WO2020158924A1

WO2020158924A1 - Method for inducing feto-maternal immune tolerance

Info

Publication number: WO2020158924A1
Application number: PCT/JP2020/003676
Authority: WO
Inventors: 克人玉井; 敬史新保; 木村　正; 誠之遠藤
Original assignee: 国立大学法人大阪大学
Priority date: 2019-01-31
Filing date: 2020-01-31
Publication date: 2020-08-06
Also published as: JPWO2020158924A1

Abstract

The present invention provides a pharmaceutical to be used to induce immune tolerance in a child to a non-inherited maternal antigen or an antigen that is the same as a non-inherited maternal antigen for the child by administration to a mother gestating a child.

Description

How to induce maternal-to-infant tolerance

The present application relates to a method for inducing maternal-to-infant immune tolerance.

In recent years, transplantation therapy for transplanting stem cells has been receiving attention. Further, it is known that stem cells induce immune tolerance. For example, a method of suppressing immunoreactivity of a graft by contacting perivascular cells (mesenchymal stem cells) of human umbilical cord with the graft. It has been proposed (see Patent Document 1).

However, the immune response is often a serious problem in cell transplantation and organ transplantation from a donor to a patient (recipient). Although cell therapy is one of the effective options for certain congenital diseases, it often causes an immune response because it introduces proteins and cells that the patient does not originally possess. In other words, the therapeutic drug becomes an antigen. When a patient develops a strong immune response, treatment must be abandoned.

On the other hand, due to improvements in fetal diagnosis technology, diseases that lead to fetal death during follow-up during pregnancy and congenital diseases that cannot be expected to survive or leave extremely serious damage during postnatal treatment are discovered during the fetal period. Is starting to appear.

In the fetal period, since immunity has not been established, it is relatively easy for other people's tissues and cells to be accepted, and treatment by bone marrow cell transplantation to the fetus has been proposed (see Non-Patent Document 3). Also, as one of the treatment methods for congenital diseases, there is a high expectation for fetal stem cell transplantation. For example, maternal hematopoietic stem cells are transplanted for the indication of abnormal hemoglobinopathy such as sickle cell disease and thalassemia. Has been studied (see Non-Patent Document 1). Further, treatment by transplantation of mesenchymal stem cells to a fetus with osteogenesis imperfecta has been reported (see Non-Patent Document 2).

Stem cells transplanted in the fetal period are relatively easy to engraft, and it is thought that immune tolerance to the stem cells and the substances produced based on them can be induced. If complete remission of the disease is difficult even with fetal treatment, continued treatment after birth is required, but if immune tolerance is induced based on fetal transplantation, treatment with the same cell transplant will be required. Can be continuously performed, and the prognosis of the patient can be improved.

Special table 2009-536163

The purpose of the present application is to provide a drug for inducing immune tolerance in a child against a non-genetic mother-derived antigen or the same antigen.

The present inventors bred female mice that express GFP only in mesenchymal stem cells and male wild-type mice. The HMGB1 peptide was administered to pregnant mother mice in order to recruit mesenchymal stem cells to the peripheral blood of pregnant mother mice. The skin of a GFPTg mouse, which expresses GFP in the cells of the whole body, was transplanted to the pups that did not express GFP. As a result, it was clarified that the graft survival rate was high in the offspring mice, and it was shown that immune tolerance to GFP, which is a non-genetic mother-derived antigen, was induced. Based on such findings, by increasing the number of stem cells in the blood of the mother gestating the offspring, the immune tolerance in the offspring to the non-genetic mother-derived antigen for the offspring derived from the stem cells or the same antigen as that A drug for inducing is provided.

That is, the present application provides the following.
[0]
A medicament for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen for the child or the same antigen by administration to a mother who is gestating the child, wherein bone marrow stem cells are used as peripheral blood. A medicament comprising a substance having an activity of mobilizing therein.
[1]
A medicament for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child by administration to a mother who is gestating the child, wherein bone marrow stem cells are used as peripheral blood. A medicament comprising a substance having an activity of mobilizing therein, wherein the bone marrow-derived stem cells of the mother have the non-genetic mother-derived antigen.
[2]
The medicament according to [1], which is a medicament for use in combination with another medicament for inducing immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as that.
[2-a]
A medicament for inducing immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the other medicament, wherein the other medicament contains isolated bone marrow-derived stem cells selected from the following: The drug according to [2]:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[2-b]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother gestating a child, wherein the mother's bone marrow-derived stem cell has a non-genetic mother-derived antigen for the child, and
(2) For the mother
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(B) by administration of an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother,
A combined drug for inducing immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the non-genetic mother, comprising the substance defined in (1) and the isolated bone marrow-derived stem cell defined in (2). A combination drug comprising:
[2-c]
The other drug induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into the offspring during fetal life, including isolated bone marrow-derived stem cells selected from The drug according to [2], which is a drug for:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[2-d]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother gestating a child, wherein the mother's bone marrow-derived stem cell has a non-genetic mother-derived antigen for the child, and
(2) For the child in the fetal period
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(B) by transplantation of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother,
A combined drug for inducing immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the non-genetic mother, comprising the substance defined in (1) and the isolated bone marrow-derived stem cell defined in (2). A combination drug comprising:
[2-e]
The other medicament induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into a neonatal offspring containing isolated bone marrow-derived stem cells selected from The drug according to [2], which is the drug:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen; and
(C) An isolated bone marrow-derived stem cell derived from an allogeneic donor to the mother and having the same antigen as the non-genetic mother-derived antigen.
[2-f]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother gestating a child, wherein the mother's bone marrow-derived stem cell has a non-genetic mother-derived antigen for the child, and
(2) For the newborn baby
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen,
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen, and
(C) by transplantation of an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor allogeneic to the mother,
A combined drug for inducing immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the non-genetic mother, comprising the substance defined in (1) and the isolated bone marrow-derived stem cell defined in (2). A combination drug comprising:
[3]
The mother who is gestating the child is a mother who is gestating a child with the disease, and the medicine is a medicine for use in combination with a medicine for treating the disease in the child, [1] The medicine according to.
[4]
A mother having a child with the disease is a mother without the disease, and a medicament for treating the disease in the child comprises isolated bone marrow-derived stem cells selected from The medicament according to [3], which is a medicament for treating the disease in the offspring by transplantation into the offspring during the fetal period:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. An isolated bone marrow-derived stem cell having the same antigen as.
[4-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who does not have the disease and is pregnant with a child who has the disease, wherein the bone marrow-derived stem cells of the mother are not Having a genetic mother-derived antigen, and
(2) For the child in the fetal period
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(B) an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. By porting
A combination drug for treating the disease in the offspring, which comprises the substance defined in (1) and the isolated bone marrow-derived stem cells defined in (2).
[5]
The mother having a child with the disease is a mother with the disease, and the medicament for treating the disease in the child is derived from a donor who is syngeneic with the mother and does not have the disease. [3] A medicament for treating the disease in the offspring by transplantation into the offspring during fetal life, which comprises an isolated bone marrow-derived stem cell having the same antigen as the non-genetic mother-derived antigen. The described medicine.
[5-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother having the disease who is pregnant with a child having the disease, wherein the bone marrow-derived stem cells of the mother are non-genetic mothers for the child Having a derived antigen, and
(2) by transplantation of an isolated bone marrow-derived stem cell derived from a donor who is syngeneic with the mother and has the same antigen as the non-genetic mother-derived antigen to the offspring in the fetal period,
A combination drug for treating the disease in the offspring, which comprises the substance defined in (1) and the isolated bone marrow-derived stem cells defined in (2).
[6]
An isolated bone marrow-derived stem cell, wherein a mother having a child with the disease is a mother without the disease, and a medicine for treating the disease in the child is selected from the following: The medicament according to [3], which is a medicament comprising an isolated tissue or an isolated organ for treating the disease in the offspring after the neonatal period:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(C) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is not allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[6-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who does not have the disease and is pregnant with a child who has the disease, wherein the bone marrow-derived stem cells of the mother are not Having a genetic mother-derived antigen, and
(2) For the child after the newborn period
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen,
(B) an isolated bone marrow-derived stem cell, isolated tissue, or isolated derived from a donor who is syngeneic with the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(C) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is not allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. Selected bone marrow-derived stem cells, isolated tissue or isolated organ transplant,
A combination drug for treating the disease in the offspring, comprising the substance defined in (1) and the isolated bone marrow-derived stem cells defined in (2), isolated tissue or isolated Combination medicine containing organs.
[7]
An isolated bone marrow-derived stem cell, wherein the mother having a child with the disease is a mother with the disease, and a medicine for treating the disease in the child is selected from the following: The medicament according to [3], which is a medicament for treating the disease in the offspring by transplanting into the offspring after the neonatal period, which comprises the isolated tissue or isolated organ:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(B) an isolated bone marrow-derived stem cell, an isolated tissue or an isolated tissue derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[7-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother having the disease who is pregnant with a child having the disease, wherein the bone marrow-derived stem cells of the mother are non-genetic mothers for the child Having a derived antigen, and
(2) For the child after the newborn period
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(B) an isolated bone marrow-derived stem cell, an isolated tissue or an isolated tissue derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. Organ
Selected from, by transplantation of isolated bone marrow-derived stem cells, isolated tissue or isolated organ,
A combination drug for treating the disease in the offspring, comprising the substance defined in (1) and the isolated bone marrow-derived stem cells defined in (2), isolated tissue or isolated Combination medicine containing organs.
[8]
A medicament for treating the disease in the offspring comprises the same isolated protein as the non-genetic mother-derived antigen for treating the disease in the offspring by administration to the offspring after neonatal period. The medicine according to [3], which is a medicine.
[8-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who is pregnant with a diseased child, wherein the bone marrow-derived stem cell of the mother has a non-genetic mother-derived antigen for the child ,and,
(2) administration of the same isolated protein as the non-genetic mother-derived antigen to the offspring after neonatal period,
A combination drug for treating the disease in the offspring, comprising the substance defined in (1) and the isolated protein defined in (2).
[9]
A medicament for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child when administered to a mother who is gestating the child, selected from the following: A medicament containing isolated bone marrow-derived stem cells:
(A) an isolated bone marrow-derived stem cell derived from a mother who is gestating the offspring and having a non-genetic mother-derived antigen for the offspring;
(B) An isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen.
[10]
The medicament according to [9], which is a medicament for use in combination with another medicament for inducing immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the non-genetic mother.
[10-a]
The other drug induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into the offspring during fetal life, including isolated bone marrow-derived stem cells selected from The drug according to [10], which is a drug for:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[10-b]
(1) Administration of an isolated bone marrow-derived stem cell selected from the following (a) and (b) to a mother gestating a child, and
(2) By transplanting the isolated bone marrow-derived stem cells selected from the following (a) and (b) to the fetus in the offspring,
A combination drug for inducing immunological tolerance in a child against a non-genetic mother-derived antigen for the child or the same antigen as the child, comprising the isolated bone marrow-derived stem cells defined in (1) and (2) A combination drug comprising the defined isolated bone marrow-derived stem cells:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[10-c]
The other medicament induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into a neonatal offspring containing isolated bone marrow-derived stem cells selected from The drug according to [10], which is the drug:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen; and
(C) An isolated bone marrow-derived stem cell derived from an allogeneic donor to the mother and having the same antigen as the non-genetic mother-derived antigen.
[10-d]
(1) Administration of an isolated bone marrow-derived stem cell selected from the following (a) and (b) to a mother gestating a child, and
(2) By transplanting the isolated bone marrow-derived stem cells selected from the following (a) to (c) to the neonatal period,
A combination drug for inducing immunological tolerance in a child against a non-genetic mother-derived antigen for the child or the same antigen as the child, comprising the isolated bone marrow-derived stem cells defined in (1) and (2) A combination drug comprising the defined isolated bone marrow-derived stem cells:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen; and
(C) An isolated bone marrow-derived stem cell derived from an allogeneic donor to the mother and having the same antigen as the non-genetic mother-derived antigen.
[11]
The mother who is gestating the child is a mother who is gestating a child with the disease, and the medicine is a medicine for use in combination with a medicine for treating the disease in the child, [9] The medicine according to.
[12]
A mother having a child with the disease is a mother without the disease, and a medicament for treating the disease in the child comprises isolated bone marrow-derived stem cells selected from The medicament according to [11], which is a medicament for treating the disease in the offspring by transplantation to the offspring in a fetal period:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. An isolated bone marrow-derived stem cell having the same antigen as.
[12-a]
(1) For mothers without the disease who are pregnant with a child with the disease
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) For the child in the fetal period
(I) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(Ii) an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor who does not have the disease and is syngeneic with the mother By porting
A combination drug for treating the disease in the offspring, which comprises the isolated bone marrow-derived stem cells defined in (1) and the isolated bone marrow-derived stem cells defined in (2).
[13]
The mother having a child with the disease is a mother with the disease, and the medicament for treating the disease in the child is derived from a donor who is syngeneic with the mother and does not have the disease. A pharmaceutical for treating the disease in the offspring by transplantation to the offspring in a prenatal period, which comprises an isolated bone marrow-derived stem cell having the same antigen as the non-genetic mother-derived antigen, [11] The described medicine.
[13-a]
(1) For mothers with a disease who are pregnant with a child with the disease
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) by transplantation of an isolated bone marrow-derived stem cell derived from a donor who is syngeneic with the mother and has the same antigen as the non-genetic mother-derived antigen to the offspring in the fetal period,
A combination drug for treating the disease in the offspring, which comprises the isolated bone marrow-derived stem cells defined in (1) and the isolated bone marrow-derived stem cells defined in (2).
[14]
A mother having a child with the disease is a mother without the disease, and a drug for treating the disease in the child has isolated bone marrow-derived stem cells selected from the following: The medicament according to [11], which is a medicament containing a separated tissue or an isolated organ for treating the disease in the offspring after the neonatal period:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(C) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is not allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[14-a]
(1) For mothers without the disease who are pregnant with a child with the disease
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) For the child after the newborn period
(I) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, an isolated tissue or an isolated organ,
(Ii) isolated bone marrow-derived stem cells, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(Iii) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is allogeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen. By transplantation of isolated bone marrow-derived stem cells, isolated tissues or isolated organs selected from
A combined medicament for treating the disease in the offspring, comprising isolated bone marrow-derived stem cells defined in (1) and isolated bone marrow-derived stem cells defined in (2), isolated A combination drug comprising a tissue or an isolated organ.
[15]
A mother having a child with the disease is a mother with the disease, and a medicament for treating the disease in the child is an isolated bone marrow-derived stem cell selected from the following: The medicament according to [11], which is a medicament for treating the disease in the offspring by transplanting into the offspring after the neonatal period, including the isolated tissue or the isolated organ:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(B) an isolated bone marrow-derived stem cell, an isolated tissue or an isolated tissue derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[15-a]
(1) For mothers with a disease who are pregnant with a child with the disease
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) For the child after the newborn period
(I) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(Ii) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. By transplantation of isolated bone marrow-derived stem cells, isolated tissues or isolated organs selected from
A combined medicament for treating the disease in the offspring, comprising isolated bone marrow-derived stem cells defined in (1) and isolated bone marrow-derived stem cells defined in (2), isolated A combination drug comprising a tissue or an isolated organ.
[16]
A medicament for treating the disease in the offspring comprises the same isolated protein as the non-genetic mother-derived antigen for treating the disease in the offspring by administration to the offspring after neonatal period. The medicine according to [11], which is a medicine.
[16-a]
(1) For mothers who are pregnant with sick children
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) administration of the same isolated protein as the non-genetic mother-derived antigen to the offspring after neonatal period,
A combination drug for treating the disease in the offspring, which comprises the isolated bone marrow-derived stem cells defined in (1) and the isolated protein defined in (2).
[17]
[3] to [8] and [11] to [16], wherein the disease is treatable as a result of induction of immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as that. The drug according to any one of claims.
[18]
[3] to [8] and [11] to [16], wherein the disease is treatable as a result of suppressing an immune reaction in the offspring to the non-genetic mother-derived antigen or the same antigen as that. The drug according to any one of claims.
[19]
The medicament according to any one of [1] to [18], wherein the stem cells are mesenchymal stem cells or hematopoietic stem cells.
[20]
The medicament according to any one of [1] to [18], wherein the stem cells are mesenchymal stem cells.
[21]
A medicinal stem cell that is used for inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child by administration to a mother who is gestating the child. A medicament comprising a substance having an activity of mobilizing in peripheral blood, wherein the bone marrow-derived stem cells of the mother have the non-genetic mother-derived antigen.
[22]
A medicament for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child when administered to a mother who is gestating the child, selected from the following: A medicament containing isolated mesenchymal stem cells:
(A) an isolated mesenchymal stem cell derived from a mother who is gestating the offspring and having a non-genetic mother-derived antigen for the offspring;
(B) An isolated mesenchymal stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen.
[A0]
A method of inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child, comprising:
A method comprising the step of administering an effective amount of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood, to a mother who is pregnant with a child.
[A1]
A method of inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child, comprising:
A step of administering an effective amount of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood, to a mother who is gestating a child, wherein bone marrow-derived stem cells of the mother are derived from a non-genetic mother A method comprising the step of having an antigen.
[A2]
The method according to [A1], which further comprises a step of using the substance in combination with another drug for inducing immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the non-genetic mother.
[A2-a]
A medicament for inducing immune tolerance in the offspring against the non-genetic mother-derived antigen or the same antigen as the other medicament, wherein the other medicament comprises isolated bone marrow-derived stem cells selected from the following: The method according to [A2]:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[A2-b]
A method of inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child, comprising:
(1) A step of administering an effective amount of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who is gestating a child, wherein the bone marrow-derived stem cells of the mother are Having a genetic mother-derived antigen, and
(2) For the mother,
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(B) a step of administering an effective amount of an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother Including the method.
[A2-c]
The other medicament induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into the offspring in the fetal period, which comprises isolated bone marrow-derived stem cells selected from The method according to [A2], which is a drug for:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[A2-d]
A method of inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child, comprising:
(1) A step of administering an effective amount of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who is gestating a child, wherein the bone marrow-derived stem cells of the mother are Having a genetic mother-derived antigen, and
(2) For the child in the fetal period,
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(B) a step of transplanting an effective amount of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother Including the method.
[A2-e]
The other medicament induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into a neonatal offspring containing isolated bone marrow-derived stem cells selected from: The method according to [A2], which is the drug:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen; and
(C) An isolated bone marrow-derived stem cell derived from an allogeneic donor to the mother and having the same antigen as the non-genetic mother-derived antigen.
[A2-f]
A method of inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child, comprising:
(1) A step of administering an effective amount of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who is gestating a child, wherein the bone marrow-derived stem cells of the mother are Having a genetic mother-derived antigen, and
(2) For the newborn baby,
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen,
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen, and
(C) transplant an effective amount of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having an antigen identical to the non-genetic mother-derived antigen, which is derived from a donor allogeneic to the mother A method including steps.
[A3]
The mother who is gestating the child is a mother who is gestating a child with the disease, and further comprises the step of using the substance in combination with a drug for treating the disease in the child [A1]. The method described in.
[A4]
A mother having a child with the disease is a mother without the disease, and a medicament for treating the disease in the child comprises isolated bone marrow-derived stem cells selected from The method according to [A3], which is a medicament for treating the disease in the offspring by transplantation into the offspring.
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. An isolated bone marrow-derived stem cell having the same antigen as.
[A4-a]
A method of treating a disease in a child, comprising:
(1) A step of administering an effective amount of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood, to a mother who does not have the disease and is pregnant with a child who has the disease. Bone marrow-derived stem cells having a non-genetic mother-derived antigen for the offspring, and
(2) For the child in the fetal period,
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(B) an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease A method comprising the step of implanting an effective amount.
[A5]
The mother having a child with the disease is a mother with the disease, and the medicament for treating the disease in the child is derived from a donor who is syngeneic with the mother and does not have the disease. A drug for treating the disease in the offspring by transplantation into the offspring at the fetal stage, comprising isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, [A3] The method described.
[A5-a]
A method of treating a disease in a child, comprising:
(1) A step of administering an effective amount of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood, to a mother having the disease who is pregnant with a child having the disease, the method being derived from bone marrow of the mother Stem cells having a non-genetic mother-derived antigen for the offspring, and
(2) Transplanting an effective amount of isolated bone marrow-derived stem cells derived from a donor free of the disease that is syngeneic to the mother to the offspring in the fetal period and has the same antigen as the non-genetic mother-derived antigen A method including steps.
[A6]
An isolated bone marrow-derived stem cell, wherein a mother having a child with the disease is a mother without the disease, and a medicine for treating the disease in the child is selected from the following: The method according to [A3], which is a medicament comprising an isolated tissue or an isolated organ for treating the disease in the offspring by transplantation to the offspring after the neonatal period:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(C) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is not allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[A6-a]
A method of treating a disease in a child, comprising:
(1) A step of administering an effective amount of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood, to a mother not having the disease who is pregnant with a child having the disease, Bone marrow-derived stem cells having a non-genetic mother-derived antigen for the offspring, and
(2) For the child after the neonatal period,
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen,
(B) an isolated bone marrow-derived stem cell, isolated tissue, or isolated derived from a donor who is syngeneic with the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(C) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is not allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells, isolated tissue or an effective amount of the isolated organ selected from the above-mentioned organs.
[A7]
An isolated bone marrow-derived stem cell, wherein the mother having a child with the disease is a mother with the disease, and a medicine for treating the disease in the child is selected from the following: The method according to [A3], which is a medicament for treating the disease in the offspring by transplanting into the offspring after the neonatal period, which comprises the isolated tissue or the isolated organ:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(B) an isolated bone marrow-derived stem cell, an isolated tissue or an isolated tissue derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[A7-a]
A method of treating a disease in a child, comprising:
(1) A step of administering an effective amount of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood, to a mother having the disease who is pregnant with a child having the disease, the bone marrow derived from the mother Stem cells having a non-genetic mother-derived antigen for the offspring, and
(2) For the child after the neonatal period,
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(B) an isolated bone marrow-derived stem cell, an isolated tissue or an isolated tissue derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. Organ
A method comprising the step of transplanting an isolated bone marrow-derived stem cell, an isolated tissue or an isolated organ in an effective amount selected from
[A8]
A medicament for treating the disease in the offspring comprises the same isolated protein as the non-genetic mother-derived antigen for treating the disease in the offspring by administration to the offspring after neonatal period. The method according to [A3], which is a medicine.
[A8-a]
A method of treating a disease in a child, comprising:
(1) A step of administering an effective amount of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who is pregnant with a diseased child, wherein the mother's bone marrow-derived stem cells are Having a non-genetic mother-derived antigen for
(2) A method comprising the step of administering to the offspring after the neonatal period an effective amount of the same isolated protein as the non-genetic mother-derived antigen.
[A9]
A method of inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child, comprising:
A method comprising the step of administering to a mother having a baby an effective amount of isolated bone marrow-derived stem cells selected from:
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring; and
(B) An isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen.
[A10]
The method according to [A9], further comprising the step of using the isolated bone marrow-derived stem cells in combination with another drug for inducing immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as that antigen. ..
[A10-a]
The other drug induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into the offspring during fetal life, including isolated bone marrow-derived stem cells selected from The method according to [A10], which is a drug for:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[A10-b]
A method of inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child, comprising:
(1) a step of administering an effective amount of the isolated bone marrow-derived stem cells selected from the following (a) and (b) to a mother gestating a baby, and
(2) A method comprising the step of transplanting an effective amount of the isolated bone marrow-derived stem cells selected from the following (a) and (b) to the fetus in the fetal period:
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring; and
(B) An isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen.
[A10-c]
The other medicament induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into a neonatal offspring containing isolated bone marrow-derived stem cells selected from The method according to [A10], which is the drug:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen; and
(C) An isolated bone marrow-derived stem cell derived from an allogeneic donor to the mother and having the same antigen as the non-genetic mother-derived antigen.
[A10-d]
A method of inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child, comprising:
(1) a step of administering an effective amount of the isolated bone marrow-derived stem cells selected from the following (a) and (b) to a mother gestating a baby, and
(2) A method comprising a step of transplanting an effective amount of the isolated bone marrow-derived stem cells selected from the following (a) to (c) to the newborn infant:
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring;
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen; and
(C) An isolated bone marrow-derived stem cell derived from an allogeneic donor to the mother and having the same antigen as the non-genetic mother-derived antigen.
[A11]
A step of using the isolated bone marrow-derived stem cells in combination with a medicine for treating the disease in the offspring, wherein the mother who is gestating the offspring is the mom who is gestating the diseased child. The method according to [A9], further comprising:
[A12]
A mother having a child with the disease is a mother without the disease, and a medicament for treating the disease in the child comprises isolated bone marrow-derived stem cells selected from The method according to [A11], which is a medicament for treating the disease in the offspring by transplanting the offspring into the offspring.
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. An isolated bone marrow-derived stem cell having the same antigen as.
[A12-a]
A method of treating a disease in a child, comprising:
(1) For mothers without the disease who are pregnant with a child with the disease,
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) a step of administering an effective amount of an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother ,and,
(2) For the child in the fetal period,
(I) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(Ii) an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor who does not have the disease and is syngeneic with the mother A method comprising the step of implanting an effective amount.
[A13]
The mother having a child with the disease is a mother with the disease, and the medicament for treating the disease in the child is derived from a donor who is syngeneic with the mother and does not have the disease. , A medicament for treating the disease in the offspring by transplantation into the offspring during fetal life, which comprises an isolated bone marrow-derived stem cell having the same antigen as the non-genetic mother-derived antigen, [A11] The method described.
[A13-a]
A method of treating a disease in a child, comprising:
(1) For a mother with the disease who is pregnant with a child with the disease,
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) a step of administering an effective amount of an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother ,and,
(2) An effective amount of the isolated bone marrow-derived stem cells derived from a donor who is syngeneic to the mother and has no disease with respect to the offspring in the fetal period, and has the same antigen as the non-genetic mother-derived antigen A method comprising the step of transplanting.
[A14]
A mother having a child with the disease is a mother without the disease, and a drug for treating the disease in the child has isolated bone marrow-derived stem cells selected from the following: The method according to [A11], which is a medicament for treating the disease in the offspring by transplanting into the offspring after neonatal period, which comprises isolated tissue or isolated organs:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(C) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is not allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[A14-a]
A method of treating a disease in a child, comprising:
(1) For mothers without the disease who are pregnant with a child with the disease,
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) a step of administering an effective amount of an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother ,and,
(2) For the child after the neonatal period,
(I) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, an isolated tissue or an isolated organ,
(Ii) isolated bone marrow-derived stem cells, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(Iii) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is allogeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen. Bone marrow-derived stem cells selected from selected organs, isolated tissues or isolated organs.
[A15]
A mother having a child with the disease is a mother with the disease, and a medicament for treating the disease in the child is an isolated bone marrow-derived stem cell selected from the following: The method according to [A11], which is a medicament for treating the disease in the offspring by transplanting into the offspring after the neonatal period, including the isolated tissue or the isolated organ:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(B) an isolated bone marrow-derived stem cell, an isolated tissue or an isolated tissue derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[A15-a]
A method of treating a disease in a child, comprising:
(1) For a mother with the disease who is pregnant with a child with the disease,
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) a step of administering an effective amount of an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother ,and,
(2) For the child after the neonatal period,
(I) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(Ii) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. Bone marrow-derived stem cells selected from selected organs, isolated tissues or isolated organs.
[A16]
A medicament for treating the disease in the offspring comprises the same isolated protein as the non-genetic mother-derived antigen for treating the disease in the offspring by administration to the offspring after neonatal period. The method according to [A11], which is a medicine.
[A16-a]
A method of treating a disease in a child, comprising:
(1) For mothers who are pregnant with a sick child,
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) a step of administering an effective amount of an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother ,and,
(2) A method comprising the step of administering to the offspring after the neonatal period the same isolated protein as the non-genetic mother-derived antigen.
[A17]
[A3] to [A8] and [A11] to [A16], which is treatable as a result of induction of immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as that of the non-genetic mother. The method described in either.
[A18]
[A3] to [A8] and [A11] to [A16], which is treatable as a result of suppressing the immune reaction in the offspring against the non-genetic mother-derived antigen or the same antigen as the non-genetic mother. The method described in either.
[A19]
The method according to any one of [A1] to [A18], wherein the stem cells are mesenchymal stem cells or hematopoietic stem cells.
[A20]
The method according to any one of [A1] to [A18], wherein the stem cells are mesenchymal stem cells.
[A21]
A method of inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child, comprising:
A step of administering an effective amount of a substance having an activity of mobilizing mesenchymal stem cells into peripheral blood, to a mother who is gestating a child, wherein bone marrow-derived stem cells of the mother are non-genetic. A method comprising the step of having a mother-derived antigen.
[A22]
A method of inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child, comprising:
A method comprising the step of administering to a mother having pups an effective amount of isolated mesenchymal stem cells selected from:
(A) an isolated mesenchymal stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring; and
(B) An isolated mesenchymal stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen.
[B0]
Administration of the offspring to a gestating mother mobilizes bone marrow stem cells into peripheral blood for use in inducing tolerance in the offspring to a non-genetic mother-derived antigen or the same antigen for the offspring. A substance that has activity.
[B1]
Administration of the offspring to a gestating mother mobilizes bone marrow stem cells into peripheral blood for use in inducing tolerance in the offspring to a non-genetic mother-derived antigen or the same antigen for the offspring. A substance having an activity, wherein said mother's bone marrow-derived stem cells have said non-genetic mother-derived antigen.
[B2]
The substance according to [B1], wherein the substance is a substance for use in combination with another drug for inducing immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the non-genetic mother antigen.
[B2-a]
A medicament for inducing immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the other medicament, wherein the other medicament contains isolated bone marrow-derived stem cells selected from the following: And the substance according to [B2]:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[B2-b]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother gestating a child, wherein the mother's bone marrow-derived stem cell has a non-genetic mother-derived antigen for the child, and
(2) For the mother
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(B) by administration of an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother,
A substance as defined in (1) and an isolated bone marrow-derived stem cell as defined in (2) for use in inducing immune tolerance in said offspring to said non-genetic mother-derived antigen or the same antigen as that A combination.
[B2-c]
The other drug induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into the offspring during fetal life, including isolated bone marrow-derived stem cells selected from The substance according to [B2], which is a drug for:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[B2-d]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother gestating a child, wherein the mother's bone marrow-derived stem cell has a non-genetic mother-derived antigen for the child, and
(2) For the child in the fetal period
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(B) by transplantation of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother,
A substance as defined in (1) and an isolated bone marrow-derived stem cell as defined in (2) for use in inducing immune tolerance in said offspring to said non-genetic mother-derived antigen or the same antigen as that A combination.
[B2-e]
The other medicament induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into a neonatal offspring containing isolated bone marrow-derived stem cells selected from The substance according to [B2], which is a drug of:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen; and
(C) An isolated bone marrow-derived stem cell derived from an allogeneic donor to the mother and having the same antigen as the non-genetic mother-derived antigen.
[B2-f]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother gestating a child, wherein the mother's bone marrow-derived stem cell has a non-genetic mother-derived antigen for the child, and
(2) For the newborn baby
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen,
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen, and
(C) by transplantation of an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor allogeneic to the mother,
A substance as defined in (1) and an isolated bone marrow-derived stem cell as defined in (2) for use in inducing immune tolerance in said offspring to said non-genetic mother-derived antigen or the same antigen as that A combination.
[B3]
The mother who is gestating the child is a mother who is gestating a child with the disease, and the substance is a substance to be used in combination with a medicine for treating the disease in the child, [B1]. Substances listed in.
[B4]
A mother having a child with the disease is a mother without the disease, and a medicament for treating the disease in the child comprises isolated bone marrow-derived stem cells selected from The substance according to [B3], which is a drug for treating the disease in the offspring by transplantation into the offspring during fetal period:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. An isolated bone marrow-derived stem cell having the same antigen as.
[B4-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who does not have the disease and is pregnant with a child who has the disease, wherein the bone marrow-derived stem cells of the mother are not Having a genetic mother-derived antigen, and
(2) For the child in the fetal period
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(B) an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. By porting
A combination comprising a substance as defined in (1) and isolated bone marrow-derived stem cells as defined in (2) for use in treating the disease in the offspring.
[B5]
The mother having a child with the disease is a mother with the disease, and the medicament for treating the disease in the child is derived from a donor who is syngeneic with the mother and does not have the disease. [B3] which is a medicine for treating the disease in the offspring by transplantation to the offspring in the fetal period, which comprises an isolated bone marrow-derived stem cell having the same antigen as the non-genetic mother-derived antigen. Listed substances.
[B5-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother having the disease who is pregnant with a child having the disease, wherein the bone marrow-derived stem cells of the mother are non-genetic mothers for the child Having a derived antigen, and
(2) by transplantation of an isolated bone marrow-derived stem cell derived from a donor who is syngeneic with the mother and has the same antigen as the non-genetic mother-derived antigen to the offspring in the fetal period,
A combination comprising a substance as defined in (1) and isolated bone marrow-derived stem cells as defined in (2) for use in treating the disease in the offspring.
[B6]
An isolated bone marrow-derived stem cell, wherein a mother having a child with the disease is a mother without the disease, and a medicine for treating the disease in the child is selected from the following: The substance according to [B3], which is a medicament containing an isolated tissue or an isolated organ for treating the disease in the offspring after the neonatal period:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(C) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is not allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[B6-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who does not have the disease and is pregnant with a child who has the disease, wherein the bone marrow-derived stem cells of the mother are not Having a genetic mother-derived antigen, and
(2) For the child after the newborn period
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen,
(B) an isolated bone marrow-derived stem cell, isolated tissue, or isolated derived from a donor who is syngeneic with the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(C) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is not allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. Selected bone marrow-derived stem cells, isolated tissue or isolated organ transplant,
A substance defined in (1) and an isolated bone marrow-derived stem cell defined in (2), an isolated tissue or an isolated organ for use in treating the disease in the offspring. A combination that includes.
[B7]
An isolated bone marrow-derived stem cell, wherein the mother having a child with the disease is a mother with the disease, and a medicine for treating the disease in the child is selected from the following: A substance according to [B3], which is a medicament for treating the disease in the offspring by transplanting into the offspring after the neonatal period, which comprises the isolated tissue or the isolated organ:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(B) an isolated bone marrow-derived stem cell, an isolated tissue or an isolated tissue derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[B7-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother having the disease who is pregnant with a child having the disease, wherein the bone marrow-derived stem cells of the mother are non-genetic mothers for the child Having a derived antigen, and
(2) For the child after the newborn period
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(B) an isolated bone marrow-derived stem cell, an isolated tissue or an isolated tissue derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. Organ
Selected from, by transplantation of isolated bone marrow-derived stem cells, isolated tissue or isolated organ,
A substance defined in (1) and an isolated bone marrow-derived stem cell defined in (2), an isolated tissue or an isolated organ for use in treating the disease in the offspring. A combination that includes.
[B8]
A medicament for treating the disease in the offspring comprises the same isolated protein as the non-genetic mother-derived antigen for treating the disease in the offspring by administration to the offspring after neonatal period. The substance according to [B3], which is a medicine.
[B8-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who is pregnant with a diseased child, wherein the bone marrow-derived stem cell of the mother has a non-genetic mother-derived antigen for the child ,and,
(2) administration of the same isolated protein as the non-genetic mother-derived antigen to the offspring after neonatal period,
A combination comprising a substance as defined in (1) and an isolated protein as defined in (2) for use in treating the disease in said offspring.
[B9]
For use in inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen for said child by administration to a mother who is gestating the child, selected from the following: Bone marrow-derived stem cells:
(A) an isolated bone marrow-derived stem cell derived from a mother who is gestating the offspring and having a non-genetic mother-derived antigen for the offspring;
(B) An isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen.
[B10]
The cell according to [B9], wherein the cell is a cell for use in combination with another drug for inducing immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the non-genetic mother.
[B10-a]
The other drug induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into the offspring during fetal life, including isolated bone marrow-derived stem cells selected from The cell according to [B10], which is a medicament for:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[B10-b]
(1) Administration of an isolated bone marrow-derived stem cell selected from the following (a) and (b) to a mother gestating a child, and
(2) By transplanting the isolated bone marrow-derived stem cells selected from the following (a) and (b) to the fetus in the offspring,
An isolated bone marrow-derived stem cell as defined in (1) and defined in (2) for use in inducing tolerance in a non-genetic mother-derived antigen or the same antigen for said child in said child A combination comprising isolated bone marrow-derived stem cells:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[B10-c]
The other medicament induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into a neonatal offspring containing isolated bone marrow-derived stem cells selected from The cell according to [B10], which is the drug of:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen; and
(C) An isolated bone marrow-derived stem cell derived from an allogeneic donor to the mother and having the same antigen as the non-genetic mother-derived antigen.
[B10-d]
(1) Administration of an isolated bone marrow-derived stem cell selected from the following (a) and (b) to a mother gestating a child, and
(2) By transplanting the isolated bone marrow-derived stem cells selected from the following (a) to (c) to the neonatal period,
An isolated bone marrow-derived stem cell as defined in (1) and defined in (2) for use in inducing tolerance in a non-genetic mother-derived antigen or the same antigen for said child in said child A combination comprising isolated bone marrow-derived stem cells:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen; and
(C) An isolated bone marrow-derived stem cell derived from an allogeneic donor to the mother and having the same antigen as the non-genetic mother-derived antigen.
[B11]
The mother who is gestating the child is a mother who is gestating a child with the disease, and the medicine is a medicine for use in combination with a medicine for treating the disease in the child, [B9]. The cell according to.
[B12]
A mother having a child with the disease is a mother without the disease, and a medicament for treating the disease in the child comprises isolated bone marrow-derived stem cells selected from The cell according to [B11], which is a drug for treating the disease in the offspring by transplantation into the offspring during fetal period:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. An isolated bone marrow-derived stem cell having the same antigen as.
[B12-a]
(1) For mothers without the disease who are pregnant with a child with the disease
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) For the child in the fetal period
(I) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(Ii) an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor who does not have the disease and is syngeneic with the mother By porting
A combination comprising the isolated bone marrow-derived stem cells defined in (1) and the isolated bone marrow-derived stem cells defined in (2) for use in treating the disease in the offspring.
[B13]
The mother having a child with the disease is a mother with the disease, and the medicament for treating the disease in the child is derived from a donor who is syngeneic with the mother and does not have the disease. [B11] is a medicine for treating the disease in the offspring by transplantation into the offspring during fetal life, which comprises an isolated bone marrow-derived stem cell having the same antigen as the non-genetic mother-derived antigen. The described cell.
[B13-a]
(1) For mothers with a disease who are pregnant with a child with the disease
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) by transplantation of an isolated bone marrow-derived stem cell derived from a donor who is syngeneic with the mother and has the same antigen as the non-genetic mother-derived antigen to the offspring in the fetal period,
A combination comprising the isolated bone marrow-derived stem cells defined in (1) and the isolated bone marrow-derived stem cells defined in (2) for use in treating the disease in the offspring.
[B14]
A mother having a child with the disease is a mother without the disease, and a drug for treating the disease in the child has isolated bone marrow-derived stem cells selected from the following: The cell according to [B11], which is a medicament for treating the disease in the offspring by transplantation into the offspring after the neonatal period, including isolated tissue or isolated organ:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(C) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is not allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[B14-a]
(1) For mothers without the disease who are pregnant with a child with the disease
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) For the child after the newborn period
(I) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, an isolated tissue or an isolated organ,
(Ii) isolated bone marrow-derived stem cells, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(Iii) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is allogeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen. By transplantation of isolated bone marrow-derived stem cells, isolated tissues or isolated organs selected from
An isolated bone marrow-derived stem cell as defined in (1) and an isolated bone marrow-derived stem cell as defined in (2), an isolated tissue, or for use in treating the disease in said offspring. A combination comprising isolated organs.
[B15]
A mother having a child with the disease is a mother with the disease, and a medicament for treating the disease in the child is an isolated bone marrow-derived stem cell selected from the following: The cell according to [B11], which is a medicament for treating the disease in the offspring, which comprises the isolated tissue or the isolated organ, and is transplanted to the offspring after the neonatal period:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(B) an isolated bone marrow-derived stem cell, an isolated tissue or an isolated tissue derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[B15-a]
(1) For mothers with a disease who are pregnant with a child with the disease
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) For the child after the newborn period
(I) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(Ii) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. By transplantation of isolated bone marrow-derived stem cells, isolated tissues or isolated organs selected from
An isolated bone marrow-derived stem cell as defined in (1) and an isolated bone marrow-derived stem cell as defined in (2), an isolated tissue, or for use in treating the disease in said offspring. A combination comprising isolated organs.
[B16]
A medicament for treating the disease in the offspring comprises the same isolated protein as the non-genetic mother-derived antigen for treating the disease in the offspring by administration to the offspring after neonatal period. The cell according to [B11], which is a medicine.
[B16-a]
(1) For mothers who are pregnant with sick children
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) administration of the same isolated protein as the non-genetic mother-derived antigen to the offspring after neonatal period,
A combination comprising the isolated bone marrow-derived stem cells defined in (1) and the isolated protein defined in (2) for use in treating the disease in the offspring.
[B17]
[B3] to [B8] and [B11] to [B16], which are treatable as a result of induction of immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the non-genetic mother. The cell according to any of the above.
[B18]
[B3] to [B8] and [B11] to [B16], wherein the disease is treatable as a result of suppression of immune reaction in the offspring to the non-genetic mother-derived antigen or the same antigen as that The cell according to any of the above.
[B19]
The cell according to any one of [B1] to [B18], wherein the stem cell is a mesenchymal stem cell or a hematopoietic stem cell.
[B20]
The cell according to any one of [B1] to [B18], wherein the stem cell is a mesenchymal stem cell.
[B21]
Mesenchymal stem cells for use in inducing tolerance in a non-genetic mother-derived antigen for the offspring or the same antigen as that of the offspring by administration to the mother of the offspring into peripheral blood. A substance having a mobilizing activity, wherein the bone marrow-derived stem cells of the mother have the non-genetic mother-derived antigen.
[B22]
For use in inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen for said child by administration to a mother who is gestating the child, selected from the following: Mesenchymal stem cells:
(A) an isolated mesenchymal stem cell derived from a mother who is gestating the offspring and having a non-genetic mother-derived antigen for the offspring;
(B) An isolated mesenchymal stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen.
[C0]
Bone marrow stem cells in the manufacture of a medicament for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen for the child or the same antigen by administration to a mother who is gestating the child. Use of substances with active mobilization in.
[C1]
Bone marrow stem cells in the manufacture of a medicament for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen for the child or the same antigen by administration to a mother who is gestating the child. Use of a substance having mobilizing activity therein, wherein the bone marrow-derived stem cells of the mother carry the non-genetic mother-derived antigen.
[C2]
The use according to [C1], wherein the drug is a drug for use in combination with another drug for inducing immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as that.
[C2-a]
A medicament for inducing immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the other medicament, wherein the other medicament contains isolated bone marrow-derived stem cells selected from the following: The use according to [C2]:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[C2-b]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother gestating a child, wherein the mother's bone marrow-derived stem cell has a non-genetic mother-derived antigen for the child, and
(2) For the mother
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(B) by administration of an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother,
The substance defined in (1) and the isolated bone marrow-derived stem cell defined in (2) in the manufacture of a combined drug for inducing immunological tolerance in said offspring to said non-genetic mother-derived antigen or the same antigen as that Use of.
[C2-c]
The other drug induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into the offspring during fetal life, including isolated bone marrow-derived stem cells selected from Use according to [C2], which is a medicament for:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[C2-d]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother gestating a child, wherein the mother's bone marrow-derived stem cell has a non-genetic mother-derived antigen for the child, and
(2) For the child in the fetal period
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(B) by transplantation of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother,
The substance defined in (1) and the isolated bone marrow-derived stem cell defined in (2) in the manufacture of a combined drug for inducing immunological tolerance in said offspring to said non-genetic mother-derived antigen or the same antigen as that Use of.
[C2-e]
The other medicament induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into a neonatal offspring containing isolated bone marrow-derived stem cells selected from The use according to [C2], which is a drug of:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen; and
(C) An isolated bone marrow-derived stem cell derived from an allogeneic donor to the mother and having the same antigen as the non-genetic mother-derived antigen.
[C2-f]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother gestating a child, wherein the mother's bone marrow-derived stem cell has a non-genetic mother-derived antigen for the child, and
(2) For the newborn baby
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen,
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen, and
(C) by transplantation of an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor allogeneic to the mother,
The substance defined in (1) and the isolated bone marrow-derived stem cell defined in (2) in the manufacture of a combined drug for inducing immunological tolerance in said offspring to said non-genetic mother-derived antigen or the same antigen as that Use of.
[C3]
The mother who is gestating the child is a mother who is gestating a child with a disease, and the medicine is a medicine for use in combination with a medicine for treating the disease in the child, [C1]. Use as described in.
[C4]
A mother having a child with the disease is a mother without the disease, and a medicament for treating the disease in the child comprises isolated bone marrow-derived stem cells selected from The use according to [C3], which is a medicament for treating the disease in the offspring by transplantation into the offspring during fetal period:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. An isolated bone marrow-derived stem cell having the same antigen as.
[C4-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who does not have the disease and is pregnant with a child who has the disease, wherein the bone marrow-derived stem cells of the mother are not Having a genetic mother-derived antigen, and
(2) For the child in the fetal period
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(B) an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. By porting
Use of the substance as defined in (1) and the isolated bone marrow-derived stem cells as defined in (2) in the manufacture of a combined medicament for treating the disease in said offspring.
[C5]
The mother having a child with the disease is a mother with the disease, and the medicament for treating the disease in the child is derived from a donor who is syngeneic with the mother and does not have the disease. A drug for treating the disease in the offspring by transplantation into the offspring at the prenatal stage, which comprises an isolated bone marrow-derived stem cell having the same antigen as the non-genetic mother-derived antigen, [C3] Use as stated.
[C5-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother having the disease who is pregnant with a child having the disease, wherein the bone marrow-derived stem cells of the mother are non-genetic mothers for the child Having a derived antigen, and
(2) by transplantation of an isolated bone marrow-derived stem cell derived from a donor who is syngeneic with the mother and has the same antigen as the non-genetic mother-derived antigen to the offspring in the fetal period,
Use of the substance as defined in (1) and the isolated bone marrow-derived stem cells as defined in (2) in the manufacture of a combined medicament for treating the disease in said offspring.
[C6]
An isolated bone marrow-derived stem cell, wherein a mother having a child with the disease is a mother without the disease, and a medicine for treating the disease in the child is selected from the following: Use according to [C3], which is a medicament comprising an isolated tissue or an isolated organ for treating the disease in the offspring by transplantation into the offspring after neonatal period:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(C) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is not allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[C6-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who does not have the disease and is pregnant with a child who has the disease, wherein the bone marrow-derived stem cells of the mother are not Having a genetic mother-derived antigen, and
(2) For the child after the newborn period
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen,
(B) an isolated bone marrow-derived stem cell, isolated tissue, or isolated derived from a donor who is syngeneic with the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(C) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is not allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. Selected bone marrow-derived stem cells, isolated tissue or isolated organ transplant,
In the manufacture of a combination drug for treating the disease in the offspring, the substance defined in (1) and the isolated bone marrow-derived stem cell defined in (2), isolated tissue or isolated Use of organs.
[C7]
An isolated bone marrow-derived stem cell, wherein the mother having a child with the disease is a mother with the disease, and a medicine for treating the disease in the child is selected from the following: Use according to [C3], which is a medicament for treating the disease in the offspring by transplantation into the offspring after the neonatal period, which comprises the isolated tissue or isolated organ:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(B) an isolated bone marrow-derived stem cell, an isolated tissue or an isolated tissue derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[C7-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother having the disease who is pregnant with a child having the disease, wherein the bone marrow-derived stem cells of the mother are non-genetic mothers for the child Having a derived antigen, and
(2) For the child after the newborn period
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(B) an isolated bone marrow-derived stem cell, an isolated tissue or an isolated tissue derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. Organ
Selected from, by transplantation of isolated bone marrow-derived stem cells, isolated tissue or isolated organ,
In the manufacture of a combination drug for treating the disease in the offspring, the substance defined in (1) and the isolated bone marrow-derived stem cell defined in (2), isolated tissue or isolated Use of organs.
[C8]
A medicament for treating the disease in the offspring comprises the same isolated protein as the non-genetic mother-derived antigen for treating the disease in the offspring by administration to the offspring after neonatal period. Use according to [C3], which is a medicine.
[C8-a]
(1) Administration of a substance having an activity of mobilizing bone marrow stem cells into peripheral blood to a mother who is pregnant with a diseased child, wherein the bone marrow-derived stem cell of the mother has a non-genetic mother-derived antigen for the child ,and,
(2) administration of the same isolated protein as the non-genetic mother-derived antigen to the offspring after neonatal period,
Use of a substance as defined in (1) and an isolated protein as defined in (2) in the manufacture of a combined medicament for treating the disease in said offspring.
[C9]
Selected from the following in the manufacture of a medicament for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen for said child by administration to a mother carrying the child: Uses Containing Isolated Bone Marrow-Derived Stem Cells:
(A) an isolated bone marrow-derived stem cell derived from a mother who is gestating the offspring and having a non-genetic mother-derived antigen for the offspring;
(B) An isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen.
[C10]
The use according to [C9], wherein the drug is a drug for use in combination with another drug for inducing immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the non-genetic mother antigen.
[C10-a]
The other drug induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into the offspring during fetal life, including isolated bone marrow-derived stem cells selected from Use according to [C10], which is a medicament for:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[C10-b]
(1) Administration of an isolated bone marrow-derived stem cell selected from the following (a) and (b) to a mother gestating a child, and
(2) By transplanting the isolated bone marrow-derived stem cells selected from the following (a) and (b) to the fetus in the offspring,
To the isolated bone marrow-derived stem cells defined in (1) and (2) in the manufacture of a combined drug for inducing immune tolerance in a child against a non-genetic mother-derived antigen for the child or the same antigen as that Defined use of isolated bone marrow-derived stem cells:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen. Isolated bone marrow-derived stem cells.
[C10-c]
The other medicament induces immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen by transplantation into a neonatal offspring, which contains isolated bone marrow-derived stem cells selected from: The use according to [C10], which is a drug of:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen; and
(C) An isolated bone marrow-derived stem cell derived from an allogeneic donor to the mother and having the same antigen as the non-genetic mother-derived antigen.
[C10-d]
(1) Administration of an isolated bone marrow-derived stem cell selected from the following (a) and (b) to a mother gestating a child, and
(2) By transplanting the isolated bone marrow-derived stem cells selected from the following (a) to (c) to the neonatal period,
To the isolated bone marrow-derived stem cells defined in (1) and (2) in the manufacture of a combined drug for inducing immune tolerance in a child against a non-genetic mother-derived antigen for the child or the same antigen as that Defined use of isolated bone marrow-derived stem cells:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen; and
(C) An isolated bone marrow-derived stem cell derived from an allogeneic donor to the mother and having the same antigen as the non-genetic mother-derived antigen.
[C11]
The mother who is gestating the child is a mother who is gestating a child with the disease, and the medicine is a medicine for use in combination with a medicine for treating the disease in the child, [C9]. Use as described in.
[C12]
A mother having a child with the disease is a mother without the disease, and a medicament for treating the disease in the child comprises isolated bone marrow-derived stem cells selected from The use according to [C11], which is a medicament for treating the disease in the offspring by transplantation into the offspring during fetal period:
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. An isolated bone marrow-derived stem cell having the same antigen as.
[C12-a]
(1) For mothers without the disease who are pregnant with a child with the disease
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) For the child in the fetal period
(I) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, and
(Ii) an isolated bone marrow-derived stem cell selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor who does not have the disease and is syngeneic with the mother By porting
Use of the isolated bone marrow-derived stem cells as defined in (1) and the isolated bone marrow-derived stem cells as defined in (2) in the manufacture of a combination drug for treating the disease in the offspring.
[C13]
The mother having a child with the disease is a mother with the disease, and the medicament for treating the disease in the child is derived from a donor who is syngeneic with the mother and does not have the disease. A drug for treating the disease in the offspring by transplantation to the offspring in the fetal period, comprising isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, [C11] Use as stated.
[C13-a]
(1) For mothers with a disease who are pregnant with a child with the disease
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) by transplantation of an isolated bone marrow-derived stem cell derived from a donor who is syngeneic with the mother and has the same antigen as the non-genetic mother-derived antigen to the offspring in the fetal period,
Use of the isolated bone marrow-derived stem cells as defined in (1) and the isolated bone marrow-derived stem cells as defined in (2) in the manufacture of a combination drug for treating the disease in the offspring.
[C14]
A mother having a child with the disease is a mother without the disease, and a drug for treating the disease in the child has isolated bone marrow-derived stem cells selected from the following: Use according to [C11], which is a medicament for treating the disease in the offspring by transplantation into the offspring after neonatal period, including isolated tissue or isolated organs:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(C) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is not allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[C14-a]
(1) For mothers without the disease who are pregnant with a child with the disease
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) For the child after the newborn period
(I) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen, an isolated tissue or an isolated organ,
(Ii) isolated bone marrow-derived stem cells, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(Iii) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is allogeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen. By transplantation of isolated bone marrow-derived stem cells, isolated tissues or isolated organs selected from
Isolated bone marrow-derived stem cells as defined in (1) and isolated bone marrow-derived stem cells as defined in (2), in the manufacture of a combination drug for treating the disease in said offspring, isolated Use of tissues or isolated organs.
[C15]
A mother having a child with the disease is a mother with the disease, and a medicament for treating the disease in the child is an isolated bone marrow-derived stem cell selected from the following: The use according to [C11], which is a medicament for treating the disease in the offspring by transplanting into the offspring after the neonatal period, including the isolated tissue or the isolated organ:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs; and
(B) an isolated bone marrow-derived stem cell, an isolated tissue or an isolated tissue derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. An organ.
[C15-a]
(1) For mothers with a disease who are pregnant with a child with the disease
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) For the child after the newborn period
(I) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen Organs, and
(Ii) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. By transplantation of isolated bone marrow-derived stem cells, isolated tissues or isolated organs selected from
Isolated bone marrow-derived stem cells as defined in (1) and isolated bone marrow-derived stem cells as defined in (2), in the manufacture of a combination drug for treating the disease in said offspring, isolated Use of tissues or isolated organs.
[C16]
A medicament for treating the disease in the offspring comprises the same isolated protein as the non-genetic mother-derived antigen for treating the disease in the offspring by administration to the offspring after neonatal period. Use according to [C11], which is a medicine.
[C16-a]
(1) For mothers who are pregnant with sick children
(A) an isolated bone marrow-derived stem cell derived from the mother and having a non-genetic mother-derived antigen for the offspring, and
(B) administration of isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor syngeneic to the mother, and
(2) administration of the same isolated protein as the non-genetic mother-derived antigen to the offspring after neonatal period,
Use of the isolated bone marrow-derived stem cells as defined in (1) and the isolated protein as defined in (2) in the manufacture of a combined medicament for treating the disease in the offspring.
[C17]
[C3] to [C8] and [C11] to [C16], which are treatable as a result of induction of immunological tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as that of the non-genetic mother. Use as described in either.
[C18]
[C3] to [C8] and [C11] to [C16], wherein the disease is treatable as a result of suppression of immune reaction in the offspring to the non-genetic mother-derived antigen or the same antigen as that. Use as described in either.
[C19]
The use according to any of [C1] to [C18], wherein the stem cells are mesenchymal stem cells or hematopoietic stem cells.
[C20]
The use according to any of [C1] to [C18], wherein the stem cells are mesenchymal stem cells.
[C21]
The administration of mesenchymal stem cells to a mother who is gestating the offspring produces mesenchymal stem cells in the manufacture of a medicament for use in inducing immune tolerance in the offspring to a non-genetic mother-derived antigen for the offspring or the same antigen as the offspring. Use of a substance having an activity of mobilizing in peripheral blood, wherein the bone marrow-derived stem cells of the mother carry the non-genetic mother-derived antigen.
[C22]
Selected from the following in the manufacture of a medicament for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen for said child by administration to a mother carrying the child: Use of isolated mesenchymal stem cells:
(A) an isolated mesenchymal stem cell derived from a mother who is gestating the offspring and having a non-genetic mother-derived antigen for the offspring;
(B) An isolated mesenchymal stem cell derived from a donor syngeneic to the mother and having the same antigen as the non-genetic mother-derived antigen.

3 is a graph showing the results of measuring the amount of anti-GFP antibody produced in mice transplanted with cells in Example 1 by ELISA. 2 is a graph showing the results of evaluation by ELISPOT of the immune response of mice transplanted with cells in Example 1. 5 is a graph showing the results of measuring the amount of anti-GFP antibody produced in mice transplanted with cells in Example 2 by ELISA. 5 is a graph showing the results of evaluating the immune response of mice transplanted with cells in Example 2 by ELISPOT. FIG. 5 is a diagram showing the results of skin graft experiments using PDGFRaKI mice in Example 3. FIG. 8 is a diagram showing the results of skin graft experiments on HMGB1-administered mouse pups in Example 4. It is the figure which plotted the number of the colonies obtained by culturing the peripheral blood 14 hours after the administration of physiological saline or peptide. The number of colonies was shown as a value converted per 1 mL of collected peripheral blood. The long bar indicates the average value, and the short bar indicates the standard deviation. It is the figure which plotted the number of the colonies obtained by culturing the peripheral blood 14 hours after the administration of physiological saline or peptide. The number of colonies was shown as a value converted per 1 mL of collected peripheral blood. The long bar indicates the average value, and the short bar indicates the standard deviation. It is the figure which plotted the number of the colonies obtained by culturing the peripheral blood 16 hours after the administration of physiological saline or peptide. The number of colonies was shown as a value per peripheral blood volume (about 800 μL) collected from one mouse. The long bar indicates the average value, and the short bar indicates the standard deviation. It is the figure which plotted the number of the colonies obtained by culturing the peripheral blood 24 hours after the administration of physiological saline or peptide. The number of colonies was shown as a value per peripheral blood volume (about 800 μL) collected from one mouse. The long bar indicates the average value, and the short bar indicates the standard deviation. It is the figure which plotted the number of the colonies obtained by culturing the peripheral blood 14 hours after the administration of physiological saline or peptide. In the graph, "saline" represents the control group and "1r10" represents the peptide 1r10 administration group. The number of colonies was shown as a value per peripheral blood volume (about 800 μL) collected from one mouse. The long bar indicates the average value, and the short bar indicates the standard deviation.

The present application relates to a medicinal agent for use in inducing tolerance to a non-genetic mother-derived antigen for a child or the same antigen as that of the child by administration to a mother who is gestating the child, which comprises bone marrow stem cells. The present invention provides a medicine (hereinafter, referred to as “first medicine”), which comprises a substance having the activity of mobilizing saccharin into peripheral blood, and the bone marrow-derived stem cells of the mother have the non-genetic mother-derived antigen.

Further, the present application is a medicine for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen for the child or the same antigen as that of the child by administration to a mother who is gestating the child,
(A) an isolated bone marrow-derived stem cell derived from a mother gestating a child and having a non-genetic mother-derived antigen for the child; and
(B) A medicament containing isolated bone marrow-derived stem cells selected from isolated bone marrow-derived stem cells that are derived from a donor syngeneic to the mother and have the same antigen as the non-genetic mother-derived antigen (hereinafter referred to as “the 2 pharmaceuticals”).

As the mechanism of action of the first or second drug, the bone marrow-derived stem cells of the mother that have been mobilized to peripheral blood by the substance that has been administered to the mother, or the isolated bone marrow-derived stem cells that have been administered to the mother are , Transplacentally transferred from mother to fetal (eg embryonic or fetal) offspring, exposed to non-genetic mother-derived antigens derived from bone marrow-derived stem cells or their lineage cells (lineage cells), A mechanism that induces immunological tolerance in the offspring to the mother-derived antigen or the same antigen as that of the genetic mother is considered, but is not limited thereto.

The first or second medicine is to surgically access the fetus in the mother's uterus, inject stem cells into the umbilical vein of the fetus, or remove the fetus from the mother's body for treatment and then return it to the mother's body. It requires extremely advanced and invasive procedures, a heavy burden on the mother's body, is dangerous for both mother and child, and there is no problem such as fetal cell transplantation.

In the present application, a sufficient amount (ie, an effective amount) of the substance or the isolated bone marrow-derived stem cells to induce immune tolerance in the offspring is administered to the mother. The substance or the isolated bone marrow-derived stem cells or the medicament containing the same can be administered to a mother by intravascular administration (intraarterial administration, intravenous administration, etc.), intramuscular administration, subcutaneous administration, intraperitoneal administration, etc. But is not limited to these. Further, the substance or the isolated bone marrow-derived stem cells or a drug containing the same can be systemically administered by injection, for example, intravenous injection, intramuscular injection, intraperitoneal injection, subcutaneous injection and the like.

When the drug to be administered to the mother is the first drug, in addition to the above-mentioned administration methods, oral administration, nasal administration, pulmonary administration, transdermal administration, etc. may be mentioned. For example, skin surface, eyeball or palpebral conjunctiva, nasal mucosa, oral and digestive tract mucosa, vaginal/intrauterine mucosa, or damaged site) can be selected.

Before administration of the substance or the isolated bone marrow-derived stem cells to the mother, the mother and offspring cells can be examined to confirm the presence or absence of the non-genetic mother-derived antigen and to identify the non-genetic mother-derived antigen. .. The offspring cell may be a fertilized egg. As a method for testing offspring, a general prenatal diagnosis method can be applied.A villus test for collecting placental villous tissue, a fetal blood test for collecting fetal blood, and a fetal origin mixed in mother peripheral blood Examples include cell tests. By comparing the genetic analysis of the offspring based on such a test with the genetic analysis result of the mother, the non-genetic mother-derived antigen for the offspring can be identified. In the present application, it is not essential to identify the non-genetic mother-derived antigen.

The non-genetic mother-derived antigen may be present in the mother's bone marrow-derived stem cells or the isolated bone marrow-derived stem cells, and there is no limitation on the timing of its expression. Also, preferably, the non-genetic mother-derived antigen is expressed on the maternal bone marrow-derived stem cells or the isolated bone marrow-derived stem cells.

In the present application, the non-genetic mother-derived antigen means a gene product (protein antigen) of a gene that was only inherited from mother to child and an antigen (peptide, sugar chain, lipid antigen, etc.) secondarily produced by the product. Any biological substance that can become). Such non-genetic mother-derived antigens include non-genetic mother-derived MHC antigens (human non-genetic mother-derived HLA antigens), non-genetic mother-derived non-major histocompatibility antigens, and gene products expressed in mothers. Examples include gene products that cannot be produced in offspring due to mutations or gene defects, or that can be produced but have altered structures.

Non-genetic mother-derived antigen in the present application, for example, non-genetic mother-derived type VII collagen antigen when the COL7A1 gene mutation is deficient in the child, type I collagen deficiency in the child due to COL1A1 or COL1A2 gene mutation Examples include, but are not limited to, a non-genetic mother-derived type I collagen antigen, a non-genetic mother-derived dystrophin protein antigen when a dystrophin protein is deficient in a child due to a mutation of the DMD gene, and the like.

The non-genetic mother-derived antigen in which immunological tolerance is induced by the drug of the present application may be one non-inherited maternal antigen (NIMA), or multiple non-genetic mother-derived antigens (non -inherited maternal antigens: NIMAs). Here, the term "a certain one (a)" may mean "at least one (a) least". From this point of view, the non-genetic mother-derived antigen in the present drug should be construed as the broadest meaning.

In the present application, the same antigen as the non-genetic mother-derived antigen means an antigen recognized in vivo as the same as the non-genetic mother-derived antigen.

The same antigen as the non-genetic mother-derived antigen includes, but is not limited to, any in-vivo substance that can be an antigen, such as protein antigen, peptide, sugar chain, and lipid antigen.

Induction of immune tolerance in children involves not only the condition in which immune tolerance has not occurred but the condition in which immune tolerance has occurred, but also the case in which the condition in which immune tolerance has occurred is maintained or has already occurred. It also includes the case of promoting.

In the offspring, whether or not immune tolerance to the non-genetic mother-derived antigen or the same antigen as that was induced was determined by the inflammatory reaction such as redness and swelling of the transplanted area, fever, blood cytokine increase, and CRP It can be evaluated by systemic inflammatory reaction such as elevation, graft-versus-host reaction and the like. In addition, it can be evaluated by detecting an antibody against an antigen in the offspring.

∙ The gestation in this application also includes surrogate gestation. In the present application, the bone marrow-derived stem cells of the surrogate mother carry the antigen for the offspring (eg, the same antigen as the non-genetic mother-derived antigen).

In the present application, the isolated bone marrow-derived stem cells derived from the mother are not limited to those collected after the mother has conceived the child, and may be those collected before the mother conceived the child.

In the present application, the isolated bone marrow-derived stem cells may be cells that have been artificially modified so as to have a non-genetic mother-derived antigen or the same antigen. As a result, the isolated bone marrow-derived stem cells, which originally do not have the non-genetic mother-derived antigen or the same antigen as the antigen, will have the antigen.

As such a modification method, a known method of introducing a gene encoding a non-genetic mother-derived antigen or the same antigen as it into a cell can be appropriately selected and used, and for example, a CRISPR system (for example, CRISPR/Cas9 , CRISPR/Cpf1), TALEN, ZFN, etc., and methods using viral vectors such as retrovirus vector, lentivirus vector, adenovirus vector, adeno-associated virus vector, etc. can be used, but not limited to these. .. It is desirable that the gene be introduced into the cell genome and be permanently expressed, but it is also possible to use a Sendai virus vector or episomal vector that is not incorporated into the genome and remains in the cytoplasm to express the gene in the medium to long term. Good. Electroporation may be used to introduce the episomal vector into the cells.

The isolated bone marrow-derived stem cell having the non-genetic mother-derived antigen or the same antigen as the non-genetic mother-derived antigen may be an isolated bone marrow-derived stem cell expressing the non-genetic mother-derived antigen or the same antigen as the non-genetic mother-derived antigen.

In the present application, “bone marrow stem cells” are stem cells existing in the bone marrow. On the other hand, “bone marrow-derived stem cells” are bone marrow stem cells mobilized from the bone marrow to the outside of the bone marrow.

With respect to isolated bone marrow-derived stem cells in the present application, for example, the term "isolated bone marrow-derived stem cells derived from a mother and having a non-genetic mother-derived antigen for the offspring" is "isolated from the mother and , Bone marrow-derived stem cells having a non-genetic mother-derived antigen for the offspring”.

The mother and offspring in this application are not particularly limited as long as they are fetal animals. Examples of the embryonic animal include human or non-human animals, and examples thereof include humans, mice, rats, monkeys, pigs, dogs, rabbits, hamsters, guinea pigs, horses, sheep, whales, and other mammals. It is not limited to.

The term “medicament” in this application is used interchangeably with “drug”, “pharmaceutical composition” or “pharmaceutical composition”.

In the present application, the first or second drug can also be used in combination with another drug for inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen. By using a plurality of drugs in combination, immune tolerance can be induced more effectively than in the case of not using the drugs in combination.

In the present application, the term “for combined use” is used interchangeably with “for combined use”, “for combined therapy” or “for use in combined therapy”. These terms relate to the use of multiple medicaments simultaneously or sequentially in any order.

In the present application, the “other drug” used in combination with the first drug may be the second drug, and the “other drug” used in combination with the second drug may be the first drug. When the first drug and the second drug are used in combination, the mother-, child- and non-genetic mother-derived antigens are the same between the first drug and the second drug.

In the present application, examples of the “other drug” used in combination with the first or second drug include a drug selected from the following (1) and (2) (hereinafter referred to as “third drug”). Examples include, but are not limited to:
(1) Induction of immunological tolerance in the offspring to a non-genetic mother-derived antigen for the offspring or the same antigen as that of the offspring by transplantation into an offspring in the prenatal period containing the isolated bone marrow-derived stem cells selected from the following Medicine for:
(A) an isolated bone marrow-derived stem cell that is derived from a mother who is gestating the offspring and has a non-genetic mother-derived antigen for the offspring; and (b) is derived from a donor syngeneic with the mother An isolated bone marrow-derived stem cell having the same antigen as the mother-derived antigen.
(2) Induction of immunological tolerance in the offspring to a non-genetic mother-derived antigen for the offspring or the same antigen as that of the offspring by transplantation into a neonatal offspring containing isolated bone marrow-derived stem cells selected from the following Medicine for:
(A) an isolated bone marrow-derived stem cell derived from a mother who is gestating the offspring and having a non-genetic mother-derived antigen for the offspring;
(B) an isolated bone marrow-derived stem cell derived from an allogeneic donor of the mother and having the same antigen as the non-genetic mother-derived antigen; and (c) derived from an allogeneic donor of the mother, An isolated bone marrow-derived stem cell having the same antigen as the genetic mother-derived antigen.

When the first or second medicine and the third medicine are used in combination, the mother-, child-, and non-genetic mother-derived antigens in the third medicine are the same as those in the first or second medicine.

For example, the application provides a medicament selected from:
A first medication for use in combination with a second medication;
A first medication for use in combination with a third medication;
A second drug for use in combination with the first drug;
A second drug for use in combination with a third drug;
A combination drug containing a first drug and a second drug;
A combination drug containing a first drug and a third drug; and a combination drug containing a second drug and a third drug.

In the present application, a sufficient amount (ie, an effective amount) of the above-mentioned isolated bone marrow-derived stem cells (third drug) for inducing immune tolerance in the offspring is transplanted into a prenatal or neonatal offspring. Examples of the method for transplanting the isolated bone marrow-derived stem cells or a drug containing the same to fetal offspring include injection of bone marrow-derived stem cells into umbilical vein and intraamniotic administration, and as a method for transplantation into neonatal offspring. In addition to intravascular administration (intraarterial administration, intravenous administration, etc.), intramuscular administration, subcutaneous administration, intraperitoneal administration, direct injection into injured tissue or diseased tissue can be mentioned.

In the present application, as an allogeneic donor with a mother, a relative or unrelated person whose MHC haplotype matches a recipient child, or a sibling whose MHC haplotype partially matches a recipient child, Examples include siblings in which the incompatible MHC is the same substance as the non-genetic mother-derived MHC antigen for the offspring, but are not limited thereto.

In the present application, when the child has a disease, the first or second medicine can also be used in combination with a medicine for treating the disease in the child (hereinafter referred to as “fourth medicine”). By using these medicines in combination, the disease can be treated more effectively as compared with the case where they are not used in combination. The first or second drug can also be used to improve the therapeutic effect of the fourth drug. Generally, in treatment involving transplantation, a donor with high MHC compatibility is selected, but finding a suitable donor is not easy. Here, for example, when the recipient is induced tolerant to a non-hereditary mother-derived MHC antigen by the first or second drug, and the mismatched MHC of the donor is a non-hereditary mother-derived MHC antigen, a mismatch occurs. Even after transplantation, the immune response in the recipient can be suppressed. In other words, the use of the first or second drug can expand the tolerance range of the donor in the recipient in which immune tolerance is induced.

In the present application, in the case where the mother is a mother without the disease, who is pregnant with a child with the disease, the fourth medicine includes a medicine selected from the following (1) to (3). , But not limited to them.
(1) A medicament for treating a disease in an offspring of a prenatal period, which comprises an isolated bone marrow-derived stem cell selected from the following:
(A) isolated bone marrow-derived stem cells derived from a non-diseased mother gestating a diseased child and carrying a non-genetic mother-derived antigen for the offspring; and (b) allogeneic with the mother. An isolated bone marrow-derived stem cell derived from a syngeneic, disease-free donor and having the same antigen as the non-genetic mother-derived antigen.
(2) For treating a disease in an offspring of a neonatal period or later, which comprises an isolated bone marrow-derived stem cell, an isolated tissue or an isolated organ selected from the following: Medicine:
(A) Isolated bone marrow-derived stem cells, isolated tissue or single cells derived from a non-diseased mother who is pregnant with a diseased child and has a non-genetic mother-derived antigen for the child. Separated organs;
(B) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen An organ; and (c) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is allogeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen. Isolated organ.
(3) A medicament for treating a disease in a child after the neonatal period, which comprises an isolated protein which is the same as the non-genetic mother-derived antigen for the child and is administered to the child after the neonatal period.

In the present application, in the case where the mother is a mother having the disease who is pregnant with a child having the disease, the fourth drug includes a drug selected from the following (1) to (3). Not limited to.
(1) Isolated from a donor who does not have the disease and is syngeneic with a mother who has the disease and is pregnant with a child who has the disease, and has the same antigen as the non-genetic mother-derived antigen for the child A medicament for treating a disease in an offspring of a prenatal period, which comprises a bone marrow-derived stem cell.
(2) For treating a disease in an offspring of a neonatal period or later, which comprises an isolated bone marrow-derived stem cell, an isolated tissue or an isolated organ selected from the following: Medicine:
(A) Isolation, which is derived from a donor who does not have the disease, which is syngeneic with the mother who has the disease, who is pregnant with the child who has the disease, and which has the same antigen as the non-genetic mother-derived antigen for the child. Bone marrow-derived stem cells, an isolated tissue or an isolated organ; and (b) an antigen identical to the non-genetic mother-derived antigen, which is derived from a donor of the disease allogeneic with the mother. Having isolated bone marrow-derived stem cells, isolated tissue or isolated organs.
(3) A medicament for treating a disease in a child after the neonatal period, which comprises an isolated protein which is the same as the non-genetic mother-derived antigen for the child and is administered to the child after the neonatal period.

When the first or second medicine and the fourth medicine are used in combination, the mother-, child-, and non-genetic mother-derived antigens in the fourth medicine are the same as those in the first or second medicine.

For example, the application provides a medicament selected from:
A first medication for use in combination with a fourth medication;
A second medication for use in combination with a fourth medication;
A combination drug containing a first drug and a fourth drug; and a combination drug containing a second drug and a fourth drug.

As the disease in the offspring, a disease that can be treated as a result of induction of immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as that, and in the offspring to the non-genetic mother-derived antigen or the same antigen as that Examples include, but are not limited to, diseases that are treatable as a result of suppressed immune responses. Such a disease may be a hereditary disease or a non-hereditary disease.

For example, a hereditary disease in a child is a disease caused by a mutant gene inherited from a mother and/or a father to a child, and the mutant gene has the same locus as the gene encoding the non-genetic mother-derived antigen. Disease, which is a mutant gene derived from the mutant gene present in. Further, for example, the inherited disease in the offspring is a disease caused by a mutation in the offspring of a wild type (normal type) gene inherited from the mother and/or the father to the offspring, and the wild type (normal type) gene Is a wild type (normal type) gene derived from a wild type (normal type) gene existing at the same locus as the gene encoding the non-genetic mother-derived antigen.

Further, for example, the non-hereditary disease in the offspring is, if immune tolerance in the offspring to the non-inherited mother-derived MHC (HLA) antigen or the same antigen as that is induced, or if the immune response in the offspring is suppressed, It is a disease that can be treated.

The disease in the offspring may be, for example, a disease in which normal production of a specific protein is impossible, a disease in which cell dysfunction occurs, a disease caused by a mutant gene mutated in a child, or a mutant gene. Diseases in which no protein is produced or abnormal proteins are produced, and diseases with a high risk of onset in the future are predicted by genetic testing, and effective treatment for the disease (non-hereditary disease) is cell transplantation or organ transplantation. There are certain diseases.

Diseases in children include epidermolysis bullosa, osteogenesis imperfecta, fibrinogen deficiency, congenital leukemia, adenosine deaminase (ADA) deficiency, and Duchenne muscular dystrophy.

In the present application, when the first or second medicine and the fourth medicine are used in combination, a test for pre-implantation diagnosis or prenatal diagnosis can be performed for a disease.

In the present application, when the first or second medicine and then the fourth medicine are successively used in this order, a child is used after using the first or second medicine and before using the fourth medicine. It is possible to confirm whether or not microchimerism is positive in. Here, the microchimerism means cell transfer from the mother to the fetus and engraftment. As a method for confirming microchimerism, for example, when the patient is a boy (XY chromosome), a method of confirming mother-derived cells by searching for XX chromosome cells in the body can be mentioned. In addition, as an indirect confirmation method, inflammatory reaction such as redness and swelling of the transplanted area that occurs after the antigen transplantation, fever, systemic inflammatory reaction such as elevation of blood cytokine, elevation of CRP, graft-versus-host reaction, etc. Can be evaluated. In addition, it can be evaluated by detecting an antibody against an antigen in the offspring.

In this application, a sufficient amount (ie, an effective amount) of isolated bone marrow-derived stem cells, isolated tissue or isolated organs to treat a disease in a child is transferred to a child of prenatal or neonatal age. Ported. As a method for transplanting the isolated bone marrow-derived stem cells, the isolated tissue or the isolated organ, or a drug containing the same into a fetus during offspring, in the case of the isolated bone marrow-derived stem cells, a umbilical vein is used. Examples include injection of bone marrow-derived stem cells and intra-amniotic fluid administration, and transplantation by surgical operation in the case of isolated tissues or isolated organs. As a method for transplantation into a child after the neonatal period, in the case of isolated bone marrow-derived stem cells, in addition to intravascular administration (intraarterial administration, intravenous administration, etc.), intramuscular administration, subcutaneous administration, intraperitoneal administration, Examples include direct injection into injured tissue or diseased tissue, and in the case of isolated tissue or isolated organ, transplantation by surgery.

In the present application, a sufficient amount (ie, an effective amount) of the isolated protein to treat the disease in the offspring is administered to the offspring after neonatal period. As a method of administering the isolated protein to a child after the neonatal period, there are oral administration, intranasal administration in addition to intravascular administration (intraarterial administration, intravenous administration, etc.), intramuscular administration, subcutaneous administration, and intraperitoneal administration. Administration, pulmonary administration, transdermal administration, and direct injection into damaged or diseased tissue.

The treatments in this application include, but are not limited to, alleviation, delay, prevention, improvement, remission, cure, cure, etc.

The stem cells in the present application are mesenchymal stem cells or hematopoietic stem cells.

The present application relates to a medicinal product for use in inducing immunological tolerance in a child against a non-genetic mother-derived antigen for the child or the same antigen as that of the mesenchyme by administration to a mother who is gestating the child. There is also provided a medicament, which comprises a substance having an activity of mobilizing lineage stem cells into peripheral blood, wherein the maternal bone marrow-derived stem cells have the non-genetic mother-derived antigen. The drug can be used as an alternative drug to the first drug.

The present application provides a medicine for use in inducing immunological tolerance in a child to a non-genetic mother-derived antigen for the child or the same antigen as that of the child by administration to a mother who is gestating the child,
(A) an isolated mesenchymal stem cell having a non-genetic mother-derived antigen for the offspring, which is derived from a mother having the offspring, and
(B) A drug containing an isolated mesenchymal stem cell selected from isolated mesenchymal stem cells having the same antigen as the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother. provide. The drug can be used as an alternative drug to the second drug.

In the present application, the “mesenchymal stem cells” are collected from bone marrow or other tissues (blood, such as cord blood, and skin, fat, dental pulp, etc.), and in vivo, between bone, cartilage, fat, muscle, etc. It is a cell having the ability to differentiate into leaf-like cells and the ability to self-renew. In addition, it can be cultured and expanded in vitro as adherent cells to a culture dish (plastic or glass), and has the ability to differentiate into mesenchymal tissues such as bone, cartilage, fat and muscle and self-renewal ability. Cells that have. In one embodiment, the mesenchymal stem cells also have the ability to differentiate into epithelial tissues and neural tissues. In one embodiment, the mesenchymal stem cells may be present as a heterogeneous cell population containing not only narrowly defined stem cells but also progenitor cells, and differentiated in addition to narrowly defined stem cells and/or progenitor cells under culture conditions. The cells may also be included. In one embodiment, the mesenchymal stem cells may be composed of only stem cells in a narrow sense, or may be a cell population composed of a plurality of types of progenitor cells.

In the present application, progenitor cells are defined as cells having unidirectional differentiation ability to specific tissue cells other than blood cells, and have the ability to differentiate into mesenchymal tissue, epithelial tissue, nerve tissue, parenchymal organ, and vascular endothelium. Including cells having.

In the present application, examples of mesenchymal stem cells include, but are not limited to, bone marrow mesenchymal stem cells and bone marrow-derived mesenchymal stem cells. "Bone marrow mesenchymal stem cells" are cells that are present in the bone marrow and are mesenchymal stem cells collected from the bone marrow. In one embodiment, the bone marrow mesenchymal stem cells also have the ability to differentiate into epithelial tissues and neural tissues. In the present application, the term "bone marrow mesenchymal stem cells" is used interchangeably with "bone marrow mesenchymal stromal cells", "bone marrow pluripotent stem cells" or "bone marrow pluripotent stromal cells".

In addition, "bone marrow-derived mesenchymal stem cells" refer to bone marrow mesenchymal stem cells mobilized from the bone marrow to the outside of the bone marrow, peripheral blood sampling, and further mesenchymal tissues such as fat, epithelial tissues such as skin, and brain. It is a cell that can be obtained by collecting from nerve tissue such as. In the present application, the term "bone marrow-derived mesenchymal stem cells" is used interchangeably with "bone marrow-derived mesenchymal stromal cells", "bone marrow-derived pluripotent stem cells" or "bone marrow-derived pluripotent stromal cells" To be

In one embodiment, the bone marrow mesenchymal stem cells and the bone marrow-derived mesenchymal stem cells are directly collected or by administering the cells once attached to a culture dish to a damaged part of a living body, for example, keratinocytes constituting the skin. It also has the characteristic of having the ability to differentiate into epithelial tissues of the brain and tissues of the nervous system that constitutes the brain.

Bone marrow mesenchymal stem cells and bone marrow-derived mesenchymal stem cells include osteoblasts (identifiable by observing calcium deposition when differentiation is induced), chondrocytes (alcian blue staining positive, safranin-O staining positive, etc.). Identifiable), adipocytes (identifiable by Sudan III staining positive, etc.), as well as mesenchymal cells such as fibroblasts, smooth muscle cells, skeletal muscle cells, stromal cells, tendon cells, nerve cells, pigment cells , Epidermal cells, hair follicle cells (express cytokeratin family, hair keratin family, etc.), epithelial cells (eg epidermal keratinocytes, intestinal epithelial cells express cytokeratin family, etc.), endothelial cells, and liver, It is preferable that the cells have the ability to differentiate into parenchymal organ cells such as kidney and pancreas, but the differentiated cells are not limited to the above cells.

As markers for human mesenchymal stem cells, PDGFRα positive, PDGFRβ positive, Lin negative, CD45 negative, CD44 positive, CD90 positive, CD29 positive, Flk-1 negative, CD105 positive, CD73 positive, CD90 positive, CD71 positive, Stro- Examples of all or part of 1-positive, CD106-positive, CD166-positive, CD31-negative, CD271-positive, and CD11b-negative can be mentioned, but the present invention is not limited thereto.

As markers for mouse mesenchymal stem cells, CD44 positive, PDGFRα positive, PDGFRβ positive, CD45 negative, Lin negative, Sca-1 positive, c-kit negative, CD90 positive, CD105 positive, CD29 positive, Flk-1 negative, CD271 All or a part of positive and CD11b negative can be exemplified, but the present invention is not limited thereto.

Examples of rat mesenchymal stem cell markers include PDGFRα-positive, CD44-positive, CD54-positive, CD73-positive, CD90-positive, CD105-positive, CD29-positive, CD271-positive, CD31-negative, and all or part of CD45-negative. Is not limited to.

In the present application, the mesenchymal stem cells, PDGFRα-positive mesenchymal stem cells, PDGFRα-positive bone marrow-derived mesenchymal stem cells, PDGFRα-positive bone marrow-derived cells, by bone marrow collection (bone marrow cell collection) or peripheral blood sampling Examples of the cells obtained as adherent cells by the mononuclear cell fractionated cell culture in the obtained blood can be exemplified, but the present invention is not limited thereto. Examples of PDGFRα-positive mesenchymal stem cells include PDGFRα and CD44 positive cells, PDGFRα and CD90 positive cells, PDGFRα and CD105 positive cells, PDGFRα and CD29 positive cells, etc. To be In one embodiment, PDGFRα positive mesenchymal stem cells may be CD44 negative cells. In one embodiment, the mesenchymal stem cell marker is PDGFRα positive and Lin negative.

The hematopoietic stem cell in the present application refers to a stem cell capable of differentiating into a hematopoietic cell. Somatic stem cells that can differentiate into white blood cells (neutrophils, eosinophils, basophils, lymphocytes, monocytes, macrophages), red blood cells, platelets, mast cells, and dendritic cells. Stem cells that can supply blood cells, for example, cells that can reproduce the hematopoietic mechanism when transplanted into immunodeficient mice. This hematopoietic stem cell can be identified by a unique marker, and markers such as CD34, c-kit, and sca-1 are known, but the marker is not limited to these.

The substance having an activity of mobilizing bone marrow stem cells into peripheral blood in the present application is a substance having an activity of mobilizing bone marrow mesenchymal stem cells or bone marrow hematopoietic stem cells into peripheral blood. The term “activity of mobilizing bone marrow stem cells into peripheral blood” is used interchangeably with “activity of increasing the abundance of bone marrow-derived stem cells in peripheral blood”.

In the present application, the activity of mobilizing bone marrow mesenchymal stem cells into peripheral blood refers to i) an individual administered with a substance having an activity of mobilizing bone marrow stem cells into peripheral blood, and peripherals from an individual not administered the substance. Blood is collected, seeded in culture dish and cultured (several days to 10 days), and the number of colonies formed is counted, and ii) the formed colonies adhere to the solid phase and grow. It can be evaluated by confirming that it has the ability (self-renewal ability) and the ability to differentiate into osteoblasts, chondrocytes and adipocytes. In i) above, erythrocytes may be removed from the peripheral blood by a desired method before seeding the collected peripheral blood on the culture dish.

As a substance having an activity of mobilizing bone marrow mesenchymal stem cells in peripheral blood in the present application, a nuclear protein or a fragment peptide thereof, a specific artificial peptide, a CXCR4 chemokine receptor inhibitor (for example, AMD-3100, ALX40-4C, T134 Etc.) (Special table 2012-525393).

In the present application, “nuclear protein” means a protein that exhibits a certain function in the nucleus.

The nuclear proteins in the present application include, but are not limited to, nuclear proteins involved in transcriptional regulation, for example. The “protein involved in transcriptional regulation” means a protein having a function of regulating any process of transcription among nuclear proteins, and examples thereof include, but are not limited to, a transcription factor and a transcription coupling factor. ..

In the present application, a transcription factor is a protein that binds to DNA alone or in the form of a complex with another protein to control transcription, and includes a basic transcription factor (protein constituting a transcriptional device), a transcriptional regulatory factor, It includes transcription elongation factors, factors that regulate transcription by participating in the transcription termination process, and the like.

In the present application, the transcription coupling factor is a protein that does not directly bind to DNA but regulates transcription by the interaction between proteins, and is, for example, interposed (bound to both) between a transcriptional regulatory factor and a basal transcription factor. Examples thereof include, but are not limited to, a coactivator and a corepressor that regulate transcription.

In the present application, “a fragment peptide of a nuclear protein” means a fragment peptide derived from the above nuclear protein. In one embodiment, the fragment peptide of the nuclear protein is a fragment peptide having an activity of mobilizing bone marrow mesenchymal stem cells into peripheral blood.

In the present application, the term "fragment peptide of nucleoprotein" means "fragment peptide derived from nucleoprotein", "partial peptide derived from nucleoprotein", "fragment peptide consisting of a part of nucleoprotein", "of nucleoprotein". It is used interchangeably with "partial peptide consisting of a part" or "partial peptide of nuclear protein".

The nuclear protein or a fragment peptide thereof in the present application, or a specific artificial sequence peptide can be obtained as a recombinant by incorporating the DNA encoding the same into an appropriate expression system, or artificially. It can also be synthesized. Thus, the nuclear protein or peptide also includes a nuclear protein or peptide produced by using a cell, and an artificially synthesized nuclear protein or peptide (that is, a synthetic nuclear protein or peptide). ..

In order to obtain the nuclear protein or a fragment peptide thereof in the present application or a specific artificial sequence peptide by a genetic engineering technique, a DNA encoding the nuclear protein or peptide may be incorporated into an appropriate expression system and expressed. ..

Hosts applicable to the present application include, but are not limited to, prokaryotic cells and eukaryotic cells. In addition, examples of hosts applicable to the present application also include bacteria (for example, Escherichia coli), yeast, animal cells (for example, mammalian cells such as HEK293 cells and CHO cells, insect cells such as silkworm cells), and plant cells. , Not limited to these.

The host/vector system applicable to the present application can include, for example, the expression vector pGEX and E. coli. pGEX can express a foreign gene as a fusion protein with glutathione S-transferase (GST) (Gene, 67:31-40, 1988), so the nuclear protein of the present application or its fragment peptide, or a specific artificial sequence Introducing pGEX incorporating the DNA encoding the peptide into an E. coli strain such as BL21 by heat shock, and adding isopropylthio-β-D-galactoside (IPTG) after an appropriate culture time to induce the expression of the GST fusion peptide To do. Since GST in the present application is adsorbed on glutathione sepharose 4B, the expression product can be easily separated and purified by affinity chromatography.

As the host/vector system for obtaining the gene recombinant of the nuclear protein of the present application or a fragment peptide thereof, or a specific artificial sequence peptide, the following may be applied in addition to this. First, when bacteria are used as hosts, vectors for expressing fusion proteins using tags and the like are commercially available. Further, the gene recombinant of the present application also includes a recombinant in which a tag or a part of the peptide is added.

The tag added to the nuclear protein of the present application or a fragment peptide thereof, or a specific artificial sequence peptide is not particularly limited as long as it does not affect the activity of the nuclear protein or peptide, and examples thereof include a histidine tag (for example, 6×His tag). , 10×His), HA tag, FLAG tag, GST tag, T7-tag, HSV-tag, E-tag, lck tag, B-tag and the like.

Regarding yeast, it is known that Pichia genus yeast is effective in expressing a protein having a sugar chain. In terms of sugar chain addition, an expression system using a baculovirus vector that hosts insect cells is also useful (Bio/Technology, 6:47-55, 1988). Furthermore, transfection of a vector using a promoter such as CMV, RSV, or SV40 has been carried out using mammalian cells, and any of these host/vector systems is a nuclear protein of the present application or a fragment thereof. It can be used as an expression system for a peptide or a specific artificial sequence peptide. Further, it is also possible to introduce the gene using a viral vector such as a plasmid vector, a retrovirus vector, a lentivirus vector, an adenovirus vector, an adeno-associated virus vector, a Sendai virus vector, a Sendai virus envelope vector, or a papillomavirus vector. However, the present invention is not limited to these. The vector may contain a promoter DNA sequence that effectively induces gene expression, a factor that controls gene expression, and a molecule necessary for maintaining DNA stability.

The obtained nuclear protein of the present application or a fragment peptide thereof, or a specific artificial sequence peptide can be isolated from inside or outside of the host cell (medium etc.) and purified as a substantially pure and homogeneous protein or peptide. it can. Separation and purification of the protein or peptide may be carried out by using the separation and purification methods used in ordinary protein purification, and is not limited at all. For example, a chromatography column, filter, ultrafiltration, salting out, solvent precipitation, solvent extraction, distillation, immunoprecipitation, SDS-polyacrylamide gel electrophoresis, isoelectric focusing, dialysis, recrystallization, etc. are appropriately selected, If combined, proteins or peptides can be separated and purified.

Examples of chromatography include affinity chromatography, ion exchange chromatography, hydrophobic chromatography, gel filtration, reverse phase chromatography, adsorption chromatography, etc. (Marshak et al., Strategies for Protein Purification and Characterization: A Laboratory Course Manual.Ed Daniel R.Cold Spring Harbor Laboratory Press, 1996). These chromatographys can be performed using liquid phase chromatography, for example, liquid phase chromatography such as HPLC and FPLC.

Further, the nuclear protein of the present application or a fragment peptide thereof, or a specific artificial sequence peptide is preferably a substantially purified nuclear protein or peptide. Here, "substantially purified" means that the degree of purification of the nuclear protein or peptide (the ratio of the nuclear protein or peptide in the whole protein component) is 50% or more, 60% or more, 70% or more, 80%. Above, it means 90% or more, 95% or more, 100% or close to 100%. The upper limit close to 100% depends on the purification technique and analytical technique of those skilled in the art, and is, for example, 99.999%, 99.99%, 99.9%, 99%.

Further, as long as it has the above-mentioned degree of purification, what is purified by any purification method is included in the substantially purified nuclear protein or peptide. For example, the above chromatography column, filter, ultrafiltration, salting out, solvent precipitation, solvent extraction, distillation, immunoprecipitation, SDS-polyacrylamide gel electrophoresis, isoelectric focusing, dialysis, recrystallization, etc. By selection or combination, a substantially purified nuclear protein or peptide can be exemplified, but the present invention is not limited thereto.

On the other hand, the nuclear protein or peptide of the present application can also be artificially synthesized. In the peptide synthesis method of the present application, peptides can be chemically synthesized by methods such as peptide liquid phase synthesis method and peptide solid phase synthesis method. The solid-phase peptide synthesis method is one of the methods generally used in chemically synthesizing peptides. Beads of polystyrene polymer gel with a diameter of about 0.1 mm whose surface is modified with amino groups are used as the solid phase, from which amino acid chains are extended one by one by a dehydration reaction. When the target peptide sequence is completed, it is cut out from the solid phase surface to obtain the target substance. By the solid-phase synthesis method, it is possible to synthesize a ribosomal peptide that is difficult to synthesize in bacteria, introduce an unnatural amino acid such as a D-form or a heavy atom-substituted form, and modify a peptide or protein main chain. When synthesizing 70 to more than 100 long peptide chains by the solid phase method, it is possible to synthesize by linking two peptide chains using a native chemical ligation method. The nuclear protein of the present application or a fragment peptide thereof, or a specific artificial sequence peptide may be in the form of a pharmaceutically acceptable salt of the protein or peptide. Examples of the pharmaceutically acceptable salt include, but are not limited to, hydrochloride, acetate, trifluoroacetate and the like. The nuclear protein of the present application or a fragment peptide thereof, or a specific artificial sequence peptide may be in the form of a solvate of the protein or peptide or a solvate of a pharmaceutically acceptable salt of the protein or peptide. Good. The solvate refers to a molecule in which an arbitrary number of solvent molecules are coordinated with a solute molecule, and includes, for example, a hydrate, but is not limited thereto.

The amino acid length of the nuclear protein or its fragment peptide or the specific artificial sequence peptide in the present application is, for example, in the range of 25 to 35 amino acids, 20 to 40 amino acids, 10 to 50 amino acids, 10 to 70 amino acids, 10 to 100 amino acids, etc. However, the present invention is not limited to these.

The nuclear protein or a fragment peptide thereof in the present application includes, for example, a nuclear protein selected from the following or a fragment peptide derived therefrom:
1. BTF3 protein (Basic transcription factor 3);
2. SUPT16H protein (Suppressor of Ty 16 Homolog; or Facilitates chromatin transcription complex subunit SPT16);
3. YBX1 protein (Y-Box binding protein 1; or Nuclease-sensitive element-binding protein 1);
4. NPM1 protein (Nucleophosmin 1);
5. PA2G4 protein (Proliferation-associated protein 2G4);
6. PFDN5 protein (Prefoldin subunit 5);
7. PSMC3 protein (Proteasome (Prosome, Macropain) 26S subunit, ATPase 3; or 26S proteasome regulatory subunit 6A);
8. HNRNPK protein (Heterogeneous nuclear ribonucleoprotein K);
9. HMGB1 protein (High mobility group box 1);
10. HMGB2 protein (High mobility group box 2);
11. HMGB3 protein (High mobility group box 3);
12. S100A8 protein (S100 calcium-binding protein A8);
13. S100A9 protein (S100 calcium-binding protein A9);
14. IL-1 family (Interleukin-1 family); and
15. A nuclear protein functionally equivalent to a protein selected from 1 to 14.

In light of the examples of the fragment peptides described in the examples of the present application and the prior art, the nuclear proteins selected from 1 to 14 above are considered to have the activity of mobilizing bone marrow mesenchymal stem cells into peripheral blood. Therefore, the term “functionally equivalent” described in 15 above means functional equivalent regarding the activity of mobilizing bone marrow mesenchymal stem cells into peripheral blood. Therefore, the nuclear protein described in 15 above can be expressed as a nuclear protein having an activity equivalent to the protein selected from 1 to 14 (activity equivalent to mobilizing bone marrow mesenchymal stem cells into peripheral blood).

The nuclear protein or fragment peptide thereof selected from 1 to 15 above has an activity of mobilizing bone marrow mesenchymal stem cells into peripheral blood, and is therefore considered to have an effect of mobilizing bone marrow mesenchymal stem cells into peripheral blood. To be

In the present application, examples of the nuclear protein or a fragment peptide thereof include a nuclear protein selected from the following or a fragment peptide derived therefrom:
(I) a nuclear protein comprising an amino acid sequence selected from SEQ ID NOs: 1 to 34 and 131 to 144;
(II) a nuclear protein consisting of an amino acid sequence selected from SEQ ID NOs: 1 to 34 and 131 to 144;
(III) A nuclear protein containing an amino acid sequence in which one or more amino acids are substituted, deleted, inserted or added in the amino acid sequence selected from SEQ ID NOs: 1 to 34 and 131 to 144;
(IV) a nuclear protein consisting of an amino acid sequence in which one or more amino acids are substituted, deleted, inserted or added in the amino acid sequence selected from SEQ ID NOs: 1 to 34 and 131 to 144;
(V) a nuclear protein comprising an amino acid sequence having a sequence identity of about 80% or more with an amino acid sequence selected from SEQ ID NOs: 1 to 34 and 131 to 144;
(VI) a nuclear protein consisting of an amino acid sequence having a sequence identity of about 80% or more with an amino acid sequence selected from SEQ ID NOs: 1 to 34 and 131 to 144;
(VII) a nuclear protein encoded by a DNA consisting of a nucleotide sequence selected from SEQ ID NOs: 57 to 90 and 147 to 160; and (VIII) a nucleotide sequence selected from SEQ ID NOs: 57 to 90 and 147 to 160 Is a nuclear protein encoded by a DNA that hybridizes with the DNA consisting of DNA under stringent conditions.

In the present application, examples of the fragment peptide of the nuclear protein include fragment peptides selected from the following:
(I) a fragment peptide of a nuclear protein consisting of a part of an amino acid sequence selected from SEQ ID NOs: 1 to 34 and 131 to 144;
(Ii) a fragment peptide of a nuclear protein containing an amino acid sequence selected from SEQ ID NOs: 35 to 56 and 145;
(Iii) a fragment peptide of a nuclear protein consisting of an amino acid sequence selected from SEQ ID NOs: 35 to 56 and 145;
(Iv) a fragment peptide of a nuclear protein consisting of a part of an amino acid sequence selected from SEQ ID NOs: 35 to 56 and 145;
(V) A fragment peptide of a nuclear protein containing an amino acid sequence in which one or more amino acids are substituted, deleted, inserted or added in the amino acid sequence selected from SEQ ID NOs: 35 to 56 and 145;
(Vi) a nuclear protein fragment peptide consisting of an amino acid sequence in which one or more amino acids are substituted, deleted, inserted or added in the amino acid sequence selected from SEQ ID NOs: 35 to 56 and 145;
(Vii) a fragment peptide of a nuclear protein containing an amino acid sequence having a sequence identity of about 80% or more with an amino acid sequence selected from SEQ ID NOs: 35 to 56 and 145;
(Viii) a fragment peptide of a nuclear protein consisting of an amino acid sequence having a sequence identity of about 80% or more with an amino acid sequence selected from SEQ ID NOs: 35 to 56 and 145;
(Ix) a nuclear protein fragment peptide encoded by a DNA consisting of a nucleotide sequence selected from SEQ ID NOs: 91 to 112 and 161; and (x) a nucleotide sequence selected from SEQ ID NOs: 91 to 112 and 161 Fragment peptide of a nuclear protein encoded by a DNA that hybridizes with the following DNA under stringent conditions.

In the present application, "a plurality" means, for example, 1 to 10, 1 to 9, 1 to 8, 1 to 7, 1 to 6, 1 to 5, 1 to There are four, one to three, or one or two.

In the present application, "about 80% or more" means, for example, about 85% or more, about 90% or more, about 91% or more, about 92% or more, about 93% or more, about 94% or more, about 95% or more, about 96. % Or more, about 97% or more, about 98% or more, or about 99% or more.

In the present application, “stringent conditions” can indicate conditions such as 6×SSC, 40% formamide, hybridization at 25° C., and washing at 1×SSC, 55° C. The stringency depends on conditions such as salt concentration, formamide concentration, or temperature, and those skilled in the art can set these conditions so as to obtain the required stringency.

When the hybridization is carried out under stringent conditions, a DNA having high homology as a nucleotide sequence is selected, and as a result, the isolated protein has amino acids selected from SEQ ID NOs: 1 to 34 and 131 to 144. It is more likely that a protein functionally equivalent to the protein consisting of the sequence (for example, homologue), or a fragment peptide functionally equivalent to the fragment peptide consisting of the amino acid sequence selected from SEQ ID NOs: 35 to 56 and 145 is included. A highly homologous base sequence can show, for example, about 60% or more, about 70% or more, or about 80% or more identity.

In the present application, examples of the fragment peptide of the nuclear protein include fragment peptides selected from the following:
(1) A fragment peptide consisting of a part of the amino acid sequence selected from SEQ ID NOs: 1 to 2, comprising the amino acid sequence of SEQ ID NO: 35;
(2) A fragment peptide consisting of a part of the amino acid sequence set forth in SEQ ID NO:3, the fragment peptide containing the amino acid sequence set forth in SEQ ID NO:36;
(3) A fragment peptide consisting of a part of the amino acid sequence of SEQ ID NO: 4, comprising an amino acid sequence selected from SEQ ID NOs: 37 to 39;
(4) A fragment peptide consisting of a part of an amino acid sequence selected from SEQ ID NOs: 5 to 7, which fragment fragment peptide contains an amino acid sequence selected from SEQ ID NOs: 40 to 41;
(5) A fragment peptide consisting of a part of the amino acid sequence set forth in SEQ ID NO:8, the fragment peptide comprising the amino acid sequence set forth in SEQ ID NO:42;
(6) A fragment peptide consisting of a part of the amino acid sequence set forth in SEQ ID NO:9, the fragment peptide containing the amino acid sequence set forth in SEQ ID NO:43;
(7) A fragment peptide consisting of a part of the amino acid sequence of SEQ ID NO: 10, comprising the amino acid sequence of SEQ ID NO: 44;
(8) A fragment peptide consisting of a part of the amino acid sequence selected from SEQ ID NOs: 11 to 16 and containing the amino acid sequence set forth in SEQ ID NO: 45;
(9) A fragment peptide consisting of a part of the amino acid sequence selected from SEQ ID NOs: 17 to 18, comprising the amino acid sequence set forth in SEQ ID NO: 46;
(10) A fragment peptide consisting of a part of the amino acid sequence set forth in SEQ ID NO: 19 and containing the amino acid sequence set forth in SEQ ID NO: 47;
(11) A fragment peptide consisting of a part of the amino acid sequence set forth in SEQ ID NO: 20, comprising an amino acid sequence selected from SEQ ID NOs: 48 to 50;
(12) A fragment peptide consisting of a part of the amino acid sequence selected from SEQ ID NOs: 21, 23 and 24, comprising the amino acid sequence of SEQ ID NO: 51;
(13) A fragment peptide consisting of a part of the amino acid sequence selected from SEQ ID NOs: 21 to 26, comprising the amino acid sequence set forth in SEQ ID NO: 52;
(14) A fragment peptide consisting of a part of the amino acid sequence of SEQ ID NO: 27, which contains the amino acid sequence of SEQ ID NO: 53;
(15) A fragment peptide consisting of a part of the amino acid sequence selected from SEQ ID NOs: 28 to 29, which comprises the amino acid sequence set forth in SEQ ID NO: 54;
(16) A fragment peptide consisting of a part of the amino acid sequence of SEQ ID NO: 30, which contains the amino acid sequence of SEQ ID NO: 55;
(17) A fragment peptide consisting of a part of the amino acid sequence selected from SEQ ID NOs: 31 to 34, which comprises the amino acid sequence set forth in SEQ ID NO: 56; and (18) from <1> below When two amino acid sequences (both are amino acid sequences in the amino acid sequence selected from SEQ ID NOs: 131 to 133) are selected from the group of <74> and the longer one is A and the shorter one is B, the SEQ ID NOs: : A peptide consisting of all or part of A in the amino acid sequence selected from 131 to 133, and containing B:
<1> an amino acid sequence consisting of amino acids 1 to 44;
<2> Amino acid sequence consisting of the 1st to 25th amino acids;
<3> Amino acid sequence consisting of the 1st to 34th amino acids;
<4> Amino acid sequence consisting of the 1st to 42nd amino acids;
<5> Amino acid sequence consisting of amino acids 1 to 43;
<6> Amino acid sequence consisting of the 1st to 45th amino acids;
<7> an amino acid sequence consisting of the 1st to 46th amino acids;
<8> an amino acid sequence consisting of the 1st to 47th amino acids;
<9> An amino acid sequence consisting of the 1st to 48th amino acids;
<10> An amino acid sequence consisting of the 1st to 49th amino acids;
<11> An amino acid sequence consisting of the 1st to 50th amino acids;
<12> An amino acid sequence consisting of the 1st to 51st amino acids;
<13> Amino acid sequence consisting of the 1st to 52nd amino acids;
<14> an amino acid sequence consisting of the 1st to 62nd amino acids;
<15> An amino acid sequence consisting of the 1st to 84th amino acids;
<16> an amino acid sequence consisting of the 10th to 25th amino acids;
<17> an amino acid sequence consisting of the 11th to 25th amino acids;
<18> an amino acid sequence consisting of the 11th to 27th amino acids;
<19> an amino acid sequence consisting of the 11th to 28th amino acids;
<20> an amino acid sequence consisting of the 11th to 29th amino acids;
<21> An amino acid sequence consisting of the 11th to 30th amino acids;
<22> an amino acid sequence consisting of the 11th to 34th amino acids;
<23> an amino acid sequence consisting of the 11th to 44th amino acids;
<24> an amino acid sequence consisting of the 12th to 25th amino acids;
<25> An amino acid sequence consisting of the 12th to 30th amino acids;
<26> An amino acid sequence consisting of the 13th to 25th amino acids;
<27> An amino acid sequence consisting of the 13th to 30th amino acids;
<28> An amino acid sequence consisting of the 14th to 25th amino acids;
<29> An amino acid sequence consisting of the 14th to 30th amino acids;
<30> an amino acid sequence consisting of the 15th to 25th amino acids;
<31> an amino acid sequence consisting of the 15th to 30th amino acids;
<32> an amino acid sequence consisting of the 16th to 25th amino acids;
<33> An amino acid sequence consisting of the 16th to 30th amino acids;
<34> an amino acid sequence consisting of the 17th to 25th amino acids;
<35> an amino acid sequence consisting of the 17th to 30th amino acids;
<36> an amino acid sequence consisting of the 45th to 74th amino acids;
<37> an amino acid sequence consisting of the 45th to 84th amino acids;
<38> an amino acid sequence consisting of amino acids 45 to 215;
<39> an amino acid sequence consisting of the 55th to 84th amino acids;
<40> an amino acid sequence consisting of the 63rd to 215th amino acids;
<41> an amino acid sequence consisting of the 1st to 70th amino acids;
<42> an amino acid sequence consisting of the 1st to 81st amino acids;
<43> an amino acid sequence consisting of the 1st to 170th amino acids;
<44> an amino acid sequence consisting of the 2nd to 25th amino acids;
<45> Amino acid sequence consisting of the 2nd to 34th amino acids;
<46> an amino acid sequence consisting of the 2nd to 42nd amino acids;
<47> an amino acid sequence consisting of the 2nd to 43rd amino acids;
<48> an amino acid sequence consisting of the 2nd to 44th amino acids;
<49> an amino acid sequence consisting of the 2nd to 45th amino acids;
<50> An amino acid sequence consisting of the 2nd to 46th amino acids;
<51> an amino acid sequence consisting of the 2nd to 47th amino acids;
<52> an amino acid sequence consisting of the 2nd to 48th amino acids;
<53> an amino acid sequence consisting of the 2nd to 49th amino acids;
<54> an amino acid sequence consisting of the 2nd to 50th amino acids;
<55> An amino acid sequence consisting of the 2nd to 51st amino acids;
<56> an amino acid sequence consisting of the 2nd to 52nd amino acids;
<57> an amino acid sequence consisting of the 2nd to 62nd amino acids;
<58> an amino acid sequence consisting of the 2nd to 70th amino acids;
<59> Amino acid sequence consisting of the 2nd to 81st amino acids;
<60> Amino acid sequence consisting of the 2nd to 84th amino acids;
<61> an amino acid sequence consisting of the 2nd to 170th amino acids;
<62> an amino acid sequence consisting of the 85th to 169th amino acids;
<63> an amino acid sequence consisting of the 89th to 185th amino acids;
<64> an amino acid sequence consisting of the 89th to 195th amino acids;
<65> an amino acid sequence consisting of the 89th to 205th amino acids;
<66> an amino acid sequence consisting of the 89th to 215th amino acids;
<67> an amino acid sequence consisting of the 93rd to 215th amino acids;
<68> an amino acid sequence consisting of the 17th to 44th amino acids;
<69> an amino acid sequence consisting of the 1st to 185th amino acids;
<70> an amino acid sequence consisting of the 1st to 195th amino acids;
<71> an amino acid sequence consisting of the 1st to 205th amino acids;
<72> an amino acid sequence consisting of the 2nd to 185th amino acids;
<73> an amino acid sequence consisting of the 2nd to 195th amino acids; and <74> an amino acid sequence consisting of the 2nd to 205th amino acids.

The peptide according to (18) above is, for example, a peptide consisting of all or part of the amino acid sequence from the 1st to 84th amino acids in the amino acid sequence selected from SEQ ID NOs: 131 to 133, and is one of the following: A peptide comprising the amino acid sequence of:
-The 17th to 25th amino acid sequences in the amino acid sequence selected from SEQ ID NOs: 131 to 133;
-The 45th to 74th amino acid sequences in the amino acid sequence selected from SEQ ID NOS: 131 to 133; and-the 55th to 84th amino acid sequences in the amino acid sequence selected from SEQ ID NOS: 131 to 133.

The amino acid sequences shown in SEQ ID NOs: 1 to 56 are the amino acid sequences of the proteins or peptides shown in Tables 1-1 and 1-2 below.

The base sequences shown in SEQ ID NOs: 57 to 112 are examples of the base sequences of DNAs encoding the proteins or peptides shown in Tables 2-1 and 2-2 below. Other DNA sequences that encode the proteins or peptides set forth in Tables 2-1 and 2-2 below, use codon tables known to those of skill in the art to identify the codons that correspond to the amino acid residues of the protein or peptide. Can be created by the method of converting to (reverse translation). Reverse translation can be carried out, if desired, using various software (including programs, algorithms, etc.) developed for analysis of amino acid and nucleic acid sequences.

In the present application, the nuclear protein or a fragment peptide thereof is, for example, a nuclear protein or a fragment peptide containing a nuclear localization signal. The nuclear localization signal (NLS) is an amino acid sequence having a certain pattern and having a function of translocating a protein/peptide having the same into the nucleus.

For example, many nuclear proteins are known to have NLS in their amino acid sequences and bind to a nuclear transport receptor (also called a nuclear transport protein or nuclear transport factor) that recognizes the NLS. It moves into the nucleus.

Examples of currently known NLS (hereinafter known NLS) include those listed in Table 3-1 and Table 3-2 below:

Also, as known NLS, sequences registered in the NLSdb database (https://rostlab.org/services/nlsdb/) can be mentioned. The sequences registered in NLSdb can be viewed and downloaded from the above website. The NLS sequences registered in NLSdb, whose Annotation type is “Experimental” or “ByExpert”, can be evaluated as having the function of translocating a protein/peptide into the nucleus, and thus are known in the present application. Treat as NLS.

The NLS in this application may be NLS predicted using a specific program (hereinafter, prediction NLS). Whether or not the predicted NLS is included in the desired amino acid sequence is determined by the following program: SeqNLS (Lin et al., PLoS One. 2013 Oct 29;8(10):e76864) or NLStradamus (Nguyen et al., BMC Bioinformatics). 2009 Jun 29;10:202).

In one aspect, the NLS is a known NLS. In one embodiment, the NLS is a known NLS selected from the group consisting of cNLS, PY-NLS, BIB domain and BIB domain-like sequences. In one aspect, the cNLS is a monopartite cNLS. In one embodiment, the monopartite cNLS is KKEK (SEQ ID NO:130).

In the present application, the amino acid sequences shown in SEQ ID NOs: 131 to 146 are the amino acid sequences of the proteins or peptides shown in Table 4 below.

The nucleotide sequences shown in SEQ ID NOs: 147 to 162 are examples of the nucleotide sequences of DNAs encoding the proteins or peptides shown in Table 5 below.

In the present application, “artificial sequence peptide” refers to a peptide having an amino acid sequence that does not exist in nature. Further, in the present application, the “artificial sequence peptide” is also simply referred to as “artificial peptide”.

In the present application, artificial sequence peptides include, for example, peptides selected from:
i. an artificial sequence peptide comprising the amino acid sequence of SEQ ID NO: 146;
ii. An artificial sequence peptide consisting of the amino acid sequence of SEQ ID NO: 146;
iii. An artificial sequence peptide consisting of a part of the amino acid sequence of SEQ ID NO: 146;
iv. An artificial sequence peptide containing an amino acid sequence in which one or several amino acids are substituted, deleted, inserted or added in the amino acid sequence of SEQ ID NO: 146;
v. An artificial sequence peptide consisting of the amino acid sequence of SEQ ID NO: 146 in which one or several amino acids have been substituted, deleted, inserted or added;
vi. An artificial sequence peptide comprising an amino acid sequence having a sequence identity of about 90% or more with the amino acid sequence of SEQ ID NO: 146;
vii. An artificial sequence peptide consisting of an amino acid sequence having about 90% or more sequence identity with the amino acid sequence of SEQ ID NO: 146;
viii. An artificial sequence peptide encoded by the DNA consisting of the nucleotide sequence of SEQ ID NO: 162; and
ix. An artificial sequence peptide encoded by a DNA that hybridizes with the DNA consisting of the nucleotide sequence of SEQ ID NO: 162 under stringent conditions.

In the present application, examples of "several" include 1 to 5, 1 to 4, 1 to 3, or 1 or 2.

In the present application, "about 90% or more" means, for example, about 90% or more, about 91% or more, about 92% or more, about 93% or more, about 94% or more, about 95% or more, about 96% or more, about 97. % Or more, about 98% or more, or about 99% or more.

The artificial sequence peptide selected from the above i to ix is an artificial sequence peptide having an activity of mobilizing bone marrow mesenchymal stem cells into peripheral blood. Therefore, it is considered that these artificial sequence peptides have an effect of mobilizing bone marrow mesenchymal stem cells into peripheral blood.

The amino acid sequence set forth in SEQ ID NO: 146 is the amino acid sequence of the artificial sequence peptide 1r10 of the present application. The base sequence shown in SEQ ID NO: 162 is an example of the base sequence of the DNA encoding the artificial sequence peptide 1r10 of the present application.

In the present application, a cell for secreting the protein or peptide, a gene therapy vector into which a DNA encoding the protein or peptide is inserted may be used instead of the artificial sequence peptide, the nuclear protein or the fragment peptide thereof.

The substance having an activity of mobilizing myelopoietic stem cells into peripheral blood in the present application includes, as cytokines, hepatocyte growth factor (HGF) (JP 2006-104177), G-CSF (granulocyte colony-stimulating factor), GM. -CSF (granulocyte-macrophage colony-stimulating factor), IL-7, IL-12, flt-3 ligand, stem cell factor (SCF), thrombopoietin, and IL-8, MIP-2 (macrophage inflammatory protein) as chemokines -2), BB-10010 (MIP-1α) and the like (WO2003/043651), but not limited thereto.

In this application, the administration method can be appropriately selected depending on the age and symptoms of the patient. For example, when protein is administered, for example, the dose can be selected within the range of 0.0000001 mg to 1000 mg per 1 kg of body weight per administration. Alternatively, for example, the dose can be selected within the range of 0.00001 to 100000 mg/body per patient. Also, when a gene therapy vector into which a DNA encoding a protein is inserted is administered, the amount of the protein can be administered within the above range. However, the drug in the present application is not limited to these doses.

In the present application, the term “patient” is used interchangeably with “subject”, “individual” and “animal”.

The drug of the present application can be formulated according to a conventional method (for example, Remington's Pharmaceutical Science, latest edition, Mark Publishing Company, Easton, USA), and includes both pharmaceutically acceptable carriers and additives. Good. For example, surfactants, excipients, coloring agents, flavoring agents, preservatives, stabilizers, buffers, suspending agents, isotonic agents, binders, disintegrating agents, lubricants, fluidity enhancing agents, flavoring agents. However, the carrier is not limited thereto, and other commonly used carriers can be appropriately used. Specifically, light anhydrous silicic acid, lactose, crystalline cellulose, mannitol, starch, carmellose calcium, carmellose sodium, hydroxypropyl cellulose, hydroxypropylmethyl cellulose, polyvinyl acetal diethylaminoacetate, polyvinylpyrrolidone, gelatin, medium-chain fatty acid triglyceride, Examples thereof include polyoxyethylene hydrogenated castor oil 60, sucrose, carboxymethyl cellulose, corn starch, and inorganic salts.

All the prior art documents cited in this specification are incorporated herein by reference.

The present invention is further illustrated by the following examples, but is not limited thereto.

(Example 1) Induction of immune tolerance by bone marrow cell transplantation of GFP Tg mouse

Preparation of transplanted cells GFP transgenic mice (GFPTg mice) expressing GFP in systemic cells were used as donors. First, bone marrow (femur) of 8-week-old GFPTg mouse was collected using a 26-G needle, and the collected bone marrow was passed through a 40-μm mesh nylon filter, and then centrifuged at 440xg for 4 minutes at 4°C. A separation operation was performed. After suspending the separated precipitate in a lysis buffer (RBC Lysis Buffer, manufactured by BioLegend) to lyse red blood cells, centrifugal force is again performed at 440 xg, 4°C for 5 minutes, and then the supernatant is removed, The precipitate cells were collected. The collected cells were suspended in phosphate buffered saline (PBS) to give a cell suspension (bone marrow cell suspension).

Multiple biotin-labeled monoclonal antibodies (biotin-labeled anti-CD5 antibody, biotin-labeled anti-CD45R antibody, biotin-labeled anti-CD11b antibody, biotin-labeled anti-Gr-1(Ly-6G/C) antibody, biotin were added to the prepared bone marrow cell suspension. Labeled anti-Ly-6B.2 (7/4) antibody, biotin-labeled anti-Ter-119 antibody) was added to the above cell suspension and mixed for a predetermined time, and then washed anti-biotin microbeads (Miltenyi Biotec Company) was added and mixed for 10 minutes, and allowed to stand at 4°C for 15 minutes. Subsequently, the cell suspension was introduced into a magnetic bead cell separation device (device name: autoMACS Pro Separator (Miltenyi Biotec)). Then, cells (Lineage (Lin)-negative cells) that were not captured by the column and flowed out were collected.

Biotin-labeled anti-Lineage antibody and fluorescently-labeled antibody were added to the collected cell suspension, and flow cytometry (device name FACSARIAII (BD Biosciences) was used to generate hematopoietic stem cells (Hematopoietic stem cells (HSC)), The cells were fractionated into mesenchymal stem cells (MSC) and bone marrow cells (non-stem cells) that are neither hematopoietic stem cells nor mesenchymal stem cells.

Specifically, the hematopoietic stem cells are antibodies (allophycocyanin (APC)-labeled anti-C-kit antibody, phycoerythrin (PE)-labeled anti-Sca-1 antibody, biotin-labeled anti-Lineage antibody (biotin-labeled anti-CD5 antibody, biotin-labeled). Anti-CD45R antibody, biotin-labeled anti-CD11b antibody, biotin-labeled anti-Gr-1(Ly-6G/C) antibody, biotin-labeled anti-Ter-119 antibody)), c-kit positive, Sca-1 positive, Lineage negative I picked ones.

For mesenchymal stem cells, BV421-labeled anti-PDGFRa antibody and biotin-labeled anti-Lineage antibody (biotin-labeled anti-CD45R antibody, biotin-labeled anti-CD31 antibody) were used, and PDGFRa-positive and Lineage-negative cells were selected.

Regarding Lineage-positive cells supplemented with biotin-labeled antibody, streptavidin PerCP-Cy5.5 (Thermo Fisher Scientific) was used as a secondary antibody and fractionated prior to selection by flow cytometry. The fractionated cells (Lineage positive cells) were used as non-stem cells (BMC-HSC-MSC) in the subsequent experiments. BMC-HSC-MSC indicates that non-stem cells are bone marrow cells (BM) excluding hematopoietic stem cells (HSC) and mesenchymal stem cells (MSC).

For each of bone marrow cell suspension (bone marrow cells), fractionated hematopoietic stem cells, mesenchymal stem cells, and non-stem cells, 10 ² cells were suspended in 20 μl of PBS to further improve skin engraftment. 2.5×10 ⁶ cells of wild-type mouse bone cells (WT-BMC) were added to prepare an injection.

Cell transplantation to recipient mice Newborn mice up to about 10th of age, where the immune system has not been established, can be regarded as human fetuses, so we decided to adopt mouse newborns at this time as recipients. In this example, mice within 24 hours after birth were used as recipients. Then, using a glass needle of φ75 μm under a microscope, the injection prepared as described above into the superficial temporal vein or facial vein of the prepared newborn mouse (within 24 hours after birth) of the wild-type mouse (C57BL/6) A total volume of 20 μl of the agent was injected. By doing so, the GFP protein derived from the GFPTg mouse serves as a model of the non-genetic mother-derived antigen for the recipient mouse.

Skin-transplanted GFPTg (C57BL/6) mouse tail skin at the age of 6 weeks for each recipient mouse transplanted with hematopoietic stem cells, mesenchymal stem cells, non-stem cells, or bone marrow cells A 10 mm square graft was prepared and transplanted to the back skin of each recipient mouse. For comparison, skin transplantation was also performed on 6-week-old wild-type mice (untreated mice) in which GFPTg mouse cells had not been transplanted. The grafts were sutured with 3-0 absorbent thread and then fixed with band aid for 7 days. After transplantation, the remaining grafts were observed weekly using a GFP fluorescent light.

ELISA evaluation method Blood was collected from the orbital venous plexus of recipient mice 4 weeks after skin transplantation, and serum was stored at -20°C. Recombinant GFP was adjusted to a concentration of 0.2 μg/ml, 100 μl PBS was adjusted, injected into each well of a 96-well plate and kept at 4°C overnight, and then washed buffer (washing buffer, 0.05% Tween20 (registered) The cells were washed twice with (trademark)-containing PBS, SIGMA ALDLICH) to remove the recombinant GFP that did not adhere to the wells. Thereafter, 200 μl of a blocking buffer (PBS containing 1% BSA) was added, and the mixture was kept at room temperature for 1 hour and then washed with a washing buffer.

1 μl of the stored recipient mouse serum was diluted 200 times, 100 μl was added to each well to which recombinant GFP was fixed, and the mixture was kept at room temperature for 1 hour. Then, after washing with a washing buffer, 100 μl of horseradish peroxidase-labeled anti-mouse IgG antibody (anti-mouse IgG, horseradish peroxidase, diluted 1:2500 in PBST, SIGMA ALDLICH) was added, and the mixture was kept at room temperature for 1 hour. Then, after washing with a washing buffer to remove excess unreacted anti-mouse IgG antibody, 3,3',5,5'-tetramethylbenzidine (3,3',5,5'-Tetramethyl benzidine, TMB) was added to each well, and then the absorbance at 450 nm was measured using an absorptiometer (device name: TriStar ² Multimode Reader LB 942, Berthold Technologies), and the measured absorbance was produced in the skin-transplanted mouse. The amount of anti-GFP antibody was calculated.

ELISA Evaluation Results FIG. 1 shows the results of the ELISA evaluation of Example 1. FIG. 1 shows the relative ratio of the amount of anti-GFP antibody on the vertical axis. The amount of anti-GFP antibody produced in each group of recipient mice was compared with that of the untreated mouse with the highest antibody titer. The maximum value is corrected to 1, the average value is represented by a bar graph, and the standard deviation is represented by an error bar. The bar graph shown by "BMC" is the antibody production amount of the group of recipient mice transplanted with bone marrow cells of GFP Tg mouse, and the bar graph shown by "HSC" is the antibody produced by transplantation of hematopoietic stem cells of GFP Tg mouse. The amount of production, and the bar graph indicated by "MSC" is the amount of antibody production of recipient mice transplanted with mesenchymal stem cells of GFPTg mice. The bar graph shown by "non-stem cells" is the antibody production amount of the recipient mice transplanted with the non-stem cells of GFPTg mouse, and the bar graph shown by "control" is the cell transplantation shown for comparison. Results of treated mice.

As can be seen from Fig. 1, the amount of anti-GFP antibody produced was significantly reduced in the group of mice that had undergone some kind of stem cell transplantation, compared to untreated mice. In addition, compared with non-stem cell transplantation, the group of mice transplanted with bone marrow cells, hematopoietic stem cells, and mesenchymal stem cells had lower anti-GFP antibody production, that is, immune tolerance was effectively induced. Admitted. In addition, when evaluated by Student's T test, each of the recipient mouse transplanted with bone marrow cells, the recipient mouse transplanted with hematopoietic stem cells, the recipient mouse transplanted with mesenchymal stem cells, and the untreated mouse were examined. , A significant difference was observed, and the results of the T-test were p=0.005, p=0.003, and p=0.0043, respectively (in FIG. 1, "significant difference" is indicated by "asterisk (*)"). In addition, in the comparison between the untreated mouse and the recipient mouse transplanted with the non-stem cells, it was shown that there was no significant difference (indicated by N.S. in FIG. 1). That is, it was revealed that not only bone marrow cells and hematopoietic stem cell transplants but also mesenchymal stem cell transplants contribute to the induction of B cell line immunological tolerance, whereas the immunological tolerance cannot be induced by transplantation of differentiated cells. It was also revealed (the ability to induce is extremely low).

Immune response evaluation to GFP by ELISPOT using Cytotoxic T-Cell (CTL) After transplanting GFP Tg mouse cells immediately after birth, at the timing of reaching 10 weeks and 11 weeks old, it was intraperitoneally injected into the recipient mouse. Bone marrow cells of GFPTg mice were administered. The bone marrow cells to be administered are 1 to 2×10 ⁷ cells divided into two doses and administered at 10 weeks and 11 weeks, respectively, so that the total of 2 administrations will result in 1 to 2×10 ⁷ cells. Went to.

After that, the spleen was collected at the age of 13 weeks after the recipient mouse was born. The collected spleen was fragmented using a filter, and then centrifuged at a centrifugal force of 300×g for 10 minutes. The precipitate was treated with a lysis buffer (RBC Lysis Buffer, manufactured by BioLegend) for 2 minutes, and then centrifuged again at a centrifugal force of 440xg at 4°C for 5 minutes. The supernatant was removed from the sample after centrifugation, and the cells of the precipitate (spleen cells) were collected.

Bone marrow was collected from the GFPTg mouse, passed through a 40 μm mesh nylon filter, and then centrifuged at 440×g at 4° C. for 5 minutes. After suspending the separated precipitate in a lysis buffer (RBC Lysis Buffer, BioLegend) to lyse the red blood cells, centrifuge again at 440 xg at 4°C for 5 minutes to remove the supernatant. The precipitate was recovered, mitomycin C was added, and the mixture was kept for 30 minutes. After that, 10 ml of PBS in which the collected splenocytes (5×10 ⁶ cells/ml) were suspended and 10 ml of PBS in which GFPTg mouse bone marrow cells (5×10 ⁵ cells/ml) were suspended were mixed to make a total of 20 ml. , 1 μl of recombinant human interleukin 2 (rhIL-2) was added, and coculture was performed at 37° C. for 4 days.

After 4 days of co-culture, only splenocytes were collected as floating cells, and the collected splenocytes:GFPTg mouse bone marrow cells were mixed at a ratio of 25:1, and anti-interferon γ was coated as the primary antibody. It was added to each well of the well plate and co-cultured at 37° C. for 48 hours. After the supernatant was aspirated and washed 4 times, anti-interferon γ secondary antibody (biotin-labeled antibody) was coated at 100 µ/well and left still (4°C overnight). After the supernatant was aspirated and washed 3 times, streptavidin-AP (R&D Systems) 100 μ/well was added and left standing at room temperature for 2 hours. After washing 3 times with Washing buffer, the mixture was protected from light and BCIP/NBT chromogen (R&D systems) was added at 100 μ/well and left standing for 30 minutes. It was washed with distilled water, heated at 37° C. for 15-30 minutes, and IFN-γ stained on the bottom of the plastic plate was counted as the number of spots.

For comparison, bone marrow cells of GFPTg mice were intraperitoneally administered to wild-type mice (untreated mice) that had not been transplanted with GFPTg mice, and treatment and evaluation similar to those of recipient mice were performed. I went.

ELISPOT Evaluation Results Figure 2 shows the results of the ELISPOT evaluation. FIG. 2 is a box and whisker plot in which the number of spots is shown on the vertical axis of the graph, and the number of spots observed is shown by box and whiskers for each group of recipient mice. Observation images of spots are displayed above the graph in association with each group.

Box whiskers shown in "BMC" are the results of a group of recipient mice transplanted with bone marrow cells of GFP Tg mice, and box whiskers shown in "HSC" are the results of recipient mice transplanted with hematopoietic stem cells of GFP Tg mice. The box-and-whiskers indicated by “MSC” are the results of the recipient mice transplanted with the mesenchymal stem cells of the GFPTg mouse. Box whiskers shown as “non-stem cells” are the results of recipient mice transplanted with non-stem cells of GFPTg mice, and box whiskers shown as “control” are cells that were not transplanted as shown for comparison. Results of treated mice.

As can be seen from FIG. 2, untreated mice had a large number of spots and were shown to have high immunoreactivity to GFP. In addition, it was shown that even in the mice transplanted with "non-stem cells", many spots were generated, and high immunoreactivity to GFP was obtained. On the other hand, it was confirmed that in the mice transplanted with bone marrow cells, hematopoietic stem cells, and mesenchymal stem cells, spots were scarcely generated, and immunoreactivity was significantly reduced.

In addition, when the result was evaluated by a statistical test (Student T test), the recipient mouse transplanted with bone marrow cells, the recipient mouse transplanted with hematopoietic stem cells, and the recipient mouse transplanted with mesenchymal stem cells were respectively evaluated. And that of untreated mice were significantly different, and the results of T test were shown to be p=0.0042, p=0.0260, p=0.0244, respectively (in Fig. 2, there is a significant difference). Displayed with "asterisk (*)". On the other hand, it was shown that there was no significant difference between recipient mice transplanted with non-stem cells and untreated mice (indicated by N.S. in FIG. 2). That is, it was revealed that not only bone marrow cells and hematopoietic stem cell transplants but also mesenchymal stem cell transplants contribute to the induction of T cell line immunological tolerance, whereas the immunological tolerance cannot be induced by transplantation of differentiated cells. It was also revealed (the ability to induce is extremely low).

(Example 2) HSC/MSC transplantation experiment from PDGFRaKI mouse to recipient mouse

Preparation of transplanted cells (hematopoietic stem cells and mesenchymal stem cells), cell transplantation into recipients, skin transplantation, and ELISA evaluation were performed in the same manner as in Example 1, except that the donor mouse was changed from the GFPTg mouse to the PDGFRaKI mouse. , ELISPOT was evaluated.

PDGFRaKI mice are heterozygous knock-in mice in which PDGFRa-positive cells (eg, mesenchymal stem cells) express GFP (a histone H2B-GFP fusion gene is inserted at the PDGFRa gene locus). Mouse expressing GFP). The results of experiments using this PDGFRaKI mouse as a donor are shown in FIGS. 3 and 4. In this example, the PDGFRaKI mouse-derived GFP protein serves as a model of the non-genetic mother-derived antigen for the recipient mouse.

Results of ELISA Evaluation FIG. 3 is a diagram showing the results of ELISA evaluation in Example 2. FIG. 3 shows the relative ratio of the amount of anti-GFP antibody on the vertical axis thereof. The maximum value is corrected to 1, the average value is represented by a bar graph, and the standard deviation is represented as an error bar. The bar graph shown by "HSC" is the antibody production amount of the recipient mouse transplanted with the hematopoietic stem cells of the PDGFRaKI mouse, and the bar graph shown by "MSC" is the antibody of the recipient mouse transplanted with the mesenchymal stem cells of the PDGFRaKI mouse. The production amount. The bar graph labeled "control" is the result of untreated mice that did not undergo cell transplantation shown for comparison.

As can be seen from FIG. 3, when GFP-expressing cells (mesenchymal stem cells) were transplanted into recipient mice, the amount of anti-GFP antibody produced was significantly suppressed, and immunity to GFP was effectively suppressed. It was observed that tolerance was induced.

When the results were evaluated by a statistical test (Student T test), there was a significant difference (p=0.0127) between the recipient mice transplanted with mesenchymal stem cells and the untreated mice. Was shown (in FIG. 3, a significant difference is indicated by “asterisk (*)”). On the other hand, it was shown that there was no significant difference between recipient mice transplanted with hematopoietic stem cells and untreated mice (indicated by N.S. in FIG. 3). That is, in order to induce B cell line immunological tolerance to a specific protein (GFP in this experiment), stem cells that express the protein or stem cells that differentiate into cells that express the protein (in this experiment, mesenchymal system) (Stem cell) transplantation was found to be involved.

ELISPOT evaluation result The evaluation result by ELISPOT in Example 2 is shown in FIG. FIG. 4 is a box and whisker plot in which the number of spots is shown on the vertical axis of the graph, and the observed number of spots is shown by a box and whisker for each group of recipient mice. Further, an observation image of the spot is displayed above the graph in association with each group.

The boxplots indicated by "HSC" are the results of recipient mice transplanted with hematopoietic stem cells of PDGFRaKI mice, and the boxplots indicated by "MSC" are the results of recipient mice transplanted with mesenchymal stem cells of PDGFRaKI mice. Is. Box whiskers shown as “control” are results of untreated mice that did not undergo cell transplantation shown for comparison.

As can be seen from FIG. 4, untreated mice had a large number of spots and were shown to have high immunoreactivity to GFP. It was also observed that recipient mice transplanted with hematopoietic stem cells also had an immune response to GFP. On the other hand, it was confirmed that the mice transplanted with the mesenchymal stem cells had almost no spots and the immunoreactivity was significantly reduced.

When the results were evaluated by a statistical test (Student T test), there was a significant difference (p=0.0265) between the recipient mice transplanted with mesenchymal stem cells and the untreated mice. It was shown (in FIG. 4, "asterisk (*)" indicates significant difference). On the other hand, no significant difference was observed between the recipient mouse transplanted with hematopoietic stem cells and the untreated mouse (p=0.1378) (indicated by N.S. in FIG. 4). That is, in order to induce B cell line immunological tolerance to a specific protein (GFP in this experiment), stem cells that express the protein or stem cells that differentiate into cells that express the protein (in this experiment, mesenchymal system) (Stem cell) transplantation was found to be involved.

(Example 3) Skin graft experiment using PDGFRaKI mouse

Female PDGFRaKI mice were mated with male wild-type mice, and 6-week-old F1 mice that were born were subjected to skin transplantation of GFPTg mice in the same manner as in Example 1. For comparison, skin transplantation was similarly performed on B6 wild type mice (C57BL/6). The survival of the graft was observed using a GFP fluorescent light to evaluate the survival rate. For the determination of engraftment, it was judged that it was removed when the green GFP expression was not visible by irradiating it with ultraviolet light after wearing protective sunglasses.

The results are shown in Figure 5. FIG. 5: is a figure which showed the skin-engraftment rate of the mouse|mouth which performed the skin transplant in Example 3. As F1 mice, a mouse expressing GFP and a mouse not expressing GFP appear, but in FIG. 5, the result of the mouse expressing GFP is shown as F1Pa, and the result of the mouse not expressing GFP is shown as F1WT. .. In addition, the results of wild-type mice to be compared are shown as WT. In this example, GFP for F1WT is the non-genetic mother-derived antigen.

FIG. 5(a) is a graph showing the change over time in the rate of skin engraftment, and the vertical axis represents the ratio of the number of engrafted individuals after that to the number of engrafted individuals on day 0 of skin transplantation as the engraftment rate. The horizontal axis indicates the number of weeks that have passed since the skin transplantation day was the reference day. In addition, FIG. 5(b) is a table showing the number of engrafted individuals (the number of mice) every 5 weeks for each group of mice (F1Pa, F1WT, WT).

As can be seen from FIG. 5, FIWT is an individual that does not express GFP (GFP is an antigen), but the graft survival rate of transplanted GFPTg mouse skin is higher than that of wild-type mouse, and the FIWT is transmitted via the mother. It was shown that immune tolerance was induced in mice. In logrank.test, the difference was significant (logrank.test p<0.05). In addition, this result shows that even if the protein is not originally possessed by the offspring, immune tolerance can be induced through the mother.

(Example 4) Skin graft experiment of HMGB1-administered mouse pups

A female PDGFRaKI mouse is crossed with a male wild-type mouse and consists of amino acid residues 1-44 (SEQ ID NO: 145) of the human-derived HMGB1 protein in an amount of 3.0 mg per kg of the weight of a pregnant mother mouse. The peptide (hereinafter referred to as “HMGB1”) was dissolved in 10 μl of PBS to give a single injection solution (3.0 mg/kg, 10 μl/body), and the pregnant mouse was given to the child mouse at a predetermined fetal age (E6). .5, E9.5, E13.5, E16.5), and a total of 4 times through the tail vein.

Then, at the stage when the born offspring mouse (offspring mouse not expressing GFP) was 6 weeks old, skin transplantation of the GFPTg mouse was performed in the same manner as in Example 1. The survival of the grafts was observed using a GFP fluorescent light and evaluated as a survival rate. Results are shown in FIG. In this example, GFP is a non-genetic mother-derived antigen for the offspring mouse.

Skin-grafting experiment of PBS-fed mice

Administration to mother mice and skin transplantation to offspring mice (mice that do not express GFP) were performed in the same manner as in Example 4, except that PBS containing HMGB1 was changed to PBS. The results are shown in FIG. 6 together with Example 4. In this example, GFP is the non-genetic mother-derived antigen for the offspring mouse.

FIG. 6 is a diagram showing the skin engraftment rate of the mice transplanted with skin in Example 4. In FIG. 6, the results of the HMGB1-administered group offspring mice (pup mice that do not express GFP) of Example 4 are shown as HMGB1, and the results of the PBS-administered group offspring mice (pup mice that do not express GFP) are shown as PBS. Show.

FIG. 6(a) is a graph showing the change over time in the skin engraftment rate, and the vertical axis represents the ratio of the engraftment number after that to the engraftment number on day 0 of skin transplantation as the engraftment rate. The horizontal axis indicates the number of weeks that have passed since the skin transplantation day was the reference day. Further, FIG. 6(b) is a table showing the number of engrafted individuals (mouse) every 5 weeks for the mice (HMGB1, PBS) in each group.

As can be seen from FIG. 6, the survival rate of transplanted skin (GFPTg mouse skin) in child mice in the HMGB1 administration group exceeded 20% even at 40 weeks, which was 5% of that in the PBS administration group. It was higher than 9%. The difference was evaluated as significant (logrank.test p<0.05) in logrank.test. That is, it was shown that immune tolerance to GFP (protein not expressed in the offspring mouse), which is supposed to be an antigen, was effectively induced. That is, it was shown that the administration of HMGB1 further induces immune tolerance, as compared with the immune tolerance induced by natural microchimerism from the mother.

(Example 5) Mobilization of mesenchymal stem cells by fragment peptide of nuclear protein

(1) Materials and Methods i) Production of Peptides Fragment peptides of nuclear proteins shown in the following table were chemically synthesized by the solid phase method (the obtained peptides are all in the form of trifluoroacetic acid (TFA) salt). Is).

ii) Administration of peptide C57BL/6J mice (8 weeks old, male, body weight 25 g) were prepared and divided into a group to which the peptides NP-1 to 11 described in the above table were administered and a control group. The peptide was administered by injecting a solution of each peptide adjusted to a concentration of 1 μg/μL with physiological saline as a solvent into the tail vein at a dose of 100 μL/animal (the dose of peptide was 4 mg/kg). .. For the control group, physiological saline was injected into the tail vein at a volume of 100 μL/animal.

iii) Cell recovery from peripheral blood After a certain time (NP-1 to 6 and 8 after 14 hours, NP-7, 10 and 11 after 16 hours, administration of physiological saline or peptide NP-1-11, NP, NP -9 after 24 hours), about 800-1000 μL of peripheral blood was collected from the heart under general anesthesia (using a 1 mL syringe containing heparin). To remove red blood cells, Hetasep (STEMCELL Technologies, Cat No. ST-07906) in an amount equal to that of the collected blood was added, centrifuged at 100 G for 2 minutes, incubated at room temperature for 15 minutes, and then the supernatant was recovered. The supernatant was subjected to the next experiment as a sample containing nucleated cells in peripheral blood.

iv) Colony assay The supernatant obtained by the above procedure (peripheral blood-derived cell-containing sample) was seeded on a collagen I-coated 6-well plate (Corning, Cat No. 356400), and the MesenCult Expansion Kit (STEMCELL Technologies). (Cat No. ST-05513), and 1% L-glutamine (Nacalai Tesque), 10 μM ROCK inhibitor (Y27632, Tocris Bioscience) and 1% penicillin in Expansion Medium prepared according to the manual of the kit. Using a medium containing streptomycin (Nacalai Tesque, Inc.) (the numerical values are all final concentrations), the cells were cultured for 10 days under the conditions of 37° C., 5% CO ₂ and 5% O ₂ . During the culture period, the medium was replaced with fresh one twice a week. On the 10th day of culture, the cells on the plate were stained with a Differential Quik Stain Kit (Cat No. 16920, Sysmex Corporation), and the number of colonies containing 50 or more cells was counted.

In the experiments conducted by the inventors so far, all colonies obtained as a result of culturing peripheral blood on a solid phase such as a dish or a plate have the ability to adhere to the solid phase and self-renewal ability. , PDGFRα positive, and confirmed to have the ability to differentiate into bone, cartilage, fat, epithelium, etc.

In addition, peripheral blood on a solid phase after administration of a peptide consisting of amino acid residues 1-44 of human HMGB1 protein (hereinafter, HMGB1 peptide 1-44) having an activity of mobilizing mesenchymal stem cells into peripheral blood All colonies obtained as a result of culturing have adhesion to the solid phase and self-renewal ability, and have been confirmed to be PDGFRα-positive, and the data of the transcriptome analysis are clustered for gene ontology analysis. From the results obtained, it has been confirmed that it has a gene expression profile characteristic of mesenchymal stem cells.

Furthermore, it has been confirmed that the number of colonies obtained by solid phase culture in the peripheral blood after administration of HMGB1 peptide 1-44 is larger than that in the peripheral blood after administration of physiological saline.

Therefore, the colonies obtained as a result of culturing peripheral blood on the solid phase are mesenchymal stem cells, and the increase in the number of colonies detected in the solid phase culture of peripheral blood indicates the increase in the number of mesenchymal stem cells in the peripheral blood. it is conceivable that.

Moreover, since mesenchymal stem cells do not normally exist in peripheral blood, it is considered that the increased amount of mesenchymal stem cells was mobilized in peripheral blood from a tissue other than peripheral blood (for example, bone marrow).

From the above, the number of colonies detected in solid-phase culture of peripheral blood after administration of the test substance can be used as an index of the activity of the test substance to mobilize mesenchymal stem cells into peripheral blood.

(2) Results In any of the mice administered with the peptides NP-1 to 11, the number of colonies obtained on the plate by culturing the peripheral blood-derived cells was larger than that in the mice administered with physiological saline ( 7-10).

As described above, since an increase in the number of colonies detected by the colony assay described in the present specification indicates an increase in the number of mesenchymal stem cells in peripheral blood, these results show that the fragment peptide of the nuclear protein shows mesenchymal stem cells. It demonstrates that it has an activity of mobilizing in peripheral blood.

(Example 6) Recruitment of mesenchymal stem cells by artificial sequence peptides

(1) Materials and Methods i) Production of Peptide An artificial sequence peptide consisting of the amino acid sequence of SEQ ID NO: 146 was chemically synthesized (the obtained peptide is in the form of trifluoroacetic acid (TFA) salt). Hereinafter, the peptide is referred to as "artificial sequence peptide 1r10" or "peptide 1r10".

ii) Administration of peptide C57BL/6J mice (8 weeks old, male, weight 25 g) were prepared and divided into a group to which artificial sequence peptide 1r10 was administered and a control group. The peptide was administered by injecting a solution of peptide 1r10 adjusted to a concentration of 1 μg/μL with physiological saline as a solvent into the tail vein at a dose of 100 μL/animal (4 mg/kg as a peptide dose). .. For the control group, physiological saline was injected into the tail vein at a volume of 100 μL/animal.

iii) Cell Recovery from Peripheral Blood 14 hours after the administration of physiological saline or artificial sequence peptide 1r10, about 800-1000 μL of peripheral blood was collected from the heart under general anesthesia (using a 1 mL syringe containing heparin). To remove red blood cells, Hetasep (STEMCELL Technologies, Cat No. ST-07906) in an amount equal to that of the collected blood was added, centrifuged at 100 G for 2 minutes, incubated at room temperature for 15 minutes, and then the supernatant was recovered. The supernatant was subjected to the next experiment as a sample containing nucleated cells in peripheral blood.

As described above, the colonies obtained as a result of culturing peripheral blood on the solid phase are mesenchymal stem cells, and the increase in the number of colonies detected in the solid phase culture of peripheral blood is due to the increase in the number of mesenchymal stem cells in the peripheral blood. Is considered to indicate. Therefore, the number of colonies detected in the solid phase culture of peripheral blood after administration of the test substance can be used as an index of the activity of the test substance to recruit mesenchymal stem cells into peripheral blood.

(2) Results In the mouse administered with peptide 1r10, the number of colonies obtained on the plate by culturing cells derived from peripheral blood was larger than that in the mouse administered with physiological saline (FIG. 11).

As described above, since the increase in the number of colonies detected by the colony assay described herein indicates the increase in the number of mesenchymal stem cells in peripheral blood, these results show that peptide 1r10 induces mesenchymal stem cells in peripheral blood. It is demonstrated to have the activity of mobilizing to.

The medicine of the present application can be used as an immunotolerant for inducing immunological tolerance in the fetus of the child in preparation for future disease treatment.

Claims

A medicament for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen for the child or the same antigen by administration to a mother who is gestating the child, wherein bone marrow stem cells are used as peripheral blood. A medicament comprising a substance having an activity of mobilizing therein.
A medicament for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen for the child or the same antigen by administration to a mother who is gestating the child, wherein bone marrow stem cells are used as peripheral blood. A medicament comprising a substance having an activity of mobilizing therein, wherein the bone marrow-derived stem cells of the mother have the non-genetic mother-derived antigen.
The drug according to claim 1 or 2, wherein the drug is a drug for use in combination with another drug for inducing immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as that antigen.
The mother gestating the child is a gestational mother having a diseased child and the medicament is a medicament for use in combination with a medicament for treating the disease in the child. Or the medicine according to 2.
A mother having a child with the disease is a mother without the disease, and a medicament for treating the disease in the child comprises isolated bone marrow-derived stem cells selected from The medicament according to claim 4, which is a medicament for treating the disease in the offspring by transplantation into the offspring during the fetal period.
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. An isolated bone marrow-derived stem cell having the same antigen as.
The mother having a child with the disease is a mother with the disease, and the medicament for treating the disease in the child is derived from a donor who is syngeneic with the mother and does not have the disease. 5. A pharmaceutical for treating the disease in the offspring by transplantation to the offspring in the prenatal period, which comprises an isolated bone marrow-derived stem cell having the same antigen as the non-genetic mother-derived antigen. The described medicine.
An isolated bone marrow-derived stem cell, wherein a mother having a child with the disease is a mother without the disease, and a medicine for treating the disease in the child is selected from the following: The medicament according to claim 4, which is a medicament comprising an isolated tissue or an isolated organ for treating the disease in the offspring by transplantation to the offspring after the neonatal period:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen An organ; and (c) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is allogeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen. Isolated organ.
An isolated bone marrow-derived stem cell, wherein the mother having a child with the disease is a mother with the disease, and a medicine for treating the disease in the child is selected from the following: The medicament according to claim 4, which is a medicament for treating the disease in the offspring by transplanting into the offspring after the neonatal period, which comprises the isolated tissue or the isolated organ:
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen An organ; and (b) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue-derived stem cell derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. Isolated organ.
A medicament for treating the disease in the offspring comprises the same isolated protein as the non-genetic mother-derived antigen for treating the disease in the offspring by administration to the offspring after neonatal period. The medicine according to claim 4, which is a medicine.
The drug according to any one of claims 1 to 9, wherein the substance is HMGB1 protein or a peptide fragment thereof.
A medicament for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child when administered to a mother who is gestating the child, selected from the following: A medicament containing isolated bone marrow-derived stem cells:
(A) an isolated bone marrow-derived stem cell that is derived from a mother who is gestating the offspring and has a non-genetic mother-derived antigen for the offspring; and (b) is derived from a donor syngeneic with the mother An isolated bone marrow-derived stem cell having the same antigen as the mother-derived antigen.
The medicament according to claim 11, wherein the medicament is a medicament for use in combination with another medicament for inducing immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the non-genetic mother antigen.
12. The mother who is gestating the child is a mother who is gestating a child having the disease, and the medicine is a medicine for use in combination with a medicine for treating the disease in the child. The medicine according to.
The mother having a child with the disease is a mother without the disease, and the medicament for treating the disease in the child comprises isolated bone marrow-derived stem cells selected from: The medicament according to claim 13, which is a medicament for treating the disease in the offspring by transplantation into the offspring during the fetal period.
(A) an isolated bone marrow-derived stem cell derived from the mother and having the non-genetic mother-derived antigen; and (b) the non-genetic mother-derived antigen, which is derived from a donor who is syngeneic with the mother and does not have the disease. An isolated bone marrow-derived stem cell having the same antigen as.
The mother having a child with the disease is a mother with the disease, and the medicament for treating the disease in the child is derived from a donor who is syngeneic with the mother and does not have the disease. 14. A medicine for treating the disease in the offspring by transplantation to the offspring in the prenatal period, which comprises an isolated bone marrow-derived stem cell having the same antigen as the non-genetic mother-derived antigen. The described medicine.
A mother having a child with the disease is a mother without the disease, and a medicine for treating the disease in the child has isolated bone marrow-derived stem cells selected from the following: The medicament according to claim 13, which is a medicament for treating the disease in the offspring by transplantation into the offspring after the neonatal period, which includes isolated tissue or an isolated organ.
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated organ derived from the mother and having the non-genetic mother-derived antigen;
(B) an isolated bone marrow-derived stem cell, isolated tissue or isolated derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen An organ; and (c) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue derived from a donor who is allogeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen. Isolated organ.
A mother having a child with the disease is a mother with the disease, and a medicament for treating the disease in the child is an isolated bone marrow-derived stem cell selected from the following: 14. The medicament according to claim 13, which is a medicament for treating the disease in the offspring by transplanting into the offspring after the neonatal period, including the isolated tissue or the isolated organ.
(A) an isolated bone marrow-derived stem cell, isolated tissue or isolated, which is derived from a donor who is syngeneic to the mother and does not have the disease and has the same antigen as the non-genetic mother-derived antigen An organ; and (b) an isolated bone marrow-derived stem cell, an isolated tissue, or an isolated tissue-derived stem cell derived from a donor who does not have the disease allogeneic to the mother and has the same antigen as the non-genetic mother-derived antigen. Isolated organ.
A medicament for treating the disease in the offspring comprises the same isolated protein as the non-genetic mother-derived antigen for treating the disease in the offspring by administration to the offspring after neonatal period. The medicine according to claim 13, which is a medicine.
The medicine according to any one of claims 4 to 9 and 13 to 18, wherein the disease is treatable as a result of induction of immune tolerance in the offspring to the non-genetic mother-derived antigen or the same antigen as the non-genetic mother antigen. ..
The medicine according to any one of claims 4 to 9 and 13 to 18, wherein the disease is treatable as a result of suppressing an immune reaction in the offspring against the non-genetic mother-derived antigen or the same antigen as the non-genetic mother antigen. ..
The medicine according to any one of claims 1 to 20, wherein the stem cells are mesenchymal stem cells or hematopoietic stem cells.
The medicine according to any one of claims 1 to 20, wherein the stem cells are mesenchymal stem cells.
A medicinal stem cell that is used for inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child by administration to a mother who is gestating the child. A medicament comprising a substance having an activity of mobilizing in peripheral blood, wherein the bone marrow-derived stem cells of the mother have the non-genetic mother-derived antigen.
The medicine according to claim 23, wherein the substance is HMGB1 protein or a peptide fragment thereof.
A medicament for use in inducing immune tolerance in a child to a non-genetic mother-derived antigen or the same antigen as that of the child when administered to a mother who is gestating the child, selected from the following: A medicament containing isolated mesenchymal stem cells:
(A) an isolated mesenchymal stem cell that is derived from a mother who is gestating the offspring and has a non-genetic mother-derived antigen for the offspring; and (b) is derived from a donor that is syngeneic with the mother. An isolated mesenchymal stem cell having the same antigen as the genetic mother-derived antigen.