Cord blood transplantations for inherited disease

Professor Gideon Bach - Medical Director of "Bedomaich Chayi" Cord Blood Bank

Introduction: The transplantation of hematopoietic stem cells (stem cells that are the precursors of the different blood cells) has been used for over 2 decades as an important approach for the treatment of a broad spectrum of human disorders. These diseases include various hematological malignancies (blood cancers such as leukemia, lymphoma etc), as well as a variety of inherited disorders. Although the hematological malignancy patients are still the majority of the patients treated by this approach, nevertheless, in recent years the use of this source of stem cells had been constantly increased for increasing numbers of various genetic diseases. In many instances encouraging outcomes have been reported describing significant improvements and even cure of certain disorders following stem cell transplantation, thus, the number of patients treated by this approach is constantly increasing and these patients constitute today a large proportion of the total treated patients.

The lysosomal storage disorders (LSD) are an important group of inherited disorders in which encouraging reports for successful outcomes following transplantations of these stem cells had been published. This group includes approximately 50 different inherited syndromes, mostly relatively rare, though some are found in relatively high frequency in certain defined ethnic groups such as Tay Sachs and Gaucher diseases among Ashkenazi Jews, or Metachromatic Leukodystrophy among Yemenite Jews, etc. We also know of various LSD in high frequency among various Arab large families in Israel and neighboring countries, including, Krabbe, Hurler, Niemann-Pick syndromes and many more. All the LSD are inherited by a recessive trait and all but two are autosomal, meaning that a patient (mostly young children) will be born if both his parents are carriers of this disease. It should be noted that carriers are unaffected with no clinical signs of the diseases but if both parents are carriers they have a 25% probability for an affected child in every pregnancy. Two of the LSD, namely Fabry and Hunter syndromes are inherited by an X linked mode of inheritance, thus a carrier mother will have a 50% chance for an affected son.

The basic cause for the LSD and the characteristic pheotypes

Most of the LSDs are caused due to the deficiency of one of the enzymes located in lysosomes (mostly total lack of enzyme activity) . Lysosomes are intracellular organelles in which the breakdown [degradation] of complex molecules such as long sugars, fats, proteins, etc. occur. The deficiency of a particular enzyme in these organelles results in the storage of the substrate(s) that the lacking enzyme participates in their breakdown. As a result, a constant accumulation of massive amounts of this substrate(s) occurs in the lysosomes leading to cell damage and serious clinical phenotype. The nature of the storage is determined by the identity of the lacking enzyme and thus different organs and tissues are affected in each LSD, resulting in typical clinical manifestations for each disease. Most of these disorders manifest as serious devastating fatal diseases already in very early ages and thus most LSD are pediatric diseases (children disorders). Life expectancy is usually short, characteristic for each disease. In most of the LSD the central neurologic system (brain) is seriously affected so that most patients suffer from mental and motor retardation, neurodegenaration etc., but other somatic tissues and organs are usually also seriously affected, such as liver, spleen, skeleton etc. It should be noted that less severe or milder patients are also known to exist and we know today that part of this variability stems from the nature of the mutation in the relevant gene that will determine the clinical manifestations, but even in milder patients, (juvenile and adult forms) the disease is still serious and life threatening. 

Treatment possibilities for LSD patients

As mentioned above most LSDs are caused by the deficiency of one of the enzymes normally located in lysosomes. The pioneer work by Dr Elizabeth Neufeld and her group in the U.S. opened the door for successful treatment of the LSD. Dr Neufeld and her group of investigators were the first to identify a number of the defective enzymes in various LSD.  Furthermore, they could show that the exogenous addition of the missing enzyme to patients' cells, affected with a LSD, this added enzyme was taken up by the diseased cells by specific recognition cellular markers. The internalized enzymes were found to be biologically active enabling the breakdown of the accumulated substances and normalizing the status of the cells, thus achieving in fact the correction of the defect (correction of the abnormal storage). In other words, the supply of the deficient enzyme from an external source, for instance by the injection to the blood stream of a LSD patient of sufficient amounts of the enzyme can lead to the correction of the metabolic defect which is manifested by the storage of usually massive amounts of the accumulated substance(s). In order to achieve a successful outcome sufficient amounts of the missing enzymes is required for the uptake to cells of all the damaged tissues and organs, and furthermore, a requirement to design the structure of the supplied enzyme with the appropriate recognition markers to be able to penetrate to the affected cells (as outlined above). Indeed, in recent years treatment protocols for LSD patients had been developed by this concept so that patients receive periodic injections of the relevant enzyme. Hundreds of patients had been treated by this approach, for instance Gaucher patients, and lately also Fabry, Pompe, Hurler patients to mention only a few (all of which are different LSD with a characteristic enzyme deficiency). It should be mentioned that these treatments are expensive due to the requirements for large amounts of the proper enzyme, mostly from commercial sources, and the need for repeated injections, in some cases weekly injections. A very serious limitation arising in these treatments is the inability of the injected enzyme to cross the border from the blood stream to the brain due to the blood brain barrier (BBB). Thus, the effect of these treatments for the defect in the brain is very limited if at all, and thus it usually is successful only for somatic tissues and organs. We have to bear in mind that many LSD are involved with serious damages to the brain. This limitation is also relevant in a different treatment approach of gene therapy in which the defective gene is injected rather then the enzyme, that is in fact the product of the relevant gene. Although direct injections of either the missing enzyme (protein) or the gene coding for the enzyme to the brain had been experimented by various investigators, this is a complicated and risky procedure.

Bone marrow and cord blood transplantation for LSD treatment

Due to the limitations of the treatment procedures outlined above alternative approaches had been investigated and being used. The most important alternative for the treatment of many different LSD is by the transplantation of hematopoietic stem cells, i.e bone marrow transplantations, and lately cord blood is being used in increasing numbers as another source for these cells. The rational of this concept is based on the fact that the white blood cells from the healthy donor are producing normally the different lysosomal enzymes including the one that is deficient in a particular patient. Some quantities of these enzymes are excreted out from the white blood cells and eventually recaptured by cells of the patient finally reaching the lysosomes containing the stored substances. Thus, a LSD patient receiving transplantation from a healthy donor the transplanted stem cells will eventually mature to adult white blood cells that will be transferred to the blood stream and later to the various tissues and organs of the patient. These donor cells will supply the missing enzyme leading to the removal of the stored materials and a subsequent improvement and even cure of the disease. Although the major therapeutic effect stems from the supply of the missing enzyme from the donor cells to the patient cells, these transplantations have an additional value stemming from the presence of another source of stem cells in the transplantation preparation – the mesenchymal stem cells. These stem cells are present in bone marrow and cord blood in some quantities and can differentiate to a broad spectrum of various tissue cells and thus are being used as an important source for the regeneration of damaged organs ("Regenerative medicine"). It is assumed that these cells in the cord blood or bone marrow apparently contribute to the restoration of the damaged tissue of the patients, contributing to the therapeutic efficiency. The transplanted cells have another important advantage due to the fact that a subtype of these cells - the phagocytes, reach also the brain from the blood stream and thus are of therapeutic potential also for this important organ.

The first hematopoietic stem cells transplantations were reported from the U.S.A and Europe already during the 1970th using bone marrow cells for various LSD patients. These transplantations were performed for Hurler, MPSVII, Gaucher diseases patients and other LSD in lower numbers. In most cases there were encouraging reports for the outcome of these procedure, indicating a significant slow down of the deterioration of the patients that remained either in a steady state or even improved regarding various aspects of the clinical manifestations, for instance significant reduction in liver and spleen volumes when those were affected, and even some clear improvement in neurological parameters including mental retardation.

We shall bear in mind that bone marrow transplantations is involved with a significant risk for the patient mainly due to severe or chronic immunologic responses such as GVHD (graft versus host disease – immunologic response of the donor cells against the recipient organs) that may cause high rate of mortality and morbidity. On the other hand we shall bear in mind that almost all LSD are severely devastation and fatal disorders with a short life span in most cases. Thus, these line of treatments may be the best hope for the patients, indeed from the time these procedures were first used until present time is considered as one of the most important tool to help these patients, and was already used for hundreds of patients hitherto.

Cord blood transplantations

In the last 20 years cord blood has been used in increasing numbers as an alternative source for hematopoietic stem cells transplantations, instead of bone marrow. The cord blood (CB) is the blood of the baby, in fact of the placenta, collected from the umbilical cord immediately after the birth. The blood is collected by a syringe from the vein in the umbilical cord. This blood was found to be rich in various stem cells, primarily of course the hematopoietic stem cells. Indeed this source was first used in the middle of the 90th of the last century by Prof. Eliane Gluckman from Paris, France who was the pioneer in this field reporting in 1998 a successful transplantation of CB in a child affected with Fanconi anemia, which is a disease involved with bone marrow failure. Later, Prof. Pablo Rubinstein from the New York blood bank applied this technology broadly and his contributions in this field are important breakthroughs in the use of this source of cells. He developed means for efficient processing and maintenance of CB and used these cells for patients in a variety of blood malignancies.

The major obstacle of this source for patient's transplantations is the relatively low number of stem cells as compared to bone marrow thus the earlier use of CB was primarily performed for young children where there is a need for lesser quantities of stem cells than in adults for successful outcome. Nevertheless, during the past 10 years, due to the contributions of Dr. Rubinstein and others the procedures for collecting and processing of CB units has become more and more efficient both in the collection of the cord blood as well as the preparations of the CB units in the laboratory prior to the transplantation. Indeed, today CB units are prepared so that they mostly contain sufficient stem cells also for adult patients. Thus, in recent years the proportion of adult patients treated by CB transplantation of the total treated patients is increasing constantly, comprising over 50% and in some counties over 60% of the treated patients. At present, over 20000 cord blood transplantations had been already performed worldwide for both children and adults.

Today, there are a large number of public banks in which CB units are being stored in appropriate conditions ready to be used for needed patients upon request from all over the world. These banks are known as public cord blood banks, meaning that the CB units are available for any needed patient and the relevant information regarding the CB relevant properties is kept in several central information centers. The most important information required for a successful transplantation outcome is the data regarding tissue typing characteristics (HLA) to achieve maximum compatibility in tissue specificity between the donor and the patient and prevent as much as possible any rejection reactions after transplanting the cells. The CB units are kept prior to use in the banks at -196⁰C (in liquid nitrogen) that enables to keep the specimen for at least 20 years.

Advantages of cord blood

The most important feature favoring the use of cord blood transplantations is the fact that these specimen are readily available; this is due to the fact that already today there are over 400000 CB units stored in various public banks in the world and it is relatively easy to locate suitable samples with a high rate of HLA compatibility for a patient in a few days unlike bone marrow that requires a few months to find and obtain suitable donor. Another important advantage of CB is a relatively lower risk for severe immunologic reactions such as GVHD (see above) that is a serious problem in bone marrow transplantations due to the fact that the CB cells are more naïve in their immunogenic properties. Finally, the requirement for HLA compatibility is less strict than in bone marrow so that successful outcomes are achieved also with only ca 70% compatibility, though higher compatibility up to 100% increases even more the success for these transplantations.

Published results of CB transplantations in recent years indeed clearly indicated unambiguous successes in these transplantations, with similar and even improved outcome when compared to bone marrow while the search for a suitable donor is much faster and easier for CB.

CB transplantations for Krabbe disease patients

From the beginning of its use, CB transplantations were performed not only for hematologic malignancies such as leukemia patients, but also for a variety of inherited disorders of which various LSD are one of the most prominent. CB were used instead of bone marrow that was used in the past. Dr Joanne Kurtzberg from Duke University, North Carolina, USA reported the broadest experience in this field. The first report from this medical center in 2005, described the outcome of CB transplantations for 25 patients affected with Krabbe disease, which is a severe neurodegenerative LSD with a very short life expectancy if not treated, of 2-5 years. The disease involves a serious damage to the brain that undergoes a process of dismyelinization due to the storage of a specific lipid. The Duke team transplanted 2 groups of patients with CB, the first, very young patients before the manifestations of clear signs of the disease and the 2nd group included older patients, mostly 6-12 months old after the disease had been already clearly manifested. In the first group the outcome indicated a clear success so that there were barely any abnormal clinical abnormalities and the children remained in good health for 7-8 years post transplantation, while their untreated siblings mostly died in significantly younger ages from complications of the disease. The first group (pre-onset of the disease) showed barely or only slight clinical deterioration and mostly showed normal growth and development during the follow-up period. Cognitive evaluations indicated clear improvement of neurological and cognitive parameters. The results in the 2nd group (older patients) was less favorable and these children continued to deteriorate and some died from these complications during the follow-up period. This outcome indicates the importance for treatment as early as possible in the course of the disease so that the disease is prevented rather than reversed back to normal health. This also indicates the clear advantage of CB that is much faster available for transplantation while in bone marrow the search may take a few months.

Newborn screening for Krabbe disease

The encouraging results that had been achieved for the successful treatment of Krabbe disease in very early ages by CB transplantation prompted authorities from various states in the U.S such as New York State and others to include the diagnosis of this disease in the newborn screening tests performed in many states and countries throughout the world. The aim of this project is to ascertain Krabbe patients as early as possible and perform the CB transplantation, from an appropriate donor, in the attempts to prevent the onset of the disease. The diagnosis of the patients still requires appropriate means to reliably identify only those patients that the CB transplantation is indeed required.

CB transplantation for other LSD

Similar results and outcome as described above were achieved for various other LSD including Hurler (MPS I), Hunter (MPS II), Metachromatic Leukodystrophy (MLD) and others. These results were reported by the group of Dr Kurzberg as well as from other medical centers. Despite the known risk of this procedure, over 70% of the patients survived the transplantation procedure without serious rejection complications and complications. Over 90% of the transplanted patients showed clear improvement of the disease manifestations, in many the clinical deterioration had been considerably slowed down. As already stated above, the most important factor for the success of this procedure is the treatment as early as possible in the child's life. Thus, similarly to the U.S other countries such as Australia included LSD in the newborn screening operation.

Preconception genetic diagnosis (PGD) for families with inherited disorders

In recent years a new technique had been developed for prenatal diagnosis that enables the diagnosis of embryos immediately after in-vitro fertilization (IVF) of 2-3 days old embryos, prior to the return of the embryo to the uterus and conception. This diagnosis enables to identify embryos that are not affected with an inherited disease that is present in high risk families, and alternatively also identify embryos bearing fully matched HLA compatibility to their affected live sibling. By this diagnosis the family can chose to have a healthy offspring with a perfect match to the affected child and thus can become a perfect donor of the CB after birth and save the affected child. As mentioned above, the newborn child must be also tested by this technique to ascertain that he is not affected by the disease. Indeed this procedure was used already in several cases by families with an affected child in diseases that are treatable by hemapoeitic stem cells transplantation. Of course this procedure may ensure a better and probably successful outcome of the transplantation in those patients, but questions of ethic background had been raised occasionally using this approach since the new offspring might be considered to be born specifically to save his affected sibling. Does it indeed reduce the value of this individual life is still an open question. Is this a specifically designed child?

Conclusions

Every year are born In Israel, similar to other countries, numerous children suffering from different inherited diseases, including various LSD. We should be better prepared so that indeed the diagnosis of these patients will be achieved as early as possible so that they will indeed benefit from CB transplantation in due time. This at present might be the only meaningful treatment for these patients.

 

Professor Gideon Bach - Medical Director of "Bedomaich Chayi" Cord Blood Bank, Jerusalem. Chairman of the Israel Society of Human Genetics, Member  of  the Counseling Committee of the Ministry of Health for Clinical Genetics Policies Member of the national Helsinki committee (Ministry of Health) on genetics experimentations. He headed the Human Genetics Dept. of Hadassa Ein Kerem Medical Center in Jerusalem for many years.