF) given subcutaneously. The growth factors most likely influence adhesion molecules on the stem cells and bone marrow stroma allowing the stem cells to release into the blood stream. Because of this technique, it has become common to refer to stem cell transplantations instead of bone marrow transplantations as it reflects both methods of harvesting stem cells.

Harvesting from bone marrow

For extraction of stem cells from bone marrow, the tissue-compatible donor is given general anesthesia and lies in the prone position. Bone marrow is aspirated by repeated punctures of the hip bone. The procedure lasts 1 hour on average. The aspirated bone marrow with the stem cells is transferred to blood pouches with heparin. From the donor, 2 x 108 per/kg body weight of nuclei-containing cells are transferred to the recipient.

Harvesting from blood

Harvesting stem cells from blood does not require anesthesia, but the procedure can be time-consuming. The stem cells are harvested from a peripheral vein with the help of leukapheresis machines.

Each leukapheresis requires many hours. It is necessary to have 1-3 leukaphereses and a minimum of 2 x 106 CD34 of positive cells per/kg body weight of the recipient.

There are indications that the use of stem cells from blood can lead to quicker hematopoietic reconstitution than stem cells from bone marrow.


Preparation

The application for a transplant is submitted to the Allogeneic Stem Cell Group of Norway with copy to the lymphoma group at the Oslo University Hospital, the Radium Hospital. As a minimum, the application must include a short disease history including  information about the time of remission, treatment given, precise diagnosis and evidence for it. It is also necessary to provide the results from cytogenetic and molecular genetic data. Information about complications from treatment, general health status, organ function, and if the patient has a blood-related donor should also be included.

Examination before transplantation

In addition to the different blood tests and bone marrow tests, there are a series of examinations which all patients must complete before a stem cell transplantation:

  • Lung function tests
  • Dental examination
  • Sperm examination/examination from gynecological clinic
  • Eye examination 
  • X-ray examination of heart and lungs, possibly other organs

Infertility

It is unknown how this treatment affects fertility in men and women. It is important to offer sperm banking to men, if it is possible.

Banking ovarian tissue is technically possible, but at the present time, this is experimental.

Information

The patient and their family should receive thorough information about treatment, complications, and circumstances which should be in order before the treatment.

Preparation for stem cell transplantation

Before starting conditioning, there are measures which must be taken to prevent and treat complications of chemotherapy and GVHD

Intravenous access

All patients must have a central vein catheter. If it is necessary to perform plasma replacement due to ABO incompatibility, the patient will need a two-way dialysis catheter. 

Hemotherapy

All blood products given during the time frame from one month before the transplant, to at least 12 hours after, must be radiated to hinder proliferation of any included T-lymphocytes in the immunosuppressed patient, causing GVHD. Radiation is necessary despite always using leukocyte-filtered blood products.  

All patients should receive blood products which are filtered for leukocytes. This ensures that the products are functional with CMV-negative patients and are used on both anti CMV-positive and negative patients.

Immunosuppression

Cyclosporin is started before the transplant and administered orally, or intravenously if the patient is not able to take tablets. Other immunosuppressive drugs (tacrolimus, mycophenolate mofetil, alemtuzumab, sirolimus) are also used.

Antibiotic prophylaxis

  • Pneumocystic jerovecii prophylaxis—trimetoprim sulfa as for myeloablative transplants
  • Fungal prophylaxis—fluconazole daily from the 3rd day before the transplant and for 75 days after
  • Herpes zoster virus (HZV)/varicella zoster virus (VZV) prophylaxis:
    • for serum negative donor and recipient, prophylaxis is unecessary 
    • for HSV and/or positive recipient or negative recipient/positive donor, valacyclovir is given for a minimum of 4 weeks 
    • for VZV positive donor and/or recipient, valacyclovir is given until day 360.
  • Cytomegalovirus (CMV)—follow the same routines as for a myeloablative transplantation

Implementation

Conditioning regimens

Flu/2 Gy TBI (Seattle Protocol)

  • Flubarabine is administered intravenously on days 4, 3, and 2 before the transplantation.
  • On the day of the transplantation, the patient undergoes total body irradiation with 2 Gy.

Fludarabine/cyclophosphamide (Modified NCI protocol)

  • Fludarabine and cyclophosphamide are administered intravenously on days 6, 5, 4, 3 before the transplantation.
  • Day 2 before the transplantation, the immunosuppressant sirolimus is given.

For successful conditioning regimens, support treatment in the form of a 5 HT3  receptor antagonist or metoclopramid is given to prevent nausea, which is usually moderate.

In addition, the patient should have at least 3 liters of fluid per day during conditioning, either orally or intravenously. Allopurinol is given to prevent uric acid nephropathy.

Radiation treatment given in conjunction with the Seattle protocol often causes nausea which is treated with ondansetron. Other conditioning regimens are also used.

Infusion of stem cells

The stem cells are supplied to the recipient by intravenous infusion, as in a normal blood transfusion or as a boost.  

The stem cells migrate to the recipient's bone marrow where they establish with the help of adhesion molecules and proliferate in a complicated, and far from fully clarified, interaction between cytokines, growth factors, and other cellular interactions.   

For ABO incompatibility between the donor and recipient, serious hemolysis may occur. Therefore, antibodies must be removed either by plasmapheresis, if the recipient has antibodies in a high titer against the donor erythrocytes, or by removing the plasma from the donor marrow if the donor has a high titer against the recipient.


 


Follow-up

Hospitalization is not always necessary. If hospitalization is necessary, the patient should have a private room and is monitored with the same tests as a patient in myeloablative conditioning. Patients who are under outpatient care should be seen every other day for the first weeks, and should be instructed to contact an emergency doctor immediately if their temperature is > 38°C and/or increasing malaise. The unit should have a low threshold for admitting the patient for a clinical evaluation.

Bone marrow suppression should be expected between days 7 and 21, but seldom to a serious degree. The cyclosporin concentration should be measured daily for the first 2–3 weeks, otherwise, tests for cyclosporin concentration and other tests are taken Monday, Wednesday, and Friday for ordinary transplantations. 

On days 28, 56 and 84 after the transplantation, tests are taken for chimerism studies.

The remission status is assessed every 3 months with blood and bone marrow tests as for a myeloablative stem cell transplant. This may be supplemented with cytogenetics, flow cytometry, and/or PCR tests. For extramedullary disease optimal image analysis is taken which is usually CT.

Complications

Some complications should be expected after the transplant. Complications are sometimes serious and life-threatening, and in the worst case, fatal.

Infections

Due to large doses of busulfan and cyclophosphamide, the patient will have a serious immune defect and granulocytopenia during the first few days after taking the medication.

For serious granulocytopenia, normal signs of infection will often not be present because the patient will not create pus. In the aplasia phase, bacterial infections are almost always primary, which can quickly become very serious. The only sign of infection is often fever, which must be taken very seriously for these types of patients.

A reliable microbiological diagnosis is obtained relatively rarely. In cases where there is bacterial growth, the treatment is adjusted according to the resistance pattern.

If the patient becomes afebrile from the antibiotic treatment, the treatment should continue for at least 3 days or until the granulocyte count is over 0.2 x 109/l.

Both GVHD prophylaxis and GVHD requiring treatment with steroids or another immunosupressant, increase immune deficiency. Even after the patient has received sufficient granulocytes, the immune system is still impaired, with a significant risk for infections.

Important opportunistic microbes which can cause life-threatening infections are:

  • cytomegalovirus (CMV)
  • fungus (candida og aspergillus)
  • pneumocystic jerovecii
  • bacteria, such as pneumococcus, influenza hemofilus

The patient must therefore be monitored by checking for the CMV antigen or PCR. Only leukocyte-filtered products can be used. These products rarely or never transfer CMV infections.

One should be aware of new lung infiltrates, especially in patients with chronic GVHD. It is important to try to obtain diagnostic material. The threshold for doing bronchoscopy with broncial rinsing is low.

Fungal infection

In patients who are in the aplastic phase receiving antibiotics for 5-7 days without being afebrile, or who have a new rise in temperature despite adequate antibiotic treatment, a systemic or invasive fungal infection must be considered, especially if another infection cannot be detected in a blood culture. Empirical treatment must be considered with antimycotics in a sufficient dose, often intravenously must be considered.

Pneumonia/respiratory infection

Focal infiltrates are most often due to either a bacterial or fungal infection. It is important to be aware that patients without granulocytes rarely develop obvious infiltrates. An X-ray of the lungs is often negative in the aplasia phase. HR-CT is a much more sensitive examination. Infiltrates may become visuble when the granulocytes return after the cytopenia phase, with a radiologically aggravation. This may be in spite of adequate treatment and clinical improvement. It is therefore very important with close clinical observation.

Interstitial pneumonia

Interstitial pneumonia is a lung infection with numerous small areas of infection. The condition is a feared complication in stem cell transplanted patients. Interstitial pneumonia can occur both before and many months after the engraftment. The mortality is high. The condition may be due is due to infections sensitive to treatment.

  • CMV infection
  • Pneumocystic jerovecii (observed rarely during trimetoprim-sulfa prophylaxis)

It is therefore very important to initiate the correct treatment. Patients with interstitial pneumonia can quickly become dependent on a respirator and must be followed closely with a pulse oxymeter and by monitoration of arterial blood gases.

CMV infection

A CMV infection is not uncommon after an allogeneic stem cell transplantation and is often due to reactivation of a virus in sero-positive patients. Acute GVHD and treatment of this increases the risk.

The manifestations can vary from asymptomatic virus production via thrombocytopenia and leukopenia, to unexplained fever, hepatitis or gastrointestinal symptoms with life-threatening interstitial pneumonia.

CMV pneumonitis typically occurs 40-60 days after the transplantation but can also occur later on. The fear of CMV pneumonitis is one of the reasons leukocyte-filtered products are used for allogeneic stem cell transplantations.

Detection of a CMV infection (positive pp65 or CMV-PCR) will usually lead to starting treatment with ganciclovir. The dose is reduced for reduced renal function. For granulocytes under 0.7 x 109/l, foscarnet is given instead. pp65 or CMV-PCR should be monitored at least once a week during the treatment.

Thrombotic microangiopathy

These conditions occur rarely, but are potentially serious complications which affect about 5% of patients. The pathogenesis is only partly known, but endothelial damage is a central factor.

  • Microangiopathic hemolytic anemia (MAHA)
  • Hemolytic-uremic syndrome (HUS)
  • Thrombotic thrombocytopenic purpura (TTP)

Symptoms are:

  • serious hemolysis
  • renal failure
  • thrombocytopenia
  • neurological manifestations due to reduced circulation in capillaries

The benefit of therapeutic measures is controversial. Cyclosporin A is replaced with mycopholate or tacrolimus. A problem is that tracrolimus can also cause thrombocytopenia. It is also possible, but seldom effective, to perform plasma exchange.

Acute GVHD

Graft-versus-host disease is a condition where T-lymphocytes from the donor attack cells and tissue of the patient.

Bone marrow failure

Bone marrow failure refers to lasting anemia, leukopenia, and thrombocytopenia. Causes for bone marrow failure may be:

  • rejection of bone marrow
  • delayed engraftment
  • infection (CMV among others)
  • bone marrow-toxic medications
  • GVHD

Megakaryocyte function is restored often lastly restored after the stem cell transplantation. Thrombocytopenia may also be due to enhanced consumption of platelets from infection, GVHD, and immunizing.

In addition to reduced marrow function, anemia may be due to bleeding and/or hemolysis. For ABO incompatibility, a significant delay of adequate erythropoiesis is often observed (up to one year).

Long-term anorexia

Anorexia may be due to infection, GVHD, renal dysfunction, or medications. If the patient in addition has problems with swallowing, esophagitis, gastritis, or an ulcer may be the cause. The nutritional status should be maintained intravenously, as needed, which itself may cause anorexia in some patients.


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