Short-term inhibition of p53 combined with keratinocyte growth factor improves thymic epithelial cell recovery and enhances T-cell reconstitution after murine bone marrow transplantation

Abstract

Myeloablative conditioning before bone marrow transplantation (BMT) results in thymic epithelial cell (TEC) injury, T-cell immune deficiency, and susceptibility to opportunistic infections. Conditioning regimen-induced TEC damage directly contributes to slow thymopoietic recovery after BMT. Keratinocyte growth factor (KGF) is a TEC mitogen that stimulates proliferation and, when given before conditioning, reduces TEC injury. Some TEC subsets are refractory to KGF and functional T-cell responses are not fully restored in KGF-treated BM transplant recipients. Therefore, we investigated whether the addition of a pharmacologic inhibitor, PFT-␤, to transiently inhibit p53 during radiotherapy could spare TECs from radiation-induced damage in congenic and allogeneic BMTs. IntroductionAllogeneic bone marrow transplantation (BMT) is used to treat malignant and nonmalignant disorders. 1,2 Chemoradiotherapy conditioning preceding donor graft infusion damages thymic stroma, severely delaying peripheral CD4 ϩ and CD8 ϩ T-cell reconstitution. [2][3][4][5][6] Thus, BM transplant recipients are at increased risk of opportunistic fungal and viral infections. 7 Thymic stroma is composed of a 3-dimensional matrix of thymic epithelial cells (TECs), fibroblasts, macrophages, dendritic cells (DCs), and mesenchymal cells. 8 Critical signals are supplied by TECs to developing thymocytes directing their thymic ingress, 9,10 survival, 11,12 trafficking, 11 selection, 13 and export. 11 Conditioning depletes TECs, impairing T-cell production for prolonged periods of time after BMT. [14][15][16][17] Restoring thymic function would speed peripheral T-cell recovery after BMT.Developing thymocytes can be distinguished by their CD4 and CD8 cell surface expression. CD4 Ϫ CD8 Ϫ (double-negative, DN) thymocytes mature to become CD4 ϩ CD8 ϩ (double-positive, DP) thymocytes and undergo positive selection on cortical TECs (cTECs). DP thymocytes continue their maturation into CD4 ϩ or CD8 ϩ (single-positive, SP) thymocytes and migrate into the thymic medulla where negative selection is mediated by medullary TECs (mTECs) and medullary DCs. 8 SP thymocytes complete their maturation in the medulla and are exported into the periphery as mature T cells. 13 Keratinocyte growth factor (KGF) is a fibroblast growth factor family member that controls cell migration, proliferation, and differentiation in epithelial tissues. 18 Endogenous KGF is upregulated in mucosal tissues after injury, and exogenous KGF enhances protection/repair of epithelial cells in models of chemoradiotherapy-induced injury. 18 KGF is approved by the Food and Drug Administration for prevention of oral mucositis associated with high-dose chemoradiotherapy and hematopoietic stem cell transplantation. [19][20][21] KGF is produced by thymic mesenchymal cells and binds exclusively to FGFR2-IIIb expressed by TECs. 15,22 KGF is a potent mitogen for TECs. 23 KGF pretreatment prevents thymic injury and prolonged T-cell deficiency in murine BMT models. 15,[23][24][25][26][27] KGF facilitates alloe...

In Vivo Administration of the Recombinant IL-7/Hepatocyte Growth Factor β Hybrid Cytokine Efficiently Restores Thymopoiesis and Naive T Cell Generation in Lethally Irradiated Mice after Syngeneic Bone Marrow Transplantation

“…5A-D, optimal doses of rIL-7/HGFb (15 mg) increased the numbers of total donor-origin CD4 + and CD8 + T cells by 4-7-fold above those in PBS-treated BMT mice, whereas optimal doses of rIL-7 (5 mg) increased the numbers by 2-to 3-fold. In each instance, 70-80% of the donor-origin T cells had a CD62L hi CD44 lo phenotype, characteristic of naive T cells newly derived from the thymus (37,38). Hence, rIL-7/HGFb fully restored the numbers of naive donor-origin T cells to normal, whereas rIL-7 only partially restored their numbers (∼50%).…”

“…This is consistent with previous reports that rIL-7 has a short serum t 1/2 in animals and human (40,54), unless it is administered in a liposome-encapsulated form or as an rIL-7/anti-IL-7 Ab complex (40,55). Collectively, therefore, it appears that rIL-7/HGFb can correct the major causes of delayed thymocyte development following BMT, namely decreased numbers of ETPs, damage to thymic stromal cells, decreased cytokine production, abnormal progression from the DN to the SP stages of thymocyte development, and, in the case of allogenic BMT, GVHD (1,2,37,38). Consequently, it is clear that rIL-7/HGFb holds considerable promise clinically for preventing or reversing post-BMT T cell deficiency and its complications.…”

Thymic epithelial cells require p53 to support their long-term function in thymopoiesis in mice

“…5 Moreover, the use of p53 inhibitors, such as Pifithrin-β, has been approved in clinical trials to attenuate the side effects of chemotherapy. 9 Given the described adverse impact of disrupting p53 in TECs and in T cells,5 the therapeutic use of p53 inhibitors must be implemented with care to safeguard the balance between immune reconstitution and tolerance induction. ribosomal RNA, and those in Ctr mTECs were set as 1.…”

“…Although germline deletion in wild-type p53-induced phosphatase 1, a p53-target gene, impairs the maturation of mTECs and thymic regeneration, 8 the systemic administration of p53 inhibitors moderately improves TEC recovery and thymopoiesis following BMT. 9 Because genetically engineered mouse models and pharmacological studies often hide lineage-specific functions of broadly acting genes, the cell-autonomous relevance of p53 in the dynamic differentiation of TECs in vivo remains unexplored. Our findings underscore the requirement for p53 in TECs to support their role in Tcell development and selection.…”

T cell progenitor therapy–facilitated thymopoiesis depends upon thymic input and continued thymic microenvironment interaction

“…They support homing of progenitors to the thymus (16,17), provide critical survival signals (18,19), direct trafficking of progenitors between the cortex and the medulla (18), and regulate positive and negative selection (20) and egress of single-positive T cells from the thymus (18). Several strategies have been investigated preclinically to preserve TECs during pretransplant conditioning regimens, such as prophylactic administration of keratinocyte growth factor (KGF) (21) and KGF in combination with leuprolide acetate (22) or transient inhibition of p53 prior to irradiation (23). However, these approaches have not yet led to a rapid and full restoration of peripheral T cell recovery early after transplant.…”