Limitations to the Field of Transplantation

• Drug treatment-related complications

• Chronic rejection

• Availability of organs

TOLERANCE

• Specific unresponsiveness to the donor of the recipient’s immune system.

• The donor is regarded as “self”.

• Therefore, no immunosuppressive drugs are needed to prevent rejection

Bone Marrow Induces Tolerance

• Animal studies >25 years ago showed that mixed bone marrow chimerism educates the immune system to make it tolerant of the donor

• We have aimed at making this approach less toxic and therefore clinically applicable

Bone Marrow Transplantation for Tolerance Induction:

Requirements

• Recipient treatment must have minimal toxicity

• Must work for mismatched transplants

• Graft-versus-host-disease (GVHD) unacceptable. GVHD is the major complication of bone marrow transplantation, and precludes mismatched transplantation.

USING STEM CELLS TO INDUCE TOLERANCE

1)Treatments are given to block peripheral and intrathymic rejection of donor hematopioetic cells (e.g. anti-T cell mAbs, thymic RTX).Donor stem cellscells are given i.v.

2) Donor stem cells go to recipient marrow. Stem cells in the marrow send progeny to the recipient thymus.

Blood cells are a mixture of donor and host (mixed chimera)

3) New T cells mature and become “educated” in the recipient thymus gland.

RecipientDonor

4) The emerging T cells that repopulate the immune system are tolerant of donor and recipient. A donor organ is accepted and there is no GVHD.

BMT with T cell costimulatory blockade

3 Gy TBI day 0

anti-CD40L-mAb (0.5mg i.p., day 0)CTLA4Ig (0.5mg i.p., day +2)

15x10^6 B10.A bone marrow cells i.v.(fully MHC-mismatched,unseparated day 0)

C57BL/6

Wekerle et al, JEM 1998,187:2037

n=14third party

donor

0 20 40 60 80 100 120 140 1600

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60

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100

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raft

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Days post Skin-Grafting

Donor-Specific Skin Graft Tolerance in Recipients of Non-Myeloablative BMT with Costimulatory Blockade

Bone Marrow/Stem Cell Transplantation

• The only known cure for many types of leukemia and lymphoma.

• Requires an HLA closely matched donor because of the complication of graft-versus-host disease (GVHD).

• Even with unrelated donors, about half of the patients whose only hope for cure is BMT do not have a donor.

GVHD

• Major complication of BMT • Caused by donor T lymphocytes that

see recipient antigens as “non-self”• Disease of skin, liver, intestines• Prevented by marrow T cell depletion,

but this increases relapse rates, because donor T cells also eradicate leukemia cells

Our Goal

• To perform HLA mismatched transplants without GVHD.

• To use the GVH “response” (GVHR) to attack leukemia/lymphoma without producing GVHD. We have discovered that GVHR≠GVHD.

• This will allow even better cure rates than are seen with matched transplants.

Our Strategy

• Stimulate GVHR

• Confine GVHR to the tissues where leukemias and lymphomas reside (blood and lymphoid tissues).

• i.e. avoid migration of GVHR to skin, gut liver

Step 1: Bone marrow transplant with less toxic recipient treatment that includes antibodies.

Donor marrow is T cell depleted

Wait 1-2 months. Inflammation from preparative treatment subsides.

Blood cells are a mixture of donor and host: Mixed chimerism is achieved without GVHR

Step 2: Infuse donor T cells.

Donor T cells interact with “presenting cells” of mixed chimera to maximize GVHR

Tumor is killed

Donor T cells are armed to kill tumor cells that express recipient antigens. They stay inside the blood and lymph, where tumor is.

T cells don’t go to skin/gut/liver. There is no GVHD.

42 y.o. male with disseminated Hodgkin’s Disease, refractory to chemo and radiation therapy. Received a BMT with our protocol in Sept, 1999. Results: No GVHD, complete remission.

1yr post-transplantPre-transplant

Rationale: Combined Matched Related Donor Bone Marrow and Kidney Transplantation in Multiple Myeloma With Kidney Failure

• Allogeneic BMT is the only known cure for MM. Complication rates are high with standard allogeneic BMT.

• Kidney failure is a common complication of MM, but the malignancy usually precludes kidney transplantation.

• Successful allogeneic BMT with less toxic conditioning induces transplantation tolerance (animal models).

• MGH investigators have developed a less toxic BMT protocol that is safe and effective in MM.

• Less toxic BMT combined with kidney transplantation from the same donor might induce tolerance while curing the myeloma.

Wait 1-2 months.

Blood cells are a mixture of donor and host: Mixed chimerism and tolerance to the kidney is achieved.

Step 2: Infuse donor T cells.

Donor T cells interact with “presenting cells” of mixed chimera to maximize GVHR

Tumor is killed

Donor T cells are armed to kill tumor cells that express recipient antigens.

Step 1: Bone marrow and kidney transplant with less toxic recipient treatment.

Combined Kidney and Bone Marrow Transplant: Patient 1

• 55-year-old woman presented in December, 1996 with ESRD due to multiple myeloma.

• Rx: Hemodialysis, chemotherapy• September, 1998: combined kidney and bone

marrow transplant from HLA-identical sister.• 2005: pt in remission from myeloma; normal

kidney function, off all immunosuppression since December, 1998.

Clinical course of patient 1

0

1

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0 5 15 25 35 45 55 65 75 85 95 105 115154200 300500 7009000

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35SerumCRE

CyA Levels

KappaLightChain

500

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100

CyA ng/dl

Scrmg/dl

Kappa mg/dl

Days post-transplant

CyADiscontinued

Applying our Strategy to Mismatched Transplants

• A greater challenge, because T cell depleted mismatched marrow is harder to engraft , especially when less toxic recipient treatment is given

• We have developed protocols achieving engraftment of mismatched, T cell-depleted marrow without GVHD.

• We have obtained proof of principle that our strategy can work in the mismatched setting.

78 79 80 81 82 83 84 85 86 87 88 89 90 91 92 93 94 95 960

500

1,000

1,500

2,000

2,500

3,000

3,500

Heart Transplants Performed World-wide

YEAR

Nu

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roce

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res

POTENTIAL DONOR SPECIES

SwineAdvantages

AvailabilityBreeding Characteristics

DisadvantagesPhylogenetic distanceNatural (anti-GAL) antibodies

GENETIC ENGINEERING OF PIGS AS XENOGRAFT DONORS

• Transgenics– Complement inhibition

• DAF

• CD46

• CD59

– Fucosyl transferase

– Growth factors• pIL-3, pSCF• Human GF receptors

– MHC genes• Class I (NK inhibition)

• Knock-outs 1,3-galactosyl transferase

Fertilized egg

DNA

First GalT-KO miniature swine born November 2002

From: TBRC and Immerge BioTherapeutics, Boston

Anti-CD4Anti-CD8mAbs

Days –6,-1, +7, +14

Normal mouse

Thymectomize

Day 0: Implant 1mm3 fetal miniature swine thymus tissue under kidney capsule

Reconstituted murine CD4 compartment. Tolerance to donor pig.

Replacement of Recipient Thymus With a Xenogeneic Thymus in Thymectomized, T Cell-Depleted Mice

Zhao et al, Nature Medicine 1996, 2:1211

2. Thymokidney transplantation

Tolerance by Thymus Transplantation

From: TBRC and Immerge BioTherapeutics, Boston

KBx laparotomy

Creatinine levels B134 (Thymokidney – Steroid free regimen)

0123456789

10

0 7 14 21 28 35 42 49 56 63 70 77

POD

Cr(

mg

/dl)

From: TBRC and Immerge BioTherapeutics, Boston

B134 kidney graft biopsy on POD60

x200

Normal kidneyKidney graft was pinkNo spot hemorrhage

From: TBRC and Immerge BioTherapeutics, Boston

Summary of Heart and Kidney Transplants from the first available GalT-KO Pigs

• Do not undergo HAR

• Do not require antibody absorption nor complement inhibition

• With standard immunosuppression, organ survivals improved - modestly but consistently

• With kidney plus thymus tolerance strategy, survivals increased from maximum of 30 days to

>83 days

ACKNOWLEDGEMENTS

MGH• BMT Unit (Spitzer, McAfee, Dey, Ballen, et al.)• Transplant Unit (Cosimi, Kawai, Delmonico, Ko, Hertl, et al.)• TBRC (Sachs, Sykes, Yamada, et al.)• Pathology (Colvin, Saidman, et al.)• Infectious Disease (Fishman, Basgoz, et al.)• Renal (Rubin, Williams, Goes, Wong, et al.)• Wellman Photomedicine Laboratories (Lin et al.)

OUTSIDE• ITN (NIH)• Biotransplant/Immerge• Medimmune

CONTRIBUTORS: MOUSE STUDIESCONTRIBUTORS: MOUSE STUDIESCONTRIBUTORS: MOUSE STUDIESCONTRIBUTORS: MOUSE STUDIES

BMT Section/BMT Section/Transplantation Biology Transplantation Biology Research CenterResearch Center

Ronjon ChakravertyRonjon ChakravertyHyeon-Seok EomMarkus MaparaThomas FehrYasuo TakeuchiJosef KurtzJosef KurtzDenise PearsonDenise PearsonJuanita ShafferJuanita ShafferJennifer BuchliJennifer BuchliTim HoganTim HoganPeter CotterPeter CotterGuiling ZhaoGuiling ZhaoRichard HsuRichard Hsu

Wellman Center for Wellman Center for PhotomedicinePhotomedicine

Daniel CoteDaniel CoteCostas PitsillidesCostas PitsillidesCharles LinCharles Lin