MXPA99004444A - Conditioning for allogeneic stem cell transplantation - Google Patents

Abstract

The present invention features methods for conditioning patients prior to allegeneic stem cell transplantation. A first method involves treating a patient with a lymphoablative regimen that retiains a functional population of the patient's hematopoietic stem cells. A second method involves treating a patient with a myeloablative regimen that, conversely, retains a functional population of the patient's T lymphocyte population. In both methods, the patient is administered a donor-derived stem cell preparation after the conditioning regime to induce host anti-donor unresponsiveness. The patient may also be administered allogeneic cell therapy. The invention also features a method of making a patient-specific allogeneic stem cell preparation.

Description

CONDITIONING FOR TRANSPLANTATION OF ALOGENIC TOTIPOTENTIAL CELLS BACKGROUND OF THE INVENTION The lethal conditioning or high dose regimens that use chemotherapy and / or radiotherapy followed by rescue with allogeneic totipotent cell transplantation (allo-SCT) or autologous totipotent cell transplantation (ASCT) have been the treatments of choice for patients with a variety of hematological malignancies and chemosensitive solid tumors, resistant to conventional doses of chemotherapy. A common source of totipotent cells for such procedures has been the bone marrow. Recently, totipotential peripheral blood cells (PBSC) have also been used. As such, the terms "allogeneic bone marrow transplantation" (allo-BMT) and "autologous bone marrow transplantation" (ABMT) are widely used in the literature to refer to particular types of allo-SCT and ASCT, respectively, either that the rescue be with bone marrow or PBSC. REF .: 30156 Current procedures typically employ allo-SCT or ASCT after myeloablative / lymphoablative (M / L) conditioning. As the name implies, conditioning with M / L involves the elimination, through death, blockage and / or cell sub-regulation, of substantially all the hematopoietic totipotential cells and the lymphocytes of the patient. Patients treated with allo-SCT or ASCT may develop major complications due to conditioning with M / L. In addition, patients who receive allo-SCT are susceptible to developing graft versus host disease (GVHD), as well as graft rejection. In addition, relapse is still a frequent problem in these patients. Several attempts to improve disease-free survival by increasing the intensity of conditioning with M / L have failed due to unacceptable toxicity. In addition, the increase in conditioning intensity with M / L does not seem to improve the result by decreasing the relapse rate. A wide variety of protocols of varying intensities have been used among more than 30,000 transplants around the world, reported to the International Registry of Bone Marrow Transplants. Despite these numerous attempts to vary the intensity of the conditioning regimens, there are still no significant differences documented in complete results with patients. The use of conditioning with M / L followed by rescue with allo-SCT is often accompanied by graft versus tumor responses (GVT), for example, graft versus leukemia (GVL). Over the years, immune interactions between immunocompetent T lymphocytes derived from an allogeneic donor that act against host-type tumor cells have shown that they are of major therapeutic importance. For example, significantly better anti-tumor effects have been induced by allo-SCT compared to ASCT or transplants from an identical twin. Relapse after allo-SCT or ASCT in patients has been reversed a few times by adoptive therapy of allogeneic cells (allo-CT) using infusions of donor lymphocytes (DLI). The complete eradication of tumor cells by DLI, despite the resistance of tumor cells to maximally tolerated doses of M / L conditioning, suggests that alloreactive T lymphocytes may represent a crucial weapon against tumor cells. Transplantation of allogeneic totipotential cells that leads to allogeneic totipotential cell grafting in the host may function merely to induce a state of host versus graft tolerance, allowing T lymphocytes derived from an allogeneic donor, concomitant or subsequently administered, to survive and recognize and eradicate the tumor cells derived from the host. In fact, the main therapeutic component of allo-SCT can be ascribed to the effects of GVT or GVL mediated by T lymphocytes, rather than to the physical elimination of tumor cells by conditioning with M / L before transplantation. The effects of GVL or GVT mediated by T lymphocytes generally occur in the context of allo-SCT, transplantation of totipotential cells of allogeneic peripheral blood (allo-PBSCT; for example, a form of allo-SCT) or allo-CT. However, as discussed above, these procedures can lead to complications related to conditioning with M / L, GVHD and / or rejection of the injury.

BRIEF DESCRIPTION OF THE INVENTION This invention provides new methods for the treatment of a human patient with a disease of pathogenic cells. It has been found that conditioning regimens can be designed to allow the patient to maintain relatively high levels of either totipotent cells or functional lymphocytes. Thus, in one method, the conditioning regimen is designed to eliminate T lymphocytes from the patient, but to allow the retention of a functional population of hematopoietic totipotential cells from the patient. In a second method, the conditioning regimen is designed to decrease or abate the totipotent cells of the patient, but to allow retention of a functional population of the patient's lymphocytes. In both methods, after the patient has been treated with the conditioning regimen, a preparation of allogeneic totipotent cells derived from the donor is administered to the patient. Patients treated according to the methods of the invention develop inability to respond specifically to the donor, and they also develop relatively minor complications than with standard M / L regimens. The method also provides a platform for performing allo-CT to induce GVL, GVT or the effects of graft versus autoimmunity (FVA), and allows the development of patient-specific allogeneic totipotential cell preparations. In a first aspect, the invention characterizes a method for the treatment of a human patient having a disease of pathogenic cells. The method includes the treatment of the patient with a conditioning regimen that preserves a functional population of the patient's hematopoietic totipotential cells. The method also involves the administration of a preparation that includes allogeneic totipotent cells from a donor, to the patient, under conditions effective to induce the inability of host anti-donor response. The regimen may be an M / L conditioning regimen, or a conditioning regimen - / L. Preferably, the allogeneic totipotent cells are totipotential cells of peripheral blood, totipotential cells of cord blood or totipotent cells of bone marrow. The method may also include a step for the provision of allogeneic cell therapy to the patient. Allogeneic cell therapy is provided after the induction of the inability of host anti-donor response, and in the absence of significant GVHD. Allogeneic cell therapy may include the administration of donor T lymphocytes in incremental increments, while controlling GVHD without immunosuppression. T lymphocytes can be of limited life time. The T lymphocytes can be CD8 + cells or CD4 + cells. In one embodiment, allogeneic cell therapy may include the administration of donor T lymphocytes, activated before the administration, to the patient. In yet another embodiment, allogeneic cell therapy may include in vitro administration of the T cell activator to the patient. The conditioning regimen may include the administration of one or more agents such as purine analogues, alkylating agents or anti-leukocyte globulins. In one embodiment, the purine analogue is fludarabine and the anti-leukocyte globulin is anti-T lymphocyte globulin. In yet another embodiment, the regimen includes the administration of fludarabine, anti-T lymphocyte globulin and an alkylating agent. The alkylating agent can be, for example, busulfan or cyclophosphamide. Pathogenic cell diseases treatable with methods that include malignant diseases such as chronic myelogenous leukemia, acute myelogenous leukemia, acute lymphoblastic leukemia, non-Hodgkin's lymphoma, myelodysplastic syndrome or multiple myeloma. Malignant disease can also be a solid tumor as in metastatic breast cancer. In another modality more, disease of pathogenic cells can be a non-malignant disease such as ß-thalassemia major, Blackfan Diamond anemia, Gaucher anemia, Fanconi anemia or AIDS. Non-malignant disease can also be an autoimmune disease. In a second aspect, the invention characterizes the treatment of a patient having a disease of pathogenic cells, with a conditioning regime that preserves a functional population of the patient's T lymphocytes. This second method also includes the administration of a preparation that includes allogeneic totipotent cells from a donor to the patient, under conditions effective to induce the inability of host response, anti-donor. The regime can be a conditioning regime M / - or M / l. The method may also include a step of providing a therapy regimen with allogeneic cells to the patient, after induction of host anti-donor response capacity, and in the absence of significant GVHD. The conditioning regimen may include the administration of an alkylating agent, such as busulfan or cyclophosphamide. Cyclophosphamide can be administered together with hydroxyurea. Alternatively, the conditioning regimen may include the administration of irradiation to the whole body, preferably accompanied by the administration of cyclophosphamide. In yet another aspect, the invention features a method for preparing a patient-specific allogeneic totipotential cell preparation. The patient, having been administered with a conditioning regimen, is endowed with a select veto capacity. The method includes obtaining a preparation of totipotent cells from an allogeneic donor, and adjusting the veto capacity of the preparation, to balance the selected veto capacity of the patient. The method may also include adjusting the patient's veto capacity to balance the veto capacity of the preparation. In another aspect, the invention features the use of allogeneic totipotent cells of the donor in the manufacture of a medicament for the treatment of a disease of pathogenic cells. The medicament is administered to a host patient conditioned with a regimen that preserves a functional population of the hematopoietic totipotent cells of the host, and the host is administered the drug under conditions effective to induce the inability of the host's antidonary response. The medication may also include allogeneic lymphocytes.

In a further aspect, the invention features the use of allogeneic totipotent cells of the donor, in the manufacture of a medicament for the treatment of a disease of pathogenic cells. The medicament is administered to a host patient conditioned with a regimen that preserves a functional population of the host T lymphocytes, and the host is administered the drug under conditions effective to induce the inability of host anti-donor response. The medication may also include allogeneic lymphocytes. In still another aspect, the invention features the use of allogeneic lymphocytes of the donor in the manufacture of a medicament for the treatment of a disease of pathogenic cells. The medicament is administered to a host patient conditioned with a regimen that preserves a functional population of the hematopoietic totipotent cells of the host, and the host is administered the medicament after induction of the inability of host anti-donor response. In a further aspect, the invention features the use of allogeneic lymphocytes of a donor in the manufacture of a medicament for the treatment of a disease of pathogenic cells. The medicament is administered to a host patient conditioned with a regimen that preserves a functional population of the host T lymphocytes, and the host is administered the medicament after induction of the inability of the host's antidonary response. The term "myeloablative" as used herein includes any therapy that eliminates, through cell death or cell inactivation, substantially all the hematopoietic totipotential cells of host origin. "Myeloablative" is referred to herein as "M". The term "sub-myeloablative" as used herein includes any therapy that removes a significant fraction of, but not substantially all, the totipotent hematopoietic cells of host origin. "Sub-myeloablative" is referred to herein as "m". The term "lymphoablative" as used herein includes any therapy that substantially eliminates all functional T lymphocytes of host origin. This is achieved through death, blockage and / or cell under-regulation. The elimination can be short-term or long-term. "Linfoablative" is referred to herein as "L". The term "sub-lymphoablative" as used herein includes any therapy that removes a significant fraction of, but not substantially all, the functional T lymphocytes of host origin. "Sub-lymphoablative" is referred to herein as "1". The terms M, m, L, and 1 can be combined in any way to categorize the particular conditioning regimes. For example, the term "M / L" as described above, refers to a conditioning regimen that is myeloablative and lymphoablative. The term "m / L" refers to a conditioning regimen that is sub-myeloablative and lymphoablative. The terms "M / l" and "m / l" refer equally to the corresponding conditioning regimes. The term "- / L" as used herein refers to a conditioning regimen that is lymphoablative but does not significantly affect the patient's hematopoietic totipotential cells. The term "M / -" as used herein refers to a conditioning regimen that is myeloablative but does not reduce the patient's T lymphocytes. The grafting effect versus pathogenic cells as used herein, refers to the graft response against any pathogenic cell including a cancer cell, in general an abnormal totipotential cell, autoreactive T lymphocyte as in an autoimmune disease, and an infected cell derived of the host, such as a T cell infected by HIV-1 or reticuloendothelial cell. The term "cancer" as used herein includes all pathological conditions involving malignant cells; This may include "solid" tumors that arise in solid tissues or organs, as well as hematopoietic tumors such as leukemias and lymphomas. The main advantages have to be anticipated as a result of the clinical application of the methods described herein, with a relatively low incidence of short-term and long-term complications in the first place. In this way, patients should experience fewer episodes of infection, and are at reduced risk of bleeding due to thrombocytopenia, veno-occlusive liver disease, and interstitial pneumonitis. Patients treated with the methods described also generally experience low rates of acute and chronic GVHD, severe. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of ordinary skill in the art to which this invention pertains. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, suitable methods and materials are described below. All publications, patent applications, patents and other references mentioned herein are incorporated by reference in their entirety. In the case of conflict, it will control the present specification, including the definitions. In addition, the materials, methods and examples are illustrative only and are not intended to be limiting. Other features and advantages of the invention will be apparent from the following detailed description, and from the claims.

BRIEF DESCRIPTION OF THE FIGURES Figure 1 is a series of graphs showing the duration and degree of pancytopenia and the application of allografts of blood totipotential cells mobilized with G-CSF, identical to HLA, after a m / L conditioning regimen.

Figure 2 is a graph showing the probability of disease-free survival at various times after allo-SCT.

DETAILED DESCRIPTION OF THE INVENTION Two general procedures have been developed for conditioning regimens that circumvent the need to substantially eliminate the patient's hematopoietic totipotential cells and T lymphocytes. In the first procedure (Method 1), the patient is provided with a conditioning regimen (- / L ) or (m / L) that allows the retention of a functional population of hematopoietic totipotent cells but that eliminates substantially all the T lymphocytes of the patient. In the second procedure (Method 2), the patient is provided with a myeloablative (M / -) or (M / l) conditioning regimen that substantially eliminates all of the patient's hematopoietic totipotential cells, but allows the retention of a functional population of the T lymphocyte population of the patient. After the patient is treated with one of the conditioning regimens described above, the patient is given a preparation derived from the donor, which includes allogeneic totipotent cells. The preparation is administered under effective conditions to induce the ability of the host's anti-donor response. In an optional third step, the patient may be administered an allo-CT regimen after the induction of antidonor response capacity by the host, and in the absence of significant GVHD.

Human patients with a variety of pathogenic cellular diseases can be treated by the methods of the invention. Diseases of pathogenic cells include malignant diseases such as chronic myelogenous leukemia (CML), acute myelogenous leukemia (AML), acute lymphoblastic leukemia, non-Hodgkin's lymphoma (NHL), myelodysplastic syndrome (MDS), multiple myeloma (MM), primary lymphoma of the central nervous system (CNS lymphoma), as well as solid tumors such as metastatic breast cancer. Diseases of pathogenic cells also include non-malignant diseases, for example genetic disorders. Examples of non-malignant diseases include β-thalassemia major, Fanconi anemia, Gaucher's disease, Blackfan Diamond syndrome, acquired immunodeficiency syndrome (AIDS), as well as autoimmune diseases.

METHOD 1 A human patient with a disease of pathogenic cells is treated with a conditioning regimen that preserves a functional population of the patient's hematopoietic totipotential cells. Retention of a population of functional totipotential cells allows the patient to avoid the commonly encountered clinical effects of cytopenia, for example sepsis and fungal infections due to neutropenia, susceptibility to life-threatening parasites such as Pne umoci s ti s cari ni and hemorrhage due to thrombocytopenia. Preferably, the patient retains at least 20% of the population of functional hematopoietic totipotential cells, more preferably at least about 50%, and more preferably at least about 90%, of the population of functional totipotential cells. With respect to the patient's hematopoietic totipotential cells, the conditioning regimen can be submyeloablative. Alternatively, the regimen may lack any conditioning that decreases the patient's functional hematopoietic totipotential cells, for example the regimen may allow the retention of substantially all of the patient's functional hematopoietic totipotential cells.

With respect to the population of functional T lymphocytes of the patient, the conditioning regimen of Method 1 includes an intense lymphoablative regimen that transiently eliminates the population of functional T lymphocytes from the patient. The lymphoablative regimen can also temporarily eliminate the population of natural killer (NK) cells. This severe lymphoablation reduces or suppresses the population of functional T lymphocytes to a level that allows the grafting of allogeneic donor cells in a great majority of patients subject to conditioning regimen. Preferably, the infoablative regimen 1 is sufficiently severe to allow the grafting of allogeneic totipotent cells in at least about 90% of the patients. More preferably, the lymphoablative regimen is sufficiently severe to allow the grafting of allogeneic totipotent cells in almost 100% of patients. Preferably, the lymphoablative regime transiently decreases the population of functional T lymphocytes of the host, by at least about 90%. More preferably, the lymphoablative regimen transiently decreases the population of functional T lymphocytes of the host by at least 95%, and more preferably, by at least about 99%. In this way, the conditioning regimes applicable to Method 1 can be classified as conditioning regimes m / L or - / L. Examples of useful agents for sub-myeloablative components of the conditioning regimen include alkylating agents such as busulfan, cyclophosphamide, hydroxyurea, for example, carmustine (BCNU), etoposide (VP16), chlorambucil, thiotepa, carboplatin, cisplatin and melphalan. Other agents may also be useful for submyeloablative conditioning such as cytosine arabinoside (ara-C) and anthracyclines such as idarubicin. Ionizing radiation at low dose distributed by an exogenous radiation source, such as cobalt or linear accelerator, can also be used. An internal source can be used by the provision of a radiolabeled compound that searches for the marrow, such as strontium or a radioactive compound directed to the totipotent cells by antibodies specific to the totipotent cells.

Examples of useful agents for the lymphoablative components of the conditioning regimen include purine analogs such as methotrexate, cladribine (2-CDA) and fluodarabine (FLU). Alkylating agents such as cyclophosphamide can also be used for lymphoablative conditioning. Melphalan, thiotepa and busulfan are alkylating agents that are weakly immunosuppressive and can also be used. Anti-leukocyte, polyclonal and monoclonal globulins, such as anti-lymphocyte globulin, can also be used.

(ALG), anti-1-galactone globulin, anti-T lymphocyte globulin (ATG) and antibodies against well-defined T lymphocyte subgroups. The lymphoid irradiation can also be used. Doses of agents that have myeloablative and lymphoablative activities can be adjusted to provide the appropriate myeloablative, submyeloablative, lymphoablative, or sublipoblad activities. This is illustrated, for example, in the patient protocols summarized below in the examples. When a conditioning regimen with m / L is used, a combination of alkylating agents, purine analogs and / or anti-leukocyte globulins can be used as conditioning agents. A preferred m / L conditioning regimen includes the administration of sub-myeloablative amounts of bisulfan, together with sufficient amounts of FLU plus ATG to achieve severe lymphoablation. If the day on which the totipotent cells are infused is established as day 0, then a typical conditioning regimen under Method 1 can be summarized as follows: FLU: 30 mg / m2 / day for 6 consecutive days (days -10 to -5) Busulfan: 4 mg / kg / day for two consecutive days (days -6 and -5) ATG: 10 mg / kg / day for 4 consecutive days (days -4 to -1) In an alternative modality, cyclophosphamide at 10-60 mg / kg / day for two consecutive days at days -6 and -5 can be replaced by busulfan. It should be understood that these agents and dosing schemes are illustrative only, and that other agents and dosage schemes having similar effects on the patient may be employed. When a conditioning regimen - / L is employed, the agents administered may include alkylating agents and anti-leukocyte globulins. A preferred / / L conditioning regimen employs appropriate doses of cyclophosphamide and ATG to achieve severe lymphoablation without significant myeloablation. For example, ATG can be provided at 10 mg / kg / day on days -8 to -5, and cyclophosphamide is given at 50 mg / kg for one, two, three or four consecutive days starting on day -4. Two to four consecutive doses of cyclophosphamide are preferred in this case, since this leads to a greater probability of totipotential cell grafting than if cyclophosphamide is limited to a single dose. Again, it should be understood that these agents and dosage schemes are illustrative only, and that other agents and dosage schemes having similar effects on the patient may be employed. After the patient is treated with one of the conditioning regimens described above, the patient is given a preparation derived from a donor, which includes allogeneic totipotent cells. The preparation of totipotent cells may also include T lymphocytes derived from the donor. Although it may be desirable in many circumstances to use a donor with compatible HLAs, in other cases it is permissible to have a non-compatibility in one or more of the histocompatibility loci. In this way, the donor can be a compatible sibling in the HLA-A, B, C, DR, DRB1 loci. In other modalities, however, the donor can be one, two or three locus not compatible with the patient either in class I, class II or both. T lymphocytes in the preparation of totipotential cells derived from the donor, and to a lesser degree the same totipotential cells (see below), can act as "veto" cells to produce a veto effect. Veto cells as used herein include T lymphocytes, especially CD8 + T lymphocytes, or other cells that result in underregulation instead of stimulation of other T lymphocytes against cells containing the same alloantigens as the veto cells themselves. . Other proliferating hematopoietic cells that include decreased totipotential cells in T lymphocytes, which are poorly immunogenic, can also induce veto effects against T lymphocytes. In this context, most or all cells that possess cell surface alloantigens can serve as veto cells. The veto capacity of a cell can be transient. For example, the veto capacity of a cell may be higher during cell division than during periods of rest. Even with severe lymphoablation, there may be small numbers of residual lymphocytes in the patient. In addition, residual lymphocytes can expand in number in response to alloantigens, leading to a host versus graft effect. Through the veto phenomenon, the residual T lymphocytes of the host can be down-regulated by the veto cells derived from the donor, including totipotential cells and / or T lymphocytes. Other cells derived from the donor, in replication, can also veto the derived T lymphocytes. of the host, if they are provided in relatively high concentrations. Conversely, immunocompetent T lymphocytes present in the donor preparation can be down-regulated by veto cells of host origin. In this way, a balanced balance can be achieved that minimizes or even eliminates graft versus host responses, thereby minimizing or also preventing possible GVHD. A balanced balance can also minimize or eliminate the effects of host versus graft, thereby reducing the chance of graft rejection. A balanced balance is achieved by balancing the alloreactive T cells of the donor with the veto cells of host origin, which represent the appropriate antigens (antigens of class I and / or class II or other minor histocompatibility antigens in a non-immunogenic form). Similarly, the host's alloreactive cells can be balanced by the veto cells of donor origin, which present the corresponding antigens (class I and / or class II antigens or other minor histocompatibility antigens in a non-specific form). immunogenic). The adjustment of the veto capacity of the host-derived cells presenting host antigens and donor-derived cells, which present donor antigens, results in the state of bilateral non-response that is induced spontaneously and consistently in stable mixed chimeras. To achieve a balanced balance of veto capacity, the veto capacity of the preparation of totipotential cells derived from the donor can be adjusted to correspond to the patient's conditioning regimen. As the intensity of the conditioning regimen of the patient decreases, especially the lymphoablative component, a greater proportion of residual alloreactive cells remains capable of causing rejection. In this way, larger numbers of donor cells with relatively strong veto capacity are required. In the preparation of totipotent cells of the donor, the T cells and the totipotential cells can downregulate the host's alloreactive cells and prevent rejection to the graft, hence the need for greater numbers of donor cells as the conditioning regimen decreases in intensity. Guest. Host veto cells can similarly prevent GVHD despite the inoculation of a greater number of alloreactive T cells derived from a donor. This explains the requirement for an increased number of totipotent donor cells when the graft is decreased in T cells, since the lack of T cells that act as veto cells must be compensated by an increased number of totipotential cells. In this case, the totipotent cells provide the required proportions of tolerance agents in non-immunogenic form, necessary for the down-regulation of residual host alloreactive T cells. The totipotent hematopoietic cells of the donor can be obtained by direct extraction from the bone marrow or from the peripheral circulation after mobilization from the bone marrow. The latter can be achieved by treatment of the donor with the granulocyte colony stimulation factor (G-CSF) or other appropriate factors that induce the mobilization of totipotent cells from the bone marrow to the peripheral circulation. The mobilized totipotential cells can be collected from the peripheral blood by any appropriate cellular pheresis technique, for example through the use of a blood collection device, commercially available as exemplified by the CS3000 Plus blood cell collection device marketed by Fenwal Division of Baxter Healthcare Corporation. Methods for performing apheresis with the CS 3000 Plus machine are described in Williams et al., Bone Marrow Transplantation 5: 129-133 (1990) and Hillyer et al., Transfusion 33: 316-321 (1993). Alternative sources of totipotential cells include totipotent neonatal cells (e.g., totipotential cells from umbilical cord blood) and fetal totipotential cells (e.g., fetal liver or yolk sac cells). Totipotential cells that have been expanded by a mixture of hematopoietic cytokines can also be used. Other useful totipotential cell preparations include totipotential cells that have been transduced with genes encoding the MHC class I or class II molecules of the donor type, as well as preparations of totipotential cells containing totipotent cells and / or T cells. transduced with herpes simplex thymidine kinase or other "suicide" genes to make mature T cells sensitive to ganciclovir or other appropriate drugs in the case of severe GVHD. Similarly, T cells derived from the donor or host, transduced with herpes simplex thymidine kinase or other "suicide" genes to make mature T cells susceptible to ganciclovir or other appropriate drugs, can be used to vet T cells of the host or donor, thereby controlling rejection or GVHD, respectively. The hematopoietic totipotential cells in the preparation are administered under conditions effective to induce the inability of the donor to respond specifically, by the host. Such inability to respond results when the totipotent cells of the donor are able to engraft in the patient as a result of an appropriate conditioning regimen. As described above, the preparation of totipotential cells derived from the donor may include T lymphocytes from the donor in addition to the hematopoietic totipotential cells. The preparation of the donor hematopoietic totipotential cells can be even enriched for T lymphocytes, particularly if a relatively high veto capacity is needed to balance a robust residual population of T lymphocytes in the host patient. It is well known that T lymphocytes in the preparation of totipotent cells can generate a graft effect versus pathogenic cells within the context of the donor-specific inability to respond, which results from grafting the hematopoietic donor totipotential cells. In some modalities, it can be administered to the anti-GVHD patient. Since an anti-GVHD agent suppresses T lymphocytes, this can also suppress the graft effect versus pathogenic cells and the veto capacity of T lymphocytes. Therefore, it is advantageous to stop administering the anti-GVHD agent as early as possible, in order to facilitate the effect of grafting versus pathogenic cells, as well as the veto capacity of the totipotent donor cell preparation. The administration of the anti-GVHD agent can be gradually diminished when the patient shows signs of grafting of totipotent cells of the donor, with the proviso that the indications of GVHD are absent or negligible. Although the anti-GVHD agent can be administered for more than three months if necessary, preferably the anti-GVHD agent is administered to the patient for no more than about ninety days, even more preferably for no more than about thirty days. Cyclosporin A is a particularly useful anti-GVHD agent. Other effective agents for preventing GVHD are within the scope of the invention. Examples of other anti-GVHD agents include methotrexate, imuran, celcept (mycophenolate mofetil), ALG, anti-lymphocyte and anti-adhesion / host antibodies, and corticosteroids. Alternatively, an anti-GVHD agent may not be necessary if the veto capacity of the preparation derived from the donor is suitably balanced by the veto capacity of the host patient. In such a case, since the donor's veto cells are in balance with the host, they "veto" one another leading to substantial reduction or even total avoidance of GVHD. As discussed above, veto capabilities can be balanced by adjusting the veto capacity of the donor derived preparation to correspond to the patient's conditioning regimen, or vice versa. In situations of mixed chimeras, where hematopoietic cells derived from the donor and derived from the host coexist, the veto capacities can be inherently balanced. The patient can be administered with other compounds such as antibiotics, which can be administered prophylactically. For example, septrin, acyclovir and ganciclovir can be administered prophylactically for the prevention of Pneumocystis carinii, herpes simplex virus and cytomegalovirus, respectively, prior to the administration of the totipotential cell preparation derived from the donor. The grafting of totipotent donor cells into the host can be detected by any number of standard methods. The presence of donor markers, such as specific markers of the sex chromosome, in the host can be determined, for example, using standard cytogenetic analysis, the polymerase chain reaction (PCR) with appropriate primers, variable number of repetitions in tandem-PCR (VNTR-PCR), microsatellite markers or other genetic fingerprint printing techniques, or fluorescence hybridization in si tu (FISH). The host-donor chimerism can also be determined by determining the percentage of donor-type cells in the blood of the host using, for example, standard complement-dependent microcytotoxicity tests. After grafting the cells derived from the donor, the patient may be provided with an allo-CT regimen to supplement any graft effects versus pathogenic cells of the donor lymphocytes, infused with the preparation of totipotent cells. Alo-CT is described, for example, in PCT Publication Nos. WO 95/24910 and WO 96/37208. Alo-CT involves the administration of allogeneic peripheral blood T lymphocytes derived from the donor, to the host, either alone or in combination with a T-cell activator such as interleukin-2 (IL-2). Preferably, allogeneic peripheral blood T lymphocytes are administered in incremental increments to prevent or control GVHD. The allogeneic peripheral blood lymphocytes can be "preactivated" using a T-cell activator such as IL-2., then administered either alone or in combination with the same or a different T-cell activator. Alternatively, allogeneic peripheral blood lymphocytes manipulated with a suicide gene, such as the herpes simplex virus thymidine kinase gene, can be administered at an early stage after the transplant. T lymphocytes manipulated in such a manner over a wide range of cell numbers can be administered since such cells can be selectively removed later, if necessary. Preferably, one or more infusions of about 105 to about 109 cells / kg of allogeneic peripheral blood lymphocytes are administered. These lymphocytes can be administered as defined subgroups of T cells (eg, CD4 + or CD8 + subgroup) if desired. Various T-cell activators are suitable including, without limitation, IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-10, IL-12, IL- 13, IFNa, IFN ?, TNFa, anti-CD3, anti-CD28, phytohemagglutinin, concanavalin A and phorbol esters. In general, allo-CT is performed after a desired level of incapacity for anti-donor response has been induced in the host, with the allo-CT functioning to generate or amplify the grafting effect versus pathogenic cells. In addition, allo-CT can increase the proportion of donor cells in the host by causing additional displacement of the host's hematopoietic totipotential cells. Alo-CT typically involves the administration of T lymphocytes from the donor to the host, although allo-CT may also include the administration of natural killer (NK) cells derived from the donor. During allo-CT, GVHD can often be controlled without the use of anti-GVHD agents. Although the presence of totipotent cells of the donor, grafted on the patient, establishes a level of inability to respond to the donor, however it is prudent to administer allo-CT in gradual increments while periodically verifying for signs of GVHD. Alo-CT is typically discontinued when GVHD is indicated and / or the effects of grafting versus pathogenic cells (eg, GVT, GVL). The T lymphocytes used in allo-CT can be limited by the expansion of life. The cells limited by the expansion of life can be controlled to function only transiently in the host. T cells of limited lifespan can be generated, for example, by transformation with "suicide" gene vectors. See, for example, Bonine et al., Sci en ce 276: 1719-1724 (1997). Such vectors typically make the lymphocytes selectively susceptible to particular chemical agents, such as ganciclovir, in the case of donor T cells transduced with the herpes simplex virus carrying the thymidine kinase gene.

METHOD 2 In an alternative way to treat a human patient with a disease of pathogenic cells, a myeloablative (M / -) or (M / l) conditioning regimen may be used. These regimens substantially reduce the population of functional hematopoietic totipotential cells of the patient, while retaining substantially all (M / -) or a substantial fraction (M / l) of the functional population of T lymphocytes. As a consequence, a patient treated with the conditioning regimen M / - or M / l has a particularly high veto capacity mediated by residual T lymphocytes. Suitable myeloablative conditioning regimens may include the administration of myeloablative doses of one or more of the myeloablative agents described above. A typical conditioning regimen under Method 2 can be summarized as follows: Busulfan: 4 mg / kg / day for four consecutive days (days -4 to -1) It should be understood that the use of busulfan in this dosage scheme is illustrative only, and that other agents and dosage systems that have similar effects on the patient may also be employed. Other conditioning regimens may involve melphalan (120-240 mg / m2 in 1 or 2 divided doses), thiotepa (10 mg / kg in 1 or 2 divided doses), hydroxyurea at a high dose (3-6 g / day until myeloablation is achieved) or other variations of dies such as dimethyl mileran. The dibromomanitol (DBM). it can also be replaced by busulfan. With M / - or M / l conditioning regimes, preferably the patient retains at least about 20% of the population of functional T lymphocytes, more preferably at least about 50% of the population of functional T lymphocytes and still more preferably at less about 90% of the functional population of T lymphocytes. After the conditioning regimen, the patient is administered with a preparation derived from the donor, which includes hematopoietic totipotent cells. The preparation derived from the donor can be a preparation with a high capacity of veto, for example, the preparation can contain large numbers of T lymphocytes, since the patient treated with the conditioning regimen M / - or M / l has a high capacity of veto. If desired, the preparation derived from the donor can be enriched for the T lymphocytes or the non-immunogenic soluble antigens to down-regulate the alloreactive cells of host origin. The T lymphocytes of the donor-derived preparation not only provide veto capacity, which controls rejection to the graft, but can also generate a substantial grafting effect versus pathogenic cells. As discussed above under Method 1, allo-CT can also be administered to the patient after induction of the host's anti-donor response inability, and in the absence of significant GVHD. In view of the defined conditioning regimens that are employed in Methods 1 and 2, it is possible to specifically design a hematopoietic totipotential donor cell preparation for a particular patient. As discussed above, such preparations can be considered as having a "veto" capability defined by the ability of the cells in the preparation to veto the anti-donor activity of the T lymphocytes in the host patient. T lymphocytes in the preparation of totipotent cells have a significantly higher veto activity compared to totipotential cells. However, it seems that totipotential cells do have a relatively weak but not insignificant veto activity. Immunocompetent T cells of the host and donor that possess the most effective veto capacity can also cause disadvantageous rejection and GVHD; in this way, their numbers must be carefully balanced. The purified totipotential cells areHowever, they are relatively safe because their veto capacity is not associated with the co-favorable allogeneicity of the donor or the host, respectively. Ideally, the veto capacity of the preparation of totipotent cells is balanced or "in balance" with that of the host patient. This minimizes the risk of GVHD while maximizing the probability of grafting totipotent cells. To achieve this, the numbers of T lymphocytes or totipotential cells, or both, can be adjusted to correspond to the intensity of the lymphoablative portions of the conditioning regimes. If the patient is severely lymphoreduced (lymphoablation) as Method 1, the preparation of totipotent cells does not need to possess substantial numbers of T lymphocytes, and may even be completely depleted of T cells. In some cases the host patient may possess small numbers of lymphocytes residuals reactive with the donor, despite the intense lymphoablative measures. In these cases it is possible to provide sufficient veto capacity even in a totipotential cell preparation decreased in T cells, by infusing particularly large numbers of totipotent cells. Alternatively, relatively small numbers of T lymphocytes from the donor can be included in the preparation of totipotential cells in order to ensure the veto of the residual host T cells. In another-S additional cases, T lymphocytes can be added to a totipotential cell preparation (enrichment with T lymphocytes), particularly if the totipotential cell preparation originally had a low number of T lymphocytes, and with the proviso that the capacity The veto of the preparation of totipotential cells remains functionally balanced with the veto capacity of the patient. Consequently, the totipotential cell preparations of the donor can also be designed for the conditioning regimes of Method 2, where the patient retains a functional population of T lymphocytes. In these cases, the preparation of totipotential cells derived from the donor is typically adjusted to have a relatively high veto capacity, balancing with the high veto capacity of the host patient. This can be achieved by the inclusion of high numbers of purified totipotential cells, or totipotent cells enriched for T cells. The methods described herein can surely be offered to patients in all age groups with low anticipated incidence of immediate complications and long term that commonly result from conventional transplants when myeloablation is combined with lymphoablation (M / L). The malignancies and genetic diseases can be treated at an early stage of the disease, without undue delay, due to the interest of the attending physician for the toxicity related to the procedure, which results from the conventional BMT that involves an M / L conditioning regimen. Treatment at an early stage may also avoid the need for repeated courses of chemotherapy with cumulative toxicity over the years. Treatment in the early stage of a disease can increase the opportunity for complete eradication of all tumor cells, before drug resistance occurs, by an optimal conditioning regimen and early therapy with allogeneic cells. In genetic diseases, early treatment can prevent irreversible damage to multiple organs, especially when the central nervous system (CNS) is at risk. Similarly, patients with autoimmune diseases may also be excellent candidates for allogeneic transplantation of totipotential cells, because early treatment can prevent irreversible complications such as ankylosis in rheumatoid arthritis., remains in the central nervous system and paralysis in multiple sclerosis, and other complications. In patients with acquired immunodeficiency syndrome, early transplantation of totipotent cells with allo-CT can reconstitute the patient's immune system with immunocompetent lymphocytes derived from the donor, preventing life-threatening infections, and preventing or treating secondary malignancies. In younger individuals, the methods described herein, in contrast to traditional M / L conditioning regimens, should not lead to growth retardation and may reduce the risk of infertility due to chemo-sensitive testicular and gonadal tissues, while they avoid additional endocrine abnormalities and late complications such as cataracts. In older individuals, who may not normally be qualified for a standard totipotential cell transplantation procedure due to unacceptably high complication rates, the methods of the present invention allow a relatively safe procedure. The ability to apply innovative allo-CT procedures after transplants of relatively safe totipotent cells, using the conditioning regimens of Methods 1 or 2, is likely to convince the attending physicians to consider the use of the procedures of curative transplantation in the early stages of the disease, when the opportunity for healing may be relatively high and before the disease is irreversibly spread to multiple organs. In addition, effective treatment at an early stage of the disease can prevent the development of tumor cell clones resistant to chemotherapy, platelet resistance induced by sensitization from repeated administration of blood products, cumulative toxicity in multiple organs, and development of resistant strains of infectious agents through repeated courses of antimicrobial therapy with susceptibility to fatal bacterial and fungal infections. The therapy mediated by the methods of the present invention can result in a significant opportunity in the complete success rate for disease-free survival, as well as improvements in quality of life and cost effectiveness for future transplant candidates. of totipotent cells. In the future, the early application of similar procedures for the induction of tolerance to transplantation may prevent the sensitization of candidates for organ allografts and the more effective use of limited allograft delivery of organs that can always be harvested with totipotent cells from the body. donor from living and deceased donors. The present invention will be further described with reference to the following examples; however, it should be understood that the present invention is not limited to such examples.

Examples Example 1 Method 1: Conditioning regimen (m / L) Protocols based on Fludarabine / ATG PATIENTS AND METHODS A combination of FLU at 30 mg / m2 x 6 days, busulfan at 4 mg / kg x 2 days, and ATG was administered to 26 patients suffering from m / L conditioning before the administration of the totipotential cells of peripheral blood, mobilized with G-CSF derived from the donor, allogeneic (10 μg / kg x 5 days) obtained from donors of 1 or 2 loci not equal and HLA completely equal. The characteristics of the patient are described in Table 1. All patients could have been considered eligible candidates for a standard alo-SCT program.

Table 1 Patients Suffering Conditioning System m / L * The donor (sibling) was incompatible at loci A and C, with positive MLR in the direction of the host versus graft. The patients included 6 with chronic myelogenous leukemia in the chronic phase (CML / CP); 1 with CML in accelerated phase (CML / AP); 1 with Youth CML (JCML); 7 with acute myelogenous leukemia (AML) in complete remission (CR) one who had secondary leukemia (AML, m5) 3 years after treatment for carcinoma of the ovary, and 1 with AML in 2 ° CR; 1 with acute lymphoblastic leukemia (ALL) in 1st CR and 1 in 2 ° CR; 2 with non-Hodgkin lymphoma (NHL) resistant to frontal line chemotherapy; 1 with myelodysplastic syndrome (MDS) with excess blasts; and 1 with multiple myeloma (MM). The series also included 4 patients with non-malignant disorders including a child with severe beta thalassemia major; 1 child with Fanconi anemia; 1 child with Gaucher's disease and 1 adult with Blackfan Diamond syndrome (Table 1). The ages of the patients were in intervals between 2 to 61 (median 31) years. Conditioning prior to infusion of the allogeneic totipotential cells included a m / L conditioning regimen with 6 infusions of FLU (Schering's AG) per day for 6 consecutive days (-10 to -5), with each infusion containing 30 mg / m2 (in adults, the dose was adjusted according to the ideal body weight); oral busulfan 4 mg / kg / day for 2 consecutive days (days -6 and -5); and ATG (Fresenius) 10 mg / kg / day for 4 consecutive days (days -4 to -1). One patient (UPN 111 with Fanconi anemia) received cytoxan 10 mg / kg / day for 2 consecutive days (days -6 and -5) instead of busulfan. The totipotential blood cells mobilized by G-CSF were collected from the donor once after a 5-day treatment of G-CSF at 10 mg / kg / day. Siblings with equal or comparable HLA-A, B, C, DR, DRB1 were used as donors, with one exception; UPN 1109 was grafted with inequalities at locus A and C as indicated by the positive mixed lymphocyte reaction in the host versus donor direction. The total number of nucleated cells infused on day 0 was in the range of 3.38-16.39 (mean 8.60) x 108 / kg. Prophylaxis against GVHD included standard cyclosporin A (CSA) 1.5 mg / kg twice daily intravenously, starting on day -1, changing to an oral dose of 3 mg / kg twice daily, as soon as patients were given high, with early elevation starting as soon as the graft was confirmed without GVHD (around 4 to 6 weeks), and the patient's stabilized condition. Prophylaxis against Pneumoci s ti s carinli included trimetoprim / sulfamethoxazole (10 mg / kg / day of trimethoprin) administered preimplant (days -10 to -1) and reduced to 7 mg / kg / day of trimethoprim twice a week as soon as the absolute neutrophil counts exceeded 0.75 x 109 / L.

Statistical Evaluation: The Kaplan-Meier method was used to calculate the probability of disease-free survival as a function of time, as well as to determine the recovery time of hematopoietic reconstitution.

RESULTS The m / L conditioning regimen was better tolerated in all patients, compared to anticipated side effects after a standard M / L conditioning regimen. As can be seen in Table 2, grade 3 or grade 4 toxicity (criteria of the World Health Organization) were not observed in any of the patients. Grade 2 mucositis was documented only in 2 cases. All the patients maintained oral ingestion throughout the procedure with 8 (31%) patients who never required any parenteral caloric supplement. Episodes of septic fever were observed in 4 cases, while 22 patients did not experience evidence of severe, positive systemic infection. Severe veno-occlusive disease (VOD) of the liver was observed only in 2 cases, while 11 patients developed mild to moderate manifestations of VOD and 13 patients showed no evidence of liver abnormality at all. No pulmonary toxicity was observed in any of the patients.

Table 2 Common Complications Related to Transplantation The additional important clinical parameters after m / L conditioning are shown in Table 3 and Figure 1. In 9 patients (31%), the absolute, neutrophil count (ANC) did not decrease below 0.1 x 109 / Liter and for the whole group it took an average of 10 days for ANC to fall below 0.1 x 109 / L (Figure 1A). Two patients never experienced ANC < 0.5 x 109 / L (Table 3). The number of days with ANC < 0.1 x 109 / L in the remaining seventeen patients was in the range between 0 to 20 with a median of 4 days. ANC > 0.5 x 109 / L was achieved within 10 to 32 days (median 15). (Figure IB) The platelet counts did not decrease below 20 x 109 / L in 4 patients (Table 3), thus not requiring support with platelets at all. Among the remaining twenty-two patients a fall in platelet count below 20 x 109 / L was observed after a mean of 7 days, with a probability of 11% of the remnants with a low platelet count (<20 x 109 / L) after day 11 (95% confidence interval from 3 to 27%) (Figure 1C). Spontaneous platelet counts greater than 20 x 109 / L were achieved within 0 to 35 days (median 12). The unsupported platelet counts greater than 20 x 109 / L were observed within 36 days in 85% of patients (95% confidence interval, 69 to 95%), (Figure ID).

Table 3 Clinical Parameters After Conditioning M / L 1 First day when ANC fell below 0.1 x 109 / L. 2 First day when the platelets fell below 2 x 109 / L. 3 First day when ANC rose above 0.5 x 10"/ L. 4 First day when the platelet count rose above 20 x 109 / L; independent of the transfusion. 5 GVHD started when the patient was on therapy with CSA. 6 GVHD was initiated when the patient was out of CSA therapy. 7 The donor was a brother with simple inequality of locus A and C, with a positive culture of a single pathway of mixed host anti-donor lymphocytes. 8 Patient with hematologic relapse 8 months after conditioning with m / L currently under induction of remission with a combination of chemotherapy for tumor de-bulking and allo-CT, in parallel with the development of grade 2 GVHD. 9 Patient with hematologic relapse manifested 4 months after conditioning m / L successfully treated by a combination of chemotherapy for tumor de-bulking and allo-CT, in parallel with the development of grade 2 GVHD, without evidence of disease, with 100% of donor-type cells (female) by cytogenetic analysis and without male cells by PCR. 10 Male patient with tumor cells completely resistant to all chemotherapy at the time of admission. The patient had residual disease after m / L conditioning with marrow infiltration by the lymphoma cells. The patient was successfully treated with allo-CT in parallel with the development of grade 2 GVHD. Currently, there is no evidence of disease, with 100% donor cells (female) by cytogenetic analysis and no male cells by PCR. 11 Patient without haematological evidence of disease, treated with allo-CT for cytogenetic relapse with bcr-abl positive RT-PCR. GVHD greater than or equal to grade 1 was observed in 12 of 26 patients (Table 3). Severe GVHD (grade 3 and 4) was the only major complication, diagnosed in 6 cases (25%) and was the sole cause of mortality in 4 patients, all of whom developed the first signs of illness while out of CSA. Interestingly, acute GVHD was developed only in 4 patients while on regular maintenance therapy with CSA with only one patient who developed grade 3 GVHD while in CSA (UPN 1109, currently alive and in good condition). In 8 cases, the onset of GVHD was observed only after the sudden discontinuation of CSA in an attempt to improve the graft or displace the residual host cells that were documented by molecular or cytogenetic analysis. The 4 patients who died of severe GVHD represent the only observed losses of the complete series, with an observation period that exceeds one year (median 8 months). In one of the patients (UPN 1093) who died of GVHD grade 4, the signs of the disease developed while he was out of CSA, without access to adequate follow-up or to additional treatment with CSA, since she lived at that time a foreign country. His death, therefore, can not be considered a failure of the protocol. The second patient (UPN 1124) died of grade 4 GVHD that developed immediately after discontinuation of CSA and reinfusion on day +22 of a second inoculum of blood totipotential cells enriched with blood T cells intentionally given without CSA in an attempt to improve the graft with granulocytes, delayed. Subsequently, the granulocyte counts increased within less than a week and reached ANC >; of 0.5 x 109 / L at day +35, suggesting that the elevation of totipotent cells may not have been indispensable and may even have contributed to the development of GVHD with fatal results. The third and fourth patients with CML (UPJST 1131 and 1135) also developed grade 4 GVHD after the sudden discontinuation of CSA. The other 8 patients who developed GVHD, of which only 3 manifested GVHD greater than grade 2, responded to standard treatment with prednisone starting at 2 mg / kg with slow decrease as indicated clinically. No conclusive data can be given at this point to assess the total incidence of chronic GVHD in patients conditioned with m / L due to the relatively short observation period in the range of several months to over one year. As can be seen in Table 3, chronic GVHD was diagnosed in 9 patients, in 2 of which (UPN 1073 and 1098) signs of GVHD appeared only after the onset of allo-CT. The graft was documented in all patients by increasing blood counts as shown in Table 3, and either by amelogenin-PCR for host-donors of different sex as described in Pugatsch et al., 17: 273-275 (1996), or VNTR-PCR in donor-patient pairs of the same sex. Naka ura and collaborators, Sci en ce (1987) 235: 1616-1622. In 9 of 26 evaluable patients, a transient stage of mixed chimerism was confirmed by documentation of minimal residual host cells by cytogenetic analysis, PCR or disease-specific RT-PCR (e.g., bcr-abl in CML), and subsequently quickly discontinued CSA. Relapse was observed in 2 patients with acute leukemia (PUN 1073 and 1080) while progressive progressive disease was observed rapidly in a patient with NHL totally resistant to chemotherapy (UPN 1098). Cytogenetic relapse with normal hematological parameters was diagnosed in a patient with CML who did not develop spontaneous GVHD even after the discontinuation of CSA and is now under alo-CT therapy (UPN 1137). Successful displacement of tumor cells by allo-CT was achieved in 2 cases (UPN 1080 and 1098) while one patient is still under treatment and too early for evaluation (Table 3). UPN 1080, a male patient originally treated for ALL in 2a CR, did not characterize GVHD after conditioning m / L and administration of a totipotential cell preparation derived from the donor, developed manifest hematologic relapse at 4 months with the tumor mass that doubled within 1 to 2 days. The large tumor mass was successfully decreased in volume using a combination of cytosine arabinoside at 3 g / m2 / day in 2 doses divided by 4 days and a single dose of mitoxantron at 12 mg / m2, followed by reinfusion of blood totipotential cells of donor (female) enriched with lymphocytes, without CSA. The elimination of all detectable male cells was confirmed by PCR analysis in parallel with GVHD (grade 1-2) induced after infusion with donor lymphocytes (DLI), with stable CR that remained to date without further treatment. UPN 1098 is an elderly male patient with completely resistant NHL, with rapidly progressive malignant lymphoid infiltrate in bone marrow and bone glia. The patient did not show spontaneous GVHD after conditioning m / L and administration of the preparation derived from the donor. The elimination of all manifestations of disease was confirmed after allo-CT with DLI in parallel with the disappearance of the host / male DNA by PCR, and the resultant 100% female karyotype, in parallel with the onset of mild grade 2 GVHD. which evolved to limited, mild chronic GVHD. Another patient (UPN 1073) with AML at 2 ° CR relapsed 9 months after conditioning m / L and administration of the preparation derived from the donor. 'She is currently undergoing combination treatment with chemotherapy and allo-CT, but it is too early to be evaluated. In an observation period extending over 1 year (median 8 months), 22 of 26 patients (85%) treated with alo-m / L conditioning and administration of donor-derived preparation are alive, 21 (81% ) free of disease by all measurable criteria, including PCR, with excellent quality of life, and Karnofsky's rating of 100%. The actuarial disease-free survival at the mean follow-up of 8 months was 80.7% (at 14 months the actuarial disease-free survival was 77.5% with 95% with a confidence interval of 53 to 90% (Figure 2) Chronic mild and limited GVHD was developed in 9 out of 25 patients with a period of observation greater than 100 days, but until now none has developed clinically significant manifestations.

Example 2 Method 1: Conditioning Regimen (- / L) - Protocols Based on ATG / Cyclophosphamide Patients with various haematological malignancies received a conditioning regimen - / L of ATG (Fresenius), 10 mg / kg / day on days -8 to -5, and cyclophosphamide at 50 mg / kg / day on days -4 to - 1 or on days -2 to -1, followed by the infusion of totipotential cells of allogenic peripheral blood (allo-PBSC) from related donors equivalent or equal. All patients received ciclosporin A alone as prophylaxis with GVHD. Patients and patient protocol details are provided below.

RESULTS The parameters and important clinical outcomes of patients treated with the lymphoablative regimen - / L are shown later in Table 4. The average time for ANC > 0.5 x 109 / L was 13 days, while the average time for platelets greater than 20 x 109 / L in 5/6 patients was 18 days (in UPN 1050 the platelet count never decreased below 20 x 109 / L). Two of the patients had clinical syndromes suggestive of veno-occlusive disease, which resolved without sequelae (peak total bilirubin (TB) of UPN1042 that rose to 333 mm / L, TB peak of UPN1056 was raised to 268 mm / L The chimerism was analyzed for all patients with VNTR, and for patients with donors of different sex by amelogenin (AMG-PCR) As shown in Table 4 below, all patients became chimeric, without signs of rejection. mild to moderate cutaneous involvement was present in 3/6 patients with hepatic and bowel involvement in 3/6, UPN1003 died 2 months after transplantation with sepsis and GVHD of the intestine, while UPN1042 died 5 months after transplantation with Staphylococcal pneumonia relapsed into moderate cutaneous NHL and GVHD It is well known that UPN0944 with Ph + ALL is in CR for more than 11 months and UPN1026, with lymphoma of the central nervous system, is in CR for more than 5 months (See Example 3 following) .

Table 4 Additional patients conditioned with a regimen (- / L) are described below.

Patient 1 Diagnosis: Stage IIIc Ovarian Cancer A 49-year-old woman in good health entered a routine physical examination. A pelvic mass was noted during the examination. The laparotomy was performed; Total abdominal hysterectomy, oophorectomy, omentectomy, and dissection of the iliac nodule were performed as a measure to reduce volume, and the patient was classified surgically as minimal residual disease. The pathology showed serous cystadenocarcinoma, poorly differentiated from stage IIIc ovary. The preoperative CA-125 was 1,330, the post operative fell to 780. She then received 7 courses of cisplatin / cytoxan which resulted in a fall of CA-125 to 26. Due to neurological changes, the patient was changed to carboplatin / VP-16 for 4 cycles, but CA-125 rose to 400. CT showed a new iliac mass produced. A second laparotomy showed involvement of the para-aortic nodule with implants on the bladder, involvement of the external iliac nodule and obturator, which was reduced in volume. With CA-125 to 600, she was treated with Taxol 175 mg / m2. After 3 cycles, CA-125 fell to 27, with CT that showed no apparent disease. After 10 cycles of Taxol, CA-125 remained approximately at 22 but CT showed a mass of 2 cm below the diaphragm, without evidence of pelvic or nodal disease. Taxol was continued for a total of 28 courses, with a continued elevation of CA-125 to 152. While waiting for cell therapy, the patient received several courses of cisplatin at 20 mg / d x 3 days q3w. CA-125 continued to rise with values of 328 and 439.6. When reviewing the systems, the patient noticed recurrent dysesthesias in the fingers, especially at night, similar to the symptoms present during the initial course of cisplatin.

I. Conditioning: The patient received ATG at 10 mg / kg / day on days -4 to -1, and cyclophosphamide at 50 mg / kg on day -1.

II. Infusion with Donor Cells: The patient received 7.79 x 108 cells / kg of alo-PBSCT mobilized with G-CSF from her sister with similar HLA.

III. Inj erto: The patient did not experience marked granulocytopenia. On day +4, the patient was discharged with a WBC of 3.4 x 109 / L for follow-up as an external patient. The graft can be followed with RFLP weekly to differentiate between the graft of the donor and the recovery of the host.

IV. Ovarian Carcinoma: According to the plan, the follow-up of CA-125 was carried out as a marker of the activity of the disease. On day +15, the CA-125 was 280.

Patient 2 B.F. is a 36-year-old woman who was in a normal state of health when a mass was felt in her left breast. Although the initial biopsy was undetermined, lumpectomy revealed invasive ductal carcinoma and a mastectomy was performed. The biopsy of the lymph node showed 0/12 LN + and no additional therapy was administered for this T1N0N0 ER positive tumor. Approximately 3 years and then, recurrence of the pectoralis muscle was detected and the patient was treated by resection followed by radiation therapy.

One year later, a new mass was noted on the right side which was malignant with 7 of 13 positive nodules. The mass itself was less than 2 cm. She was then treated with 6 cycles of CAF and 5000 cGyRT. The bone scan at the same time showed a questionable TIO injury. At 11 months, TÍO collapsed, and in 8 months, the bone scan showed multiple bone involvement. Treatment was then started with Taxol and Tamoxifen at 7 months, which she received for 9 cycles, the last dose 5 weeks before admission. During this period he also developed pain in the hip that the bone scan confirmed as due to new metastases. The hip was treated with Spot RT and Tamoxifen was changed to MEGACE about 3 months. Just before admission, the CT of the head was within the upper limits, the CT of the chest showed tumor involvement in L3, bilateral iliac and in the sacrum. Abdominal CT showed that the organs were not involved with the tumor. The aspiration of the bone marrow and the biopsy were negative. His entrance to CA 15-3, was 29 (WNL). The bone scan performed on 20-6-95 (-16 days) showed new lesions in the posterior parietal skull on the right side and in the rib 11 next to the lesions present on 26-2-95 (-5 months) in the left iliac wing, right femoral, spine and ribs.

I. Conditioning: ATG at 10 mg / kg / day intravenously, days -9 to -6; cyclophosphamide at 50 mg / kg / day, days -5 to -2; II. Transplant: The patient received transplantation of totipotential cells of peripheral blood not diminished in T cells (day 0) of her sister, collected in two collections for a total of 5.22 x 108 cells / kg.

III. Prophylaxis of GHVD: On day -1 the patient was started with ciclosporin 2 mg / kg.

IV. Mucositis The patient had very mild oral mucositis during the neutropenic period.

V. Veno-occlusive liver disease (VOD): The patient had a slight elevation in total bilirubin starting on day +6, reaching a maximum of 44 on day +10 and then returning to normal. There never was a significant elevation in liver enzymes.

SAW. Interstitial pneumonitis (IP): The patient was initiated in the first protocol on day +6 with no response. Amphotericin was added on day +10 and the patient had been afebrile since then.

VII. Graft: On day +11, the patient had an ANC of 2 x 109 / Liter that was stable when the patient left G-CSF until she was discharged.

VIII. GVHD: On day 12, the patient developed marked itching, diarrhea, consistent with GVHD. The skin biopsy was consistent with GVHD. The patient was started with 2 mg / kg of steroids and cyclosporine was continued with good response of the skin and intestine. The itching on the skin was limited to the back. Bilirubin did not rise significantly from the time of onset of cutaneous GVHD. The patient died of GVHD without any signs of the disease.

Patient 3 F.E is a previously healthy 57-year-old male patient who was diagnosed with invasive gastric adenocarcinoma, CA-19-9: 20. He has had psoriasis for a number of years. The patient underwent exploratory laparotomy, and inoperable gastric cancer was found and partial resection was performed. He was admitted for the transfer of allogeneic PBSC from a sibling with the same HLA. After insertion of the Hi hm.an catheter, small right lateral pneumothorax developed and had spontaneous resolution. Before stenting, a stent was inserted into the common bile duct using ERCP. After the procedure the elevated bilirubin of 100-120 mmol / L was normalized. In the CT, he had isolated pulmonary metastases and the abdominal one was due to the conglomeration of retroperitoneal gastric hepatic lymph nodes. PE in the admission room, without jaundice, same psoriatic plaque on the skin and palpable epigastric firm mass.

Preconditioning: ATG at 10 mg / kg / day x 4 days iv; cyclophosphamide at a dose of 60 mg / kg / day x 1 day. On day 0, he received PBSC mobilized with G-CSF from his brother with identical HLA at a total of 5.3 x 108 nucleated cells / kg. After the procedure the patient was in good general condition. There was no sign of fever, bleeding or any sign of infection, and he was discharged.

Home Medication: oral Sucralent 1 gram x 4 oral Acyclovir 200 mg x 3 oral Respsin 2 grams / day twice a week The patient did not become chimeric and died of the disease.

Patient 4 A 16-month-old baby (UPN 184) was found to be suffering from ß-thalassemia major when he was ten months old. The treatment consisted of red blood cell transfusions. The patient suffered allogeneic BMT using her non-reactive grandmother to MLC with identical HLA, as a donor. There was greater ABO inequality (A +? 0+). Conditioning prior to transplantation included total lymphoid irradiation (TLl) 2 cGy x 1 / day x 4 (total dose 8 Cgy), followed by busulfan 4 mg / kg / day x 4 and cyclophosphamide 50 mg / kg. / Day x 4. The transplant consisted of decreased bone marrow in T cells that had been treated in vi x with anti-human rat monoclonal antibody Cdw52 (Campath 1M) with the donor serum that serves as the source of the complement. The protocol did not include the prophylaxis of GVHD immunosuppressure after injection. After BMT the patient received 4 intravenous injections of peripheral blood lymphocytes from the donor in an attempt to prevent rejection and / or recurrence of the disease in equivalent doses. to 103 T cells / kg on day +1, 104 T cells / kg on day +6, and 105 T cells / kg on days +12 and +28. The patient was rapidly grafted with absolute nucleated cells (ANC) reaching 0.5 x 109 / L on day +19, 1.0 x 109 on day +27 and the platelet counts of > of 2.5 x 109 / L on the day +29. On day +60 the hemoglobin level (Hb) was 9.4 g / dl without blood support and without evidence of hemolysis to come. Fourteen months after BMT Hb levels fell to 5.7-7.0 g / dl with MCV 70-72 (FL). Repeated evaluation of specific graft markers of the donor included blood group, point mutation of the ß-globin gene, semiquantitative and PCR-VNTR tests. While the PCR-VNTR test failed to demonstrate the presence of donor cells 24, 29, 45, and 51 months after BMT, analysis of the ß-globin point mutation detected 4-7% of derived DNA cells from the donor The patient was homozygous for mutation IVS-nt 110, and the donor was heterozygous for the same mutation. The blood group analysis constantly revealed the presence of donor red blood cells (type A). Despite the state of the donor cells, which are minimally chimeric, mixed, the clinical picture presented intermediate thalassemia with a low transfusion requirement (8 transfusions in a period of more than 4 years.). Hb remained low (median 6.5 g / dl, range 5.4-7.0 g / dl), and serum ferritin levels rose from 437 to 700 μg / 1. The patient showed normal growth and development (50th percentile), although the size of the spleen gradually increased to 4 cm below the left costal margin and the echocardiogram showed abnormal left ventricular function. To clarify the unusual clinical picture of this patient, a mutation test of the ß-globin gene was carried out on cultured erythroid colonies, +/- erythropoietin (Epo). The adition of Epo resulted in the propagation and differentiation of erythroid cells without nuclear ejection.

The donor cells without Epo accounted for 7%, similar to their percentage in the peripheral blood lymphocytes and nucleated BMC, rising to 16% in the presence of Epo. In view of the stable mixed chimerism, a second transplant of the same donor was performed, based on the displacement of the host's immunohepatopoietic system, without myeloablative conditioning. Ambulatory pre-transplant treatment consisted of oral hydroxyurea (1,500 mg / day x 6) followed by a single intravenous dose of 750 mg of cyclophosphamide in parallel with intravenous mesna (250 mg x 3). On the day after the conditioning program, the patient received, on an ambulatory basis, PBSC mobilized with non-manipulated G-CSF, to a total number of 8.74 x 108 / kg nucleated cells collected in three successive aphereses (Baxter CS 3000 Plus) of the original donor. No immunosuppressant treatment was administered after the transplant. The patient tolerated the conditioning regimen extremely well, without severe neutropenia or thrombocytopenia, and therefore continued to be treated in the outpatient clinic. There was a dramatic spontaneous elevation of the hemoglobin level, concomitant with a gradual increase in the percentage of cells derived from the donor. On day +52 the patient suffered signs of acute grade II GVHD, involving the skin and intestine, for which she was treated with methylprednisone and cyclosporin A with good response. More than one year after the second procedure, the patient maintains an Hb score of 12.5 g / dl and shows excellent clinical performance (Karnofsky's score of 100%). The point mutation of the ß-globin gene of donor specific markers, the PCR-VNTR and the blood group were analyzed in the peripheral blood and BM aspirates, which show 100% reconstitution with donor cells.

Patient 5 S.Z., a 31-year-old married woman with 4 children was admitted for the transplantation of allogeneic peripheral blood totipotential cells for metastatic breast cancer *. The patient had suffered breast cancer which was diagnosed during pregnancy. No multiple metastases were revealed on the examination. He was treated with a combination of chemotherapy-CAF without response. He was given hormonal therapy with Tamoxifen unanswered, and salvage chemotherapy with Taxol unanswered. He was admitted for transplantation of totipotent peripheral blood cells, allogeneic, from his brother with completely identical HLA. The physical examination revealed poor general condition and large bilateral masses in the sinuses, huge left nodule on the left axilla, without hepatic enlargement. The WBC 6500 laboratory with normal differentiation, hemoglobin 8.3 g / dl, platelets 350,000, biochemistry: alkaline phosphatase 240 units and gamma GTP-387 units. The CT scan showed multiple bone metastases and the bone marrow biopsy revealed malignant metastasis.

Conditioning for allotransplantation included ATG 10 mg / kg / day x 4 days and cyclophosphamide 50 mg / kg / day x 4 days. Transplantation of peripheral blood totipotential cells, allogeneic, was performed with 3.87 x 108 cells / kg.

Prophylaxis with GVHD: The patient received ciclosporin A from day -1 intravenously at a dose of 3 mg / kg per day.

Course after transplant: The patient developed pancytopenia. He was transfused with red blood cells, and platelets, and received total parenteral nutrition (TPN). It had a temperature with negative blood cultures. ' She received a combination of antibiotic therapy. He developed veno-occlusive disease of the liver with maximum bilirubin of 30.

Injured: On day +14, granulocytes appeared in the peripheral blood and then had white cell and platelet reconstitution. She was discharged on the day +20 after the transplant. In the outpatient clinic, the patient developed acute GVHD with involvement of the skin, liver and intestines. He proceeded on steroids with no response. The patient deteriorated rapidly with grade IV GVHD. He was hospitalized in another hospital, he was treated with a combination of steroids, cyclosporine and antibiotics. He developed an infection and died of acute GVHD and its complications.

Patient 6 G.S. She is a 50 year old woman. He had a malignant melanoma of the skin over the shoulder that was removed and received no additional therapy. Three years later, a mass was found in the left axilla. She had a wide extirpation of the mass including the auxiliary lymph node and the nuchal lymph node. After the surgery, he was treated with local radiation of 8,000 rads. The patient was treated with cyclophosphamide and allogeneic lymphocytes plus IL-2 without evidence of graft and without signs of GVHD. The disease continued to progress. A mass was removed from the urinary bladder and finally the body CT showed lymphadenopathy in the right neck and lymphadenopathy in the retroperitoneum and lymph node in the right pelvis. Due to the progression of the disease, it was decided to admit the patient for transplantation of allogeneic peripheral blood totipotential cells. Physical examinations revealed good general conditions. A small mass in the right neck of 2 cm. There was no hepatosplenomegaly.

Laboratory: Normal complete blood count.

Normal biochemistry The CT scan showed lymphadenopathy in the pelvic peritoneum and neck mass.

Conditioning: ATG at 10 mg / kg / day x 4 days followed by cyclophosphamide at 50 mg / kg / day x 4. The transplantation of totipotential cells of allogeneic peripheral blood was performed by administration of 7 x 108 cells / kg after administration. mobilization with G-CSF.

Course After Transplant: The patient had pancytopenia. She received red blood cells and platelet transfusions. On day +5, she had an episode of fever with positive blood culture with acinetobacter, sensitive to the antibiotic protocol she received since the fever began.

Prophylaxis of GVHD: - The patient received ciclosporin A from day -1 intravenously at 3 mg / kg / day.

Injured: On day +8 the first neutrophils appeared on the smear. As a result, the patient had a complete graft graft with white cell counts and normal platelets. The patient was discharged on day +11 and is currently without signs of GVHD, being treated with cyclosporin A at 7.5 mg / kg / day orally. The patient is under close monitoring in the outpatient clinic. The patient showed transient graft due to allo-CT prior to allo-PBSCT. The allo-CT sensitized the patient to the donor. Because the patient was transiently grafted, the tumor returned. However, the graft was eventually rejected and the tumor returned.

Example 3 Treatment of CNS Lymphoma with Conditioning Regimens - /! > and PBSCT A 25-year-old woman was diagnosed with non-Hodgkin's lymphoma of the central nervous system, primary, the type of large, diffuse B cells after she had suffered from nausea, vomiting and vertigo for 4 months. Computed tomography (CT) and magnetic resonance imaging (MRI) showed a right cerebellar mass measuring 4 cm in diameter. The gross total resection of the mass was performed and the histopathological evaluation revealed a cerebral lymphoma. Systemic work, including whole body CT (excluding the brain), abdominal ultrasound, bone marrow biopsy, bone scan, and repeated examinations of cerebrospinal fluid showed no signs of lymphoma, confirming the diagnosis of PCNSL.

Post-surgical chemotherapy included 2 biweekly courses of methotrexate at high dose (MTX, 3.5 g / m2) plus rescue with leucovorin, and procarbazine (150 mg / m2 / day) administered for 14 days every 4 weeks. Due to hepatic dysfunction, procarbazine was discontinued and replaced with 1- (chloroethyl) -3-cyclohexyl-1-nitrosourea (CCNU, 100 mg / m2 for additional cycles) In parallel, intra-ventricular injections of ARA-C ( 50 mg once every 2 weeks) Immediately after this, the patient received cranial irradiation (4500 Cgy) distributed to the whole brain and a shot of 1080 Cgy distributed to the tumor bed before irradiation and termination of radiotherapy, repeated MRI studies showed no evidence of disease. However, 2 months later, a follow-up CRI evaluation, routine, showed a new mass in the posterior left lateral part of the medulla oblongata, which enlarged in the subsequent image formation. The patient experienced sensory loss related to the distribution of the first and second left branches of the trigeminal nerve. Systemic chemotherapy consisted of 6 cycles of CCNU (100 mg / m2 x 1) and etoposide (50 mg / day x 14). Then, MTX was administered at high dose (2 cycles of 3.5 mg / m2) plus rescue with leucovorin, and cisplatin (40 mg / m2 / day x 3) followed by high-dose, continuous ARA-C (1 g / m2 / day x 4). After completion of the last course of chemotherapy, MRI showed no resolution of brain stem mass. The salvage treatment with high-dose chemotherapy followed by autologous BMT was judged to be ineffective in view of the chemoresistant disease in this patient. Alo-BMT using its sister with identical HLA as a donor, was therefore implemented. The preimplant conditioning consisted of rabbit anti-human thymocyte globulin (ATG-Fresenius S, Bad Homburg, Germany, 10 mg / kg / day) from day -8 to day -5 and cyclophosphamide (50 mg / kg / day ) from day -4 to day -1. CSA (3 mg / kg / day) was administered intravenously from day -1 as the only prophylactic drug against GVHD. The patient received allogeneic peripheral blood totipotential cells (2.4 x 10c CD34 + cells / kg of a total number of 5.5 x 108 nucleated cells / kg) mobilized by subcutaneous injections of G-CSF at 10 μg / kg / day for 5 days). The graft was documented on day +12, with a white blood cell count of 1 x 109 / L and granulocyte count of >; 0.5 x 109 / L. From day +13 the platelet count was above 25 x 109 / L. The post-transplant course was uneven until day 10 when the patient developed grade II acute GVHD, involving the skin as manifested by a maculopapular itching (confirmed by skin biopsy) and the liver, as determined by total bilirubin. and slightly increased and lactate dehydrogenase (LDH). On day +12 ethylprednisolone (IV, 2 mg / kg) was added to cyclosporin. By day +18, the itching of the skin stopped and the liver function had begun to normalize. After resolution of GVHD, steroids and cyclosporine were decreased and completely removed at 3 months after transplantation. The chimerism studies were based solely on the DNA analysis of the variable number of tandem repeat markers (VNTR), since the donor and the patients were of the same gender and shared the same blood group. On day +18, the VNTR analysis revealed only hematopoietic cells derived from the donor. The MRI examination performed at the same time showed marked shrinkage of the left medullary mass in increase, and 3 months later the tumor transplant was no longer discernible. To date, 13 months after implantation of allogeneic totipotent cells, the patient is in good clinical condition (100% Karnofsky performance rating) without any signs of GVHD or lymphoma, demonstrating complete donor cell chimerism. The gradual manner in which the tumor resolved, is comparable to the remission associated with graft versus leukemia cell, slowly achieved, reported in patients with chronic myeloid leukemia after BMT. The anti-lymphoma effect was associated only with acute, moderate GVHD, which responded to the glucocorticoid and treatment with CSA without approval of the tumor effect. Thirteen months after the allo-PBSCT, the patient is in complete remission, showing complete hematopoiesis derived from the donor, after resolution of GVHD.

Example 4 Method 2: Conditioning Regimes (M / l) Two patients received an M / l conditioning regimen. Patient 1 was a 24-year-old male subject with CML in an accelerated phase. This was admitted for allogeneic BMT. The conditioning was based on myeloablation with dibromomannitol (DBM) of 40 mg / kg on days -9 to -7 and cytosine arabinoside (ARAc) 20 mg / kg on days -6 to -4. The patient was weakly immunosuppressed with cytoxan 50 mg / kg on days -3 to -1. The patient did not receive radiation at all and did not receive anti-lymphocyte globulin. The patient received totipotential cells mobilized by non-diminished G-CSF in T cells (9.8 x 108 cells / kg) of their sister. The procedure was unequal and mixed chimerism was confirmed by karyotype analysis. No GVHD was observed at any stage. Perhaps due to the lack of anti-host graft reactivity on the one hand and the advanced state of the disease on the other hand, the Ph clone did not disappear and eventually complete relapse was observed. The patient responded to a combination of cyclophosphamide (60 mg / kg for 2 days) and TBI 200 cGy x 6 (total dose of 1200 cGy) of conditioning, followed by an allogeneic totipotential cell transplant and is now completely grafted with 100% female cells and negative bcr / abl by reverse transcriptase PCR. Patient 2 was an 11-year-old girl, who was 2 years old when she was first treated with BMT. She was diagnosed with ß-thalassemia major, treated with BMT and then relapsed.

She received pre-transplant ambulatory conditioning of hydroxyurea 1500 mg / day for 6 days (myeloablation) followed by a single low dose of cyclophosphamide (750 mg) in parallel with the intravenous mesna (250 mg x 3 per 1 day) for the protection of the urinary bladder. On the day after the family cyclophos, the patient received on an ambulatory basis, totipotential blood cells mobilized with unmanipulated G-CSF (10 μg / kg for 5 days subcutaneously) to a total number of 8.74 x 108 nucleated cells / kg collected in three successive apheresis (Baxter CS 3000 Plus) from the original donor (the grandmother). The patient removed the cells from the residual host (most of the cells, consisting of 95% of the DNA) so that the patient was converted to 100% host and after four years is without any evidence of thalassemia or residual host cells.

Other Modalities Although the invention has been described with reference to the currently preferred embodiments, it should be understood that various modifications may be made without departing from the spirit of the invention. Other embodiments are within the following claims.

It is noted that in relation to this date, the best method known to the applicant to carry out the aforementioned invention, is that which is clear from the present description of the invention.

Having described the invention as above, property is claimed as contained in the following:

Claims (40)

1. A method for the treatment of a human patient having a disease of pathogenic cells, characterized in that the method comprises: a) the treatment of the patient with a conditioning regimen that preserves a functional population of the patient's hematopoietic totipotential cells; and b) administering a preparation comprising allogeneic totipotent cells from a donor to the patient, under conditions effective to induce the inability of the host antidonary response.

2. The method according to claim 1, characterized in that the regime is a conditioning regime m / L.

3. The method according to claim 1, characterized in that the regime is a conditioning regime - / L.

4. The method according to claim 1, characterized in that it further comprises the provision of allogeneic cell therapy to said patient, after the induction of the host's incapacity for anti-donor response and in the absence of significant GVHD.

5. The method according to claim 4, characterized in that the therapy with allogeneic cells comprises the administration of T lymphocytes of the donor in incremental increments, while controlling the GVHD without immunosuppression.

6. The method according to claim 5, characterized in that the T lymphocytes are of limited life time.

7. The method according to claim 5, characterized in that the T lymphocytes are CD8 + cells.

8. The method according to claim 5, characterized in that the T lymphocytes are CD4 + cells.

9. The method according to claim 1, characterized in that the allogeneic totipotent cells are selected from the group consisting of totipotential cells of peripheral blood, totipotential cells of umbilical cord blood, and totipotent cells of the bone marrow.

10. The method according to claim 1, characterized in that the conditioning regimen comprises the administration of one or more agents selected from the group consisting of purine analogues, alkylating agents, and anti-leukocyte globulins.

11. The method according to claim 10, characterized in that the purine analogue is fludarabine and the antileukocyte globulin is anti-i-1 globulin T infographics.

12. The method according to claim 11, characterized in that the regimen comprises the administration of fludarabine, anti-T-lymphocyte globulin and an alkylating agent.

13. The method according to claim 12, characterized in that the alkylating agent is busulfan.

14. The method according to claim 12, characterized in that the alkylating agent is cyclophosphamide.

15. The method according to claim 4, characterized in that the therapy with allogeneic cells comprises the administration of T lymphocytes of the donor, activated in vi tro before the administration to the patient.

16. The method according to claim 4, characterized in that the therapy with allogeneic cells comprises the in vi ve administration of the T cell activator to the patient.

17. The method according to claim 1, characterized in that the disease of pathogenic cells is a malignant disease.

18. The method according to claim 17, characterized in that the malignant disease is selected from the group consisting of chronic myelogenous leukemia, acute myelogenous leukemia, acute lymphoblastic leukemia, non-Hodgkin's lymphoma, myelodysplastic syndrome, and multiple myeloma.

19. The method according to claim 17, characterized in that the malignant disease is a solid tumor.

20. The method according to claim 1, characterized in that the disease of pathogenic cells is a non-malignant disease.

21. The method according to claim 20, characterized in that the non-malignant disease is selected from the group consisting of β-thalassemia major, Blackfan Diamond anemia, Gaucher anemia, Fanconi anemia and AIDS.

22. The method according to claim 20, characterized in that the non-malignant disease is an autoimmune disease.

23. A method for the treatment of a human patient having a disease of pathogenic cells, characterized in that the method comprises: a) treating the patient with a conditioning regimen that preserves a functional population of the patient's T lymphocytes; and b) administering a preparation comprising allogeneic totipotent cells from a donor to the patient, under conditions effective to induce the inability of host anti-donor response.

24. The method according to claim 23, characterized in that the regime is a conditioning regime M / -.

25. The method according to claim 23, characterized in that the regime is a conditioning regime M / l.

26. The method according to claim 23, characterized in that it further comprises the provision of a therapy regimen with allogeneic cells to the patient, after the induction of the inability of the host antidonary response, and in the absence of significant GVHD.

27. The method according to claim 23, characterized in that the regimen comprises the administration of an alkylating agent.

28. The method according to claim 27, characterized in that the alkylating agent is busulfan.

29. The method according to claim 27, characterized in that the alkylating agent is cyclophosphamide.

30. The method according to claim 23, characterized in that the regimen comprises the administration of irradiation to the whole body.

31. The method according to claim 30, characterized in that the regimen comprises the administration of cyclophosphamide.

32. The method according to claim 29, characterized in that the regimen further comprises hydroxyurea.

33. A method for preparing a preparation of allogeneic totipotential cells, specific for the patient, said patient having been administered with a conditioning regime that endows said patient with a selected veto capacity, characterized the method because it comprises: a) obtaining a preparation of totipotent cells of an allogeneic donor; and b) the adjustment of the veto capacity of the preparation to balance the patient's veto capacity.

34. The method according to claim 33, characterized in that it comprises adjusting the patient's veto capacity to balance the veto capacity of the preparation.

35. The use of allogeneic totipotent cells of a donor, in the manufacture of a medicament for the treatment of a disease of pathogenic cells, the medicament is administered to a host patient conditioned with a regimen that preserves a functional population of totipotent hematopoietic cells of the host, the host is administered with said medicament under effective conditions to induce the inability of host anti-donor response.

36. The use according to claim 35, wherein the medicament further comprises allogeneic lymphocytes.

37. The use of allogeneic totipotent cells of the donor, in the manufacture of a medicament for the treatment of a disease of pathogenic cells, the drug is administered to the host patient conditioned with a regimen that preserves a functional population of the T lymphocytes of the host, the host it is administered with a drug under effective conditions to induce the inability of the host's anti-donor response.

38. The use according to claim 37, wherein the medicament further comprises allogeneic lymphocytes.

39. The use of allogeneic lymphocytes of the donor, in the manufacture of a medicament for the treatment of a disease of pathogenic cells, the drug is administered to a host patient conditioned with a regimen that preserves a functional population of hematopoietic totipotential cells of the host, the host is administered with said medicament after the induction of the inability to respond antidonador.

40. The use of allogeneic lymphocytes of the donor in the manufacture of a medicament for the treatment of a disease of pathogenic cells, the medicament is administered to a host patient conditioned with a regimen that preserves a functional population of the lymphocytes of the host, the host is administered with the drug after the induction of inability of the host's anti-donor response.