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BackgroundThere is a need for therapies that can mitigate bone marrow dysfunction and organ toxicity that occur following myeloablative injury and reduced intensity conditioning regimens used in patients undergoing bone marrow transplantation (BMT). The pathogenesis of adverse effects from BMT conditioning has been linked to injury to the vascular endothelium, bone marrow (BM), and other organs.ObjectiveTo evaluate the impact of the thrombopoietin mimetic drug JNJ-26366821 (TPOm) on BM vascular recovery in mice undergoing myeloablative radiation conditioning followed by BMT.Study designTPOm (doses: 0 µg, 300 µg, 1000 µg per Kg body weight) was administered on Days 0 and 7 after BMT, in mice receiving a total body irradiation (TBI) conditioning regimen (5.5 Gy x 2) before congenic BMT. BM donner cell engraftment was analyzed using flow cytometry on Days 7, 14, and 30 post-BMT. The morphological and biophysical properties of the BM vasculature were evaluated by intravital multiphoton microscopy (MPM) and immunofluorescence confocal imaging. Herein, morphological properties involve microvascular density (MVD), vessel diameter, and vascular area, while biophysical properties include transfer rate (Ktrans) of contrast within the BM vascular niche, as well as the fractional volume (vec) of extracellular extravascular tissue (EES).ResultsNo significant difference in donor chimerism was observed at days 7, 14, and 30 post-BMT, between TPOm and PBS-treated mice. TPOm intervention improved BM vasculature regeneration in transplanted mice. The MVD, Ktrans, and BM vasculature as well as vascular endothelial growth factor receptor-2 (VEGFR2) in the BM, showed a dose dependent improvement in mice treated with TPOm. On day 14 post-BMT, the group receiving 1000 µg/Kg TPOm showed significant shifts (p-value < 0.05) in MVD, Ktrans, and VEGFR2 expression from their corresponding control types (TPOm dose 0 µg) towards levels comparable to healthy controls.ConclusionTPOm intervention augments BM vascular structure and function, which may be important for hematopoietic recovery and bone marrow function in radiation conditioned hematopoietic stem cell transplant patients, in addition to enhancing platelet recovery.

Total body irradiation (TBI) is a commonly used conditioning regimen for hematopoietic stem cell transplant (HCT), but dose heterogeneity and long-term organ toxicity pose significant challenges. Total marrow irradiation (TMI), an evolving radiation conditioning regimen for HCT can overcome the limitations of TBI by delivering the prescribed dose targeted to the bone marrow (BM) while sparing organs at risk. Recently, our group demonstrated that TMI up to 20 Gy in relapsed/refractory AML patients was feasible and efficacious, significantly improving 2-year overall survival compared to the standard treatment. Whether such dose escalation is feasible in elderly patients, and how the organ toxicity profile changes when switching to TMI in patients of all ages are critical questions that need to be addressed. We used our recently developed 3D image-guided preclinical TMI model and evaluated the radiation damage and its repair in key dose-limiting organs in young (~8 weeks) and old (~90 weeks) mice undergoing congenic bone marrow transplant (BMT). Engraftment was similar in both TMI and TBI-treated young and old mice. Dose escalation using TMI (12 to 16 Gy in two fractions) was well tolerated in mice of both age groups (90% survival ~12 Weeks post-BMT). In contrast, TBI at the higher dose of 16 Gy was particularly lethal in younger mice (0% survival ~2 weeks post-BMT) while old mice showed much more tolerance (75% survival ~13 weeks post-BMT) suggesting higher radio-resistance in aged organs. Histopathology confirmed worse acute and chronic organ damage in mice treated with TBI than TMI. As the damage was alleviated, the repair processes were augmented in the TMI-treated mice over TBI as measured by average villus height and a reduced ratio of relative mRNA levels of amphiregulin/epidermal growth factor (areg/egf). These findings suggest that organ sparing using TMI does not limit donor engraftment but significantly reduces normal tissue damage and preserves repair capacity with the potential for dose escalation in elderly patients.

Sickle cell disease (SCD) is a serious global health problem, and currently, the only curative option is hematopoietic stem cell transplant (HCT). However, myeloablative total body irradiation (TBI)-based HCT is associated with high mortality/morbidity in SCD patients. Therefore, reduced-intensity (2–4 Gy) total body radiation (TBI) is currently used as a conditioning regimen resulting in mixed chimerism with the rescue of the SCD disease characteristic features. However, donor chimerism gradually reduces in a few years, resulting in a relapse of the SCD features, and organ toxicities remained the primary concern for long-term survivors. Targeted marrow irradiation (TMI) is a novel technique developed to deliver radiation to the desired target while sparing vital organs and is successfully used for HCT in refractory/relapsed patients with leukemia. However, it is unknown if TMI will be an effective treatment for a hematological disorder like SCD without adverse effects seen on TBI. Therefore, we examined preclinical feasibility to determine the tolerated dose escalation, its impact on donor engraftment, and reduction in organ damage using our recently developed TMI in the humanized homozygous Berkley SCD mouse model (SS). We show that dose-escalated TMI (8:2) (8 Gy to the bone marrow and 2 Gy to the rest of the body) is tolerated with reduced organ pathology compared with TBI (4:4)-treated mice. Furthermore, with increased SCD control (AA) mice (25 million) donor BM cells, TMI (8:2)-treated mice show successful long-term engraftment while engraftment failed in TBI (2:2)-treated mice. We further evaluated the benefit of dose-escalated TMI and donor cell engraftment in alleviating SCD features. The donor engraftment in SCD mice completely rescues SCD disease features including recovery in RBCs, hematocrit, platelets, and reduced reticulocytes. Moreover, two-photon microscopy imaging of skull BM of transplanted SCD mice shows reduced vessel density and leakiness compared to untreated control SCD mice, indicating vascular recovery post-BMT.

Background: There is a need for therapies that can mitigate bone marrow dysfunction and organ toxicity that occur following myeloablative injury and reduced intensity conditioning regimens used in patients undergoing bone marrow transplantation (BMT). The pathogenesis of adverse effects from BMT conditioning has been linked to injury to the vascular endothelium, bone marrow (BM), and other organs. Objective: To evaluate the impact of the thrombopoietin mimetic drug JNJ-26366821 (TPOm) on BM vascular recovery in mice undergoing myeloablative radiation conditioning followed by BMT. Study design: TPOm (doses: 0 µg, 300 µg, 1000 µg per Kg body weight) was administered on Days 0 and 7 after BMT, in mice receiving a total body irradiation (TBI) conditioning regimen (5.5 Gy x 2) before congenic BMT. BM donner cell engraftment was analyzed using flow cytometry on Days 7, 14, and 30 post-BMT. The morphological and biophysical properties of the BM vasculature were evaluated by intravital multiphoton microscopy (MPM) and immunofluorescence confocal imaging. Herein, morphological properties involve microvascular density (MVD), vessel diameter, and vascular area, while biophysical properties include transfer rate (Ktrans) of contrast within the BM vascular niche, as well as the fractional volume (vec) of extracellular extravascular tissue (EES). Results: No significant difference in donor chimerism was observed at days 7, 14, and 30 post-BMT, between TPOm and PBS-treated mice. TPOm intervention improved BM vasculature regeneration in transplanted mice. The MVD, Ktrans, and BM vasculature as well as vascular endothelial growth factor receptor-2 (VEGFR2) in the BM, showed a dose dependent improvement inmice treated with TPOm. On day 14 post-BMT, the group receiving 1000 µg/Kg TPOm showed significant shifts (p-value < 0.05) in MVD, Ktrans, and VEGFR2 expression from their corresponding control types (TPOm dose 0 µg) towards levels comparable to healthy controls. Conclusion: TPOm intervention augments BM vascular structure and function, which may be important for hematopoietic recovery and bone marrow function in radiation conditioned hematopoietic stem cell transplant patients, in addition to enhancing platelet recovery. day 14 post-BMT, the group receiving 1000 µg/Kg TPOm showed significant shifts (p-value < 0.05) in MVD, Ktrans, and VEGFR2 expression from their corresponding control types (TPOm dose 0 µg) towards levels comparable to healthy controls. Conclusion: TPOm intervention augments BM vascular structure and function, which may be important for hematopoietic recovery and bone marrow function in radiation conditioned hematopoietic stem cell transplant patients, in addition to enhancing platelet recovery. [ABSTRACT FROM AUTHOR]

Sickle cell disease (SCD) is a serious global health problem, and currently, the only curative option is hematopoietic stem cell transplant (HCT). However, myeloablative total body irradiation (TBI)-based HCT is associated with high mortality/morbidity in SCD patients. Therefore, reduced-intensity (2-4 Gy) total body radiation (TBI) is currently used as a conditioning regimen resulting in mixed chimerism with the rescue of the SCD disease characteristic features. However, donor chimerism gradually reduces in a few years, resulting in a relapse of the SCD features, and organ toxicities remained the primary concern for long-term survivors. Targeted marrow irradiation (TMI) is a novel technique developed to deliver radiation to the desired target while sparing vital organs and is successfully used for HCT in refractory/relapsed patients with leukemia. However, it is unknown if TMI will be an effective treatment for a hematological disorder like SCD without adverse effects seen on TBI. Therefore, we examined preclinical feasibility to determine the tolerated dose escalation, its impact on donor engraftment, and reduction in organ damage using our recently developed TMI in the humanized homozygous Berkley SCD mouse model (SS). We show that dose-escalated TMI (8:2) (8 Gy to the bone marrow and 2 Gy to the rest of the body) is tolerated with reduced organ pathology compared with TBI (4:4)-treated mice. Furthermore, with increased SCD control (AA) mice (25 million) donor BM cells, TMI (8:2)-treated mice show successful long-term engraftment while engraftment failed in TBI (2:2)-treated mice. We further evaluated the benefit of dose-escalated TMI and donor cell engraftment in alleviating SCD features. The donor engraftment in SCD mice completely rescues SCD disease features including recovery in RBCs, hematocrit, platelets, and reduced reticulocytes. Moreover, two-photon microscopy imaging of skull BM of transplanted SCD mice shows reduced vessel density and leakiness compared to untreated control SCD mice, indicating vascular recovery post-BMT. [ABSTRACT FROM AUTHOR]