Low-dose Total Body Irradiation Research Articles

Is Low Dose Total Body Irradiation Necessary in Peripheral Blood Reduced Intensity Conditioning Haploidentical Transplantation?

Is Low Dose Total Body Irradiation Necessary in Peripheral Blood Reduced Intensity Conditioning Haploidentical Transplantation?

Comparison of fludarabine-based conditioning regimens in adult cord blood transplantation for myeloid malignancy: A retrospective, registry-based study.

Fludarabine/busulfan and fludarabine/melphalan are viable options as conditioning regimens. However, the optimal fludarabine-based conditioning in cord blood transplantation (CBT) remains unclear. Therefore, this retrospective, registry-based study aimed to analyze the impact of five fludarabine-containing conditioning regimens on 1395 adult patients (median age, 61 years) with acute myeloid leukemia, myelodysplastic syndrome, and chronic myeloid leukemia who underwent their first CBT. Treatment outcomes of fludarabine combined with melphalan (100-140 mg/m2 ) and low-dose total body irradiation (TBI; FM140T); melphalan (80-99 mg/m2 ) and TBI (FM80T); busulfan (12.8 mg/kg) and melphalan (FB4M); busulfan (12.8 mg/kg) and TBI (FB4T); and busulfan (6.4 mg/kg) and TBI (FB2T) were compared. The 3-year survival rate was 67%, 53%, 44%, 36%, and 39%, respectively (p < .0001). The FM140T survival rate was the most favorable after adjusting for confounders, and the hazard ratios (vs. FM140T) for overall mortality were as follows: FM80T, 1.6 (95% confidence interval [CI], 1.2-2.2); FB4M, 2.1 (95% CI, 1.6-2.8); FB4T, 2.7 (95% CI, 2.0-3.7); and FB2T, 2.2 (95% CI, 1.6-3.1). The better survival observed with FM140T, regardless of the disease, disease risk, age, or transplant year, was attributed to the lower relapse rate and lower non-relapse mortality (NRM) associated with fewer infectious deaths. Conversely, FB4T was associated with a higher relapse rate and higher NRM. The findings indicate that the outcomes of CBT in myeloid malignancies were highly dependent on both the alkylating agent and its dose in combination with fludarabine. Therefore, compared with fludarabine/busulfan-based conditioning, FM140T may be the preferred regimen.

Low Dose Total Body Irradiation Improves CD19 CAR-T Expansion, Persistence, and Trafficking, Leading to Enhanced Efficacy in Large B-Cell Lymphoma

Anti-CD19 chimeric antigen receptor T-cell (CART19) is associated with high response rates in large B-cell lymphoma (LBCL). However, a significant subset of patients relapses due to several mechanisms that include poor expansion, lack of CART19 persistence, and immunosuppressive tumor microenvironment. We hypothesized that low dose total body irradiation (LDTBI) may interfere with these mechanisms and thus enhance the efficacy of CART19 cells in LBCL. To test this hypothesis, we utilized a syngeneic mouse model of LBCL, as they are immunocompetent, to best emulate patients treated in clinic. This model involves transplanting BALB/c mice with GFP/Luciferase transduced mouse derived B cell lymphoma (A20) cells and generating murine CART19 (mCART19). We tested mCART19 cytotoxicity in vitro against A20 cells with or without radiation. We found that exposing A20 cells to 1 Gy radiation prior to co-culturing them with mCART19 led to a significant improvement of the mCART19 cytotoxicity (E:T ratio 2.5:1; p=0.029). This is consistent with our in vitro findings for human LBCL cell lines, where weobserved that exposing LBCL cell lines to 1 Gy radiation significantly improved human CART19 cytotoxicity (E:T ratio 2.5:1: OCI-Ly10 cells: p=0.009; SU-DHL-4 cells: p=0.048). Next, we assessed the activity of mCART19 cells in vivo. Animals were lymphodepleted using cyclophosphamide (Cy) administered 24 hours before mCART19 treatment. LDTBI at 1 Gy was delivered 4 hours before mCART19 administration. In addition to evaluating the benefit of single LDTBI prior to mCART19, we also delivered a second dose of LDTBI 9 days after mCART19 treatment. Thus, we evaluated 6 cohorts as follows: (1) Untreated n=4, (2) Cy/LDTBI n=5 (3) Cy/LDTBI/LDTBI n=3, (4) Cy/mCART19 n=5, (5) Cy/LDTBI/mCART19 n=5, (6) Cy/LDTBI/mCART19/LDTBI n=5. Adding LDTBI to Cy provided a minor improvement compared to untreated controls (median overall survival [OS]= 21 days vs 16.5 days; p=0.009). As expected, Cy/mCART19 improved median OS compared to controls (median OS= 35 day; p=0.003). Strikingly, 8/10 of the mice treated with Cy/LDTBI/mCART19 remained in complete remission (CR) at 6 months post-treatment ( p&amp;lt;0.001), Figure 1. Since one dose of LDTBI prior to mCART19 was very effective, adding the second dose of LDTBI did not result in additional survival benefit. Peripheral blood (PB) was monitored regularly, we observed a 2.8-fold increase of mCART19 expansion in the Cy/LDTBI/mCART19 cohort when compared with the Cy/mCART19 cohort ( p&amp;lt;0.001) 9 days post treatment. Importantly, bone marrow aspirates were performed 6 months post treatment in animals that remained alive, demonstrating sustained B-cell aplasia in 7/8 mice (Cy/LDTBI/mCART19 cohorts) which strongly suggests mCART19 persistence. One animal showed B-cell recovery but remained in CR with no relapse suggesting potential cure. Mice that received Cy/mCART19 with no LDTBI showed evidence of B-cell reappearance 7 days prior to overt relapse. Next, we assessed mCART19 trafficking into the tumor tissues by establishing new cohorts of (1) Cy/mCART19 n=4, (2) Cy/LDTBI/ mCART19 n=4 and harvesting tumor-infiltrated spleen and liver 3 days post treatment. Flow cytometry revealed that LDTBI cohort had significant increase in the amount of mCART19 in the spleen and the liver (3.15-fold; p=0.041, and 2-fold; p=0.012, respectively) compared to Cy/mCART19 cohort, suggesting that LDTBI improves mCART19 tumor-trafficking ability. We have previously reported that exposing human LBCL cells to 1Gy radiation in vitro increases expression of FAS when compared to non-irradiated controls and that FAS blockade impairs CART19 cytotoxicity. Hence, we evaluated the expression of FAS in murine LBCL A20 cells harvested from the syngeneic mouse model 3 days post LDTBI. Consistently, we observed 3.1-fold increase in FAS expression, ( p=0.033). In conclusion, our results show a novel and efficacious approach to harnessing LDTBI with CART19. Based on our findings, LDTBI enhances the efficacy of CART19 leading to improved survival in a syngeneic mouse model. Mechanistically, radiation improves CART19 efficacy by boosting CART19 expansion, persistence, trafficking into tumor sites, and upregulation of FAS in tumor cells, pointing to a clinically translatable benefit.

Pharmacokinetic-Targeting and Dose-Adjustment of I.V. Busulfan for Myeloablative Conditioning in Allogeneic Hematopoietic Cell Transplantation

Introduction: Pharmacokinetically (PK)-targeting intravenous busulfan improves outcomes following allogeneic hematopoietic cell transplantation (HCT). However, the optimal busulfan area under the curve (AUC) likely differs based on the specific conditioning regimen, and the target busulfan exposure is unknown in combination with fludarabine and low-dose total body irradiation (TBI). Methods: This study included 1019 adult HCT recipients that received myeloablative conditioning including fludarabine, busulfan, anti-thymocyte globulin, and low-dose (4cGy) TBI. Busulfan was administered as a total dose of ~3.2mg/kg given equally from days -5 to -2 pre-transplant. Total AUC was estimated using measurements of serial serum samples. Multivariate Cox and Fine-Gray regression were used for comparison of AUC subgroups. The primary outcomes of interest were relapse-free survival (RFS) and overall survival (OS). Results: Median AUC was 62.3 mg∙hr/L (range: 39.4-128.0 mg∙hr/L). Total AUC exposure of 49.3-57.5 mg∙hr/L was associated with greater RFS (67% vs. 47%, HR=1.82, P=0.014) and OS (71% vs. 46%, HR=1.99, P=0.008) compared to patients with higher AUCs of 57.5-73.9 mg∙hr/L. Although very low (&amp;lt;49.3 mg∙hr/L) or very high (&amp;gt;73.9 mg∙hr/L) AUCs trended towards worse RFS and OS compared to 49.3-57.5 mg∙hr/L, this analysis was limited by the small number of patients with extreme AUCs and did not reach statistical significance. Except potentially for patients with a high/very high HCT disease risk index, 49.3-57.5 mg∙hr/L appeared to be the optimal AUC regardless of patient sex, age, or primary disease. Conclusion: Within the evaluated AUC range, 49.3-57.5 mg∙h/L appeared to be associated with the most favourable survival. Pharmacokinetic-targeting to this range may improve outcomes.

Haploidentical Hematopoietic Cell Transplant with Post-Transplant Cyclophosphamide Expedites Donor-Derived Immune Reconstitution in a Pediatric Severe Aplastic Anemia Cohort

Severe aplastic anemia (SAA) is a life-threatening, idiopathic process involving the immune-mediated destruction of hematopoietic stem cells. Treatment involves immunosuppressive therapy (IST) and/or allogeneic hematopoietic cell transplantation (HCT). The latter has recently been optimized with use of a post-transplant cyclophosphamide (PTCy) based graft-versus-host disease (GvHD) platform, permitting successful HCT with only half-HLA-matched haploidentical donors. In pediatrics, this platform results in &amp;gt;90% overall survival with low rates of GvHD and graft failure. However, reports on donor chimerism and immune reconstitution are limited. We hypothesized that a PTCy platform would expedite CD3 donor engraftment and CD4 T cell recovery. In a phase III clinical trial, we treated three cohorts of pediatric (age &amp;lt;/=25 years) patients with SAA requiring alloHCT: MUD, MSD, and HAPLO. The MUD cohort included n=15 recipients of 7-8/8 HLA-matched unrelated donor grafts (n=5 7/8, n=10 8/8) with median age 14 years (range 2-17 years), following cyclophosphamide (Cy) 50 mg/kg, fludarabine (Flu) 200 mg/m 2, rabbit anti-thymocyte globulin (rATG) 9 mg/kg total dose rabbit and low dose total body irradiation (TBI) at 200 cGy. The MSD cohort included n=9 recipients of 8/8 HLA-matched sibling donor grafts with median age 15 years (range 12-24 years), following Cy 200 mg/kg and 9 mg/kg rATG. The HAPLO cohort included n=4 recipients of 4/8 HLA-matched related donor grafts with median age 8.5 years (range 2-21 years) following Cy 29 mg/kg, Flu 150 mg/m 2, 9 mg/kg rATG, and low dose TBI at 200 cGy. GvHD prophylaxis consisted of 30 days of mycophenolate mofetil, and 100 days of a calcineurin inhibitor (followed by an 8-10 week taper), with the HAPLO cohort additionally receiving post-transplant Cy (PTCy) 50 mg/kg/day on days +3 and +4. Peripheral blood lymphocyte counts and peripheral blood and bone marrow donor chimerism were evaluated on days +100, +180 and +365. Recipient characteristics were largely equivalent between cohorts, with 43% receiving IST prior to HCT. Clonality was overrepresented in the HAPLO cohort (2 of 4, 1 with monosomy 7 myelodysplastic syndrome and 1 with a large PNH clone), with only 2 of 15 in the MUD and none in the MSD cohorts. Active controlled infections at the time of HCT were additionally overrepresented in the HAPLO cohort, with disseminated pseudomonas in 1. The MUD cohort had 1 with Rhizopus invasive sinusitis (none in the MSD cohort). Achievement of neutrophil engraftment was statistically significantly delayed in the HAPLO cohort at a median of 17 days compared to 12 for MSD and 14 for MUD, p=0.004), however there were no episodes of primary or secondary graft failure. With only a single late death from intracranial hemorrhage in the MUD cohort, OS was equivalent at 2 years (100% in MSD and HAPLO cohorts, 93.3% in MUD). GvHD rates were low with only 1 (MUD) experiencing grade II acute GvHD, and 1 (MUD) experiencing mild chronic GvHD. Immune reconstitution varied by cohort with statistically significantly greater CD3 peripheral blood donor chimerism (p&amp;lt;0.001) as well as CD4 (p=0.04) and CD8 (p=0.03) T cell counts over time for the HAPLO recipients (CD3 chimerism in Fig 1, CD4 count in Fig 2; 2-way ANOVA fixed effects model). Considering a CD4 &amp;gt;200 cell/uL as clinically meaningful, we assessed the proportion of each cohort achieving this milestone at each timepoint. While each cohort had an increase in proportion achieving CD4&amp;gt;200 over time, the HAPLO cohort reached 50% with CD4 &amp;gt;200 at day +100, compared to 11% MUD and 0% MSD (p=0.09). At day +180, the proportion with CD4 &amp;gt;200 were 36% MUD, 25% MSD and 75% HAPLO (1 yr: 90% MUD, 100% MSD and HAPLO). Donor availability remains a challenge to successful transplant for patients with SAA, and haploidentical matches overcome donor limited accessibility. PTCy as GvHD prophylaxis is associated with low incidences of graft failure and severe GvHD. While PTCy might be anticipated to inhibit donor T cells and immune reconstitution, our data, albeit limited by small cohorts, demonstrates superior and more efficient CD3 donor engraftment and CD4 T cell count recovery following HAPLO HCT with PTCy compared to non-PTCy based MUD and MSD cohorts.

Impact of Graft Source [Cord Blood Supported By CD34+ Selected Cells (Haplo-Cord) Vs. Matched Donor] on Transplant Outcomes in Myelofibrosis

Introduction: Allogeneic hematopoietic cell transplantation (allo-HCT) is curative for patients with myelofibrosis (MF). Here, we evaluated the impact of donor type [cord blood transplant supported by CD34+ cells (haplo-cord) vs adult matched donor] on outcomes in MF. Methods: All adult patients with primary or secondary (post-ET or post-PV) MF, including those progressing to acute leukemia (AML) who underwent their first allo-HCT at NewYork-Presbyterian Hospital/Weill Cornell Medicine between 01/2015 and 06/2022 were included. Results: 40 patients were transplanted. Median age was 63 years (range 21-77). 23% had a Karnofsky performance scale (KPS) of &amp;lt;80. Two had undergone splenectomy prior to HCT. The median spleen size was 18cm (10-28). Median number of pre-HCT molecular mutations was 2 (range 0-7). 16 (40%) had MF-3 (Table 1). 24 (60%) underwent a matched sibling donor (MSD) (n=9) or unrelated donor (MUD) (n=15) HCT, and 16 (40%) underwent a cord-based (CB) HCT. 39 received Fludarabine/Melphalan +/- low dose total body irradiation (TBI), one underwent myeloablative conditioning with TBI/etoposide based on extensive extramedullary disease. The median time to neutrophil engraftment in the MSD/MUD group was 14 days (IQR 14-18) vs 30 days in the CB group (IQR 15-36) and (p=0.1). The median time to platelet engraftment was 22 days (IQR 17-38) in MSD/MUD vs 49 days (IQR 26-60) in the CB group (P=0.07). The CI of absolute neutrophil count (ANC) and platelet engraftment by day 60 for MSD/MUD vs CB was 92% (95% CI 65-98%) vs 80% (95% CI 45-94%) (p=0.09) and 75% (95% CI 51%- 88%) vs 59% (95% CI 27-81%) (p=0.36), respectively. One patient in the MSD/MUD died without neutrophil engraftment compared to 3 in CB group. 5 patients in each group died without platelet engraftment. Median follow-up for all alive patients was 53 months. Overall survival (OS) for the cohort was 62% at 1 year (95% CI 49-79) and 34% at 3 years (95% CI 21-55). OS at 3 years was similar between MSD/MUD group and CB (32% vs 37%, P=0.9) (figure 1). Graft versus host disease relapse free survival (GRFS) at 3 years was not significantly different in MSD/MUD group, compared to CB group (15% vs 33%, P=0.64). Cumulative incidence of relapse at 3 years in MSD/MUD group was 28% (95% CI 10-49), and 12% (95% CI 1.8-34) in CB group (p=0.356). Non-relapse mortality (NRM) at 1 year was 33% (95% CI 15-52) in MSD/MUD group and 38% (95% CI 15-61) in CB group. NRM at 3 years was 54% (95% CI 29-73) in MSD/MUD group and 48% (95% CI 19-72) on CB group (P=0.95). Discussion: In our small cohort, we showed that OS after haplo-cord transplant was comparable to adult matched donors' grafts (MSD or MUD). While the haplo-cord platforms have been shown to decrease the time to engraftment (11 days for neutrophils and 22 days for platelets) compared with double cord transplants or haploidentical transplants (van Besien K et al . Haematologica 2016, van Besien K et al. Blood Adv 2019), in patients with MF, we saw a trend towards delayed engraftment compared to patients transplanted with adult matched donors' grafts (MSD or MUD). This suggests that the CD34 selected cells may not be able to provide an adequate myeloid bridge and overcome the low cellular content in the CB for adult patients with extensive marrow fibrosis. Larger studies are needed to compare best alternative graft (CB-based versus haploidentical or mismatched unrelated donors) for patients with MF that lack a matched donor.

Proinflammatory Cytokine Release and Infusional Side-Effects after Anti-Thymocyte Globulin Serotherapy for Graft-Versus-Host Disease Prophylaxis in Allogeneic Hematopoietic Cell Transplantation

Background: Anti-thymocyte globulin (ATG) serotherapy for graft-versus-host disease (GVHD) prophylaxis in allogeneic hematopoietic cell transplantation (HCT) is frequently associated with infusional side-effects (ISEs) including fever, tachycardia, respiratory distress, and hypotension/shock. Although this constellation closely resembles cytokine release syndrome, the mechanism behind ATG ISEs is poorly understood. The aim of this study was to characterize the frequency and severity of ATG ISEs, and determine the association of ISEs with cytokine levels and clinical outcomes. Methods: This study included 118 adult HCT recipients receiving myeloablative conditioning including fludarabine, busulfan, low-dose total body irradiation, and 4.5mg/kg ATG (0.5mg/kg on day -2 and 2mg/kg on days -1 and 0). Patient electronic medical records were retrospectively reviewed for clinical data collection. ISE were classified as thermal (fever/rigors), respiratory (tachypnea/hypoxemia/increased O 2 supplementation), tachycardia, or circulatory (hypotension/fluid resuscitation requirement). ISEs were further characterized into mild or severe (e.g., mild tachycardia = 105-120bpm, severe ≥120bpm). Serum levels of 31 cytokines were determined pre- and post-1 st ATG infusion in all patients (and up to 13 timepoints in select patients) using Luminex. Cox and Fine-Gray regression were used to determine the association of ISEs with clinical outcomes including grade 2-4 acute GVHD (aGVHD), moderate-severe chronic GVHD (cGVHD), cumulative incidence of relapse (CIR), and overall survival (OS). Results: At least one ISE occurred in 74% (87/118) of patients, with a maximum severity of mild in 56 (47%) and severe in 31 (26%). Patients experienced an average of 1.4 ISEs (median 1), most commonly tachycardia (57%) and thermal (50%), while respiratory (26%) and circulatory (7%) ISEs were less frequent. ISE frequency was 62%, 60%, and 35% after the 1 st-3 rd ATG infusions, respectively, with the median time from infusion to first ISE being ~4.5 hours. Compared to pre-infusion levels, there was a &amp;gt;1-fold change in 11/34 cytokines (all P&amp;lt;0.001) [Table 1]. Of greatest interest was IL-6, which demonstrated a progressive increase in median levels between patients with no vs. mild vs. severe ISEs (P&amp;lt;0.05). Severe ISEs were associated with a lower incidence of grade 2-4 aGVHD (SHR=0.12, P=0.032), but not with any other outcomes. Patient age, sex, primary disease, or pre-infusion peripheral blood cell counts (leukocytes, lymphocytes, neutrophils, and monocytes) did not appear to influence ISE frequency or severity. Conclusion: ISEs occur in the majority of ATG recipients, with one-fourth experiencing severe ISEs. The concomitant rise in serum cytokine levels (particularly IL-6) suggests that anti-cytokine antibodies (e.g., tocilizumab) may assist in the prevention or treatment of ISEs, however, the impact this may have on outcomes is unclear and warrants investigation in upcoming clinical studies.

Quality of Life and Comorbidities of Adult Survivors of Allogeneic Hematopoietic Cell Transplant Versus Their Siblings

Background: Survivors of allogeneic hematopoietic cell transplant (allo-HCT) are at risk of developing chronic health conditions. Prior studies have shown that survivors of allo-HCT in childhood/young adulthood who mainly received high dose total body irradiation (TBI)-based conditioning experience excess health conditions and more distress vs. their siblings. In recent years, allo-HCT is offered to adults of increasing age and there has been increasing use of chemotherapy +/- low dose TBI conditioning rather than high dose TBI. We set out to describe and compare the QoL and comorbidities of adults who underwent allo-HCT with chemotherapy +/- low dose TBI conditioning vs. their siblings. Methods: Allo-HCT recipients attending 1 of 2 survivorship clinics at ≥2 years post-HCT without active cGVHD or relapse, were enrolled. Beginning in 2023, participants were asked to forward an electronic version of the study survey to their biological sibling(s). The survey includes the PROMIS-Global Health instrument: raw scores are converted to a mental (MH) and physical health (PH) subscale T-score (population mean 50/SD 10). Comorbidities were scored with a published post allo-HCT morbidity index. Associations between MH and PH-related QoL, comorbidities, and patient/transplant-related variables were investigated with linear regression. Patient and sibling comparisons were made with paired sample and chi-square tests. A significance level of &amp;lt;0.05 was statistically significant &amp; a difference in QoL T-score of 5 was clinically significant. Results: Of 330 patients approached during their clinic visit, 320 consented to participate (97%). To date, 105 siblings of 57 recipients have been sent surveys and 47 siblings (45%) of 32 recipients (56%) have completed the survey. Among all recipients, median age at HCT is 47 (IQR 35-57) with 21% ≥60 years old, 48% are female, 78% are Caucasian, 69% are college educated or more &amp; 51% are working full/part-time. Median time post-HCT is 9 years (IQR 6-14). The mean PH T-score is 48.5 (SD 8.6). In the multivariable model, total number of comorbidities (p&amp;lt;0.01) and a work status of disability (p&amp;lt;0.01) are associated with inferior PH QoL. The mean MH T-score was 49.1 (SD 8.4). In the multivariable model, total number of comorbidities (p&amp;lt;0.01) and a work status of disability (p&amp;lt;0.01) are associated with inferior MH QoL. HCT characteristics, including history of moderate-severe cGVHD, and demographic characteristics are not associated with either QoL subscale. Among all recipients, the median number of comorbidities is 3 (IQR 1.5-4). The most common comorbidities include hyperlipidemia (39%), hypertension (25%) &amp; hypothyroidism (24%). At least one severe comorbidity (vascular disease, chronic heart/lung/liver/kidney disease or subsequent malignancy) is present in 23% of patients. In multivariable analysis, a greater number of years post-HCT is associated with a greater number of comorbidities (p=0.03) and full/part-time work status is associated with fewer comorbidities (p=0.01). In the sibling-recipient comparison, median age of siblings and recipients is 55 (IQR 37-66) &amp; 54 (IQR 40-64), respectively. The MH QoL of recipients is statistically, but not clinically, better than their siblings: T-score 52.9 (SD 7.1) versus 49.9 (SD 6.9), respectively, p=0.04. The PH QoL of recipients was not significantly different than their siblings: T-score 52.6 (SD 5.6) versus 50.7 (SD 8.2), respectively, p=0.23. Recipients are suffering from significantly more comorbidities vs. their siblings: median 2.5 (IQR 1.8-4) versus 1 (IQR 0-3), respectively, p=0.03. The prevalence of ≥1 severe comorbidity is greater in recipients vs. siblings, but not statistically significant (13% vs. 2%, p=0.06). Conclusion: Adult recipients of allo-HCT, living without cGVHD or relapse and conditioned without high dose TBI, in aggregate experience similar QoL but suffer excess chronic health conditions as compared to their siblings. One in 4 recipients are suffering from ≥1 severe chronic health condition(s). Time post-HCT is associated with comorbidities, suggesting that recipients continue to accumulate health conditions in the years post-HCT. Number of comorbidities, rather than transplant and demographic factors (including prior cGVHD), is associated with impaired MH and PH QoL. Preventative care after allo-HCT is critical, even in those receiving HCT in adulthood without high dose TBI.

Low-dose total body irradiation enhances systemic anti-tumor immunity induced by local cryotherapy.

Strategies that restore the immune system's ability to recognize malignant cells have yielded clinical benefits but only in some patients. Tumor cells survive cryotherapy and produce a vast amount of antigens to trigger innate and adaptive responses. However, because tumor cells have developed immune escape mechanisms, cryotherapy alone may not be enough to induce a significant immune response. The mice were randomly divided into four groups: Group A: low-dose total body irradiation combined with cryotherapy (L-TBI+cryo); Group B: cryotherapy (cryo); Group C: low-dose total body irradiation(L-TBI); Group D: control group (Control). The tumor growth, recurrence, and survival time of mice in each group were compared and the effects of different treatments on systemic anti-tumor immunity were explored. L-TBI in conjunction with cryotherapy can effectively control tumor regrowth, inhibit tumor lung metastasis, extend the survival time of mice, and stimulate a long-term protective anti-tumor immune response to resist the re-challenge of tumor cells. The anti-tumor mechanism of this combination therapy may be related to the stimulation of inflammatory factors IFN-γ and IL-2, as well as an increase in immune effector cells (CD8+ T cells) and a decrease in immunosuppressive cells (MDSC, Treg cells) in the spleen or tumor tissue. We present unique treatment options for enhancing the immune response caused by cryotherapy, pointing to the way forward for cancer treatment.

Radiation therapy improves CAR T cell activity in acute lymphoblastic leukemia

Autologous T cells engineered to express a chimeric antigen receptor (CAR) specific for CD19 are approved for the treatment of various CD19+ hematological malignancies. While CAR T cells induce objective responses in a majority of patients, relapse frequently occurs upon loss of CD19 expression by neoplastic cells. Radiation therapy (RT) has been successfully employed to circumvent the loss of CAR targets in preclinical models of pancreatic cancer. At least in part, this reflects the ability of RT to elicit death receptor (DR) expression by malignant cells, enabling at least some degree of CAR-independent tumor killing. In a human model of CD19+ acute lymphoblastic leukemia (ALL), we also observed DR upregulation by RT, both in vitro and in vivo. Moreover, low-dose total body irradiation (LD-TBI) delivered to ALL-bearing mice prior to CAR T cell infusion considerably extended the overall survival benefit afforded by CAR T cells alone. Such an improved therapeutic activity was accompanied by a superior expansion of CAR T cells in vivo. These data encourage the initiation of clinical trials combining LD-TBI with CAR T cells in patients with hematological malignancies.

Effect of triple therapy with low-dose total body irradiation and hypo-fractionated radiation plus anti-programmed cell death protein 1 blockade on abscopal antitumor immune responses in breast cancer.

Immunostimulatory effects of radiotherapy can be synergistically augmented with immune checkpoint blockade to act both on irradiated tumor lesions and distant, non-irradiated tumor sites. Our hypothesis was that low-dose total body irradiation (L-TBI) combined with hypo-fractionated radiotherapy (H-RT) and anti-programmed cell death protein 1 (aPD-1) checkpoint blockade would enhance the systemic immune response. We tested the efficacy of this triple therapy (L-TBI+H-RT+aPD-1) in BALB/c mice with bilateral breast cancer xenografts. The L-TBI dose was 0.1Gy. The primary tumor was treated with H-RT (8Gy×3). The PD-1 monoclonal antibody was injected intraperitoneally, and the secondary tumors not receiving H-RT were monitored for response. The triple therapy significantly delayed both primary and secondary tumor growths, improved survival rates, and reduced the number of lung metastasis lesions. It increased the activated dendritic and CD8+ T cell populations and reduced the infiltration of myeloid-derived suppressor cells in the secondary tumor microenvironment relative to other groups. Thus, L-TBI could be a potential therapeutic modality, and when combined with H-RT and aPD-1, the therapeutic effect could be enhanced significantly.

Allogeneic Peripheral Blood Stem Cell Transplantation for Bone Marrow Failure Syndromes Using CD34+-Selected Progenitor Cells Co-Infused with a Reduced Number of Non-Mobilized T-Cells

Background Severe aplastic anemia (SAA) patients who fail immunosuppressive therapy (IST) may be candidates for curative allogeneic hematopoietic stem cell transplantation (HCT). Patients with SAA are often heavily transfused with underlying immune-mediated disease, which increases their risk for graft rejection when bone marrow (BM) is used as a graft source. Peripheral Blood Stem Cell (PBSC) grafts have been associated with better engraftment but at a cost of a higher incidence of graft-versus-host disease (GVHD). We developed a novel protocol combining high numbers of GCSF-mobilized CD34+ peripheral blood stem cells with a BM transplant-equivalent dose of T-cells (collected from donors before GCSF administration) to improve engraftment and decrease the risk of GVHD associated with a conventional, unmanipulated PBSC HCT. Methods In this phase II trial, eligible patients included those age 4-80 years with a diagnosis of SAA or other bone marrow failure (BMF) syndromes with ≥ 9/10 HLA-matched related (sibling/family) donors (MRD) or 10/10 matched unrelated donors (MUD). All patients received conditioning with cyclophosphamide (day -7 to -6), anti-thymocyte globulin (day -5 to -2), and fludarabine (day -5 to -1). For MUD patients, low dose total body irradiation (200 cGy) was added to the regimen on day -1. GVHD prophylaxis included cyclosporin (day -4 until at least day +30) and methotrexate (5 mg/m2 , day +1, +3 and +6). The primary outcome of the study was chronic GVHD at 1 year. Secondary outcomes included mortality, engraftment and chimerism, acute and chronic GVHD, immune reconstitution, and comparisons of MRD vs. MUD patients. Results Thirty-five patients (27 with MRD and 8 with MUD) were transplanted between February 2011 and December 2021. Patients had a median age of 30 (range 6-70) years, 46% were males and 54% were females, including 31 having SAA, 3 with SAA transformed to myelodysplastic syndrome (MDS), and one with MDS; 11 patients also had PNH clone pre-transplant. The median numbers of transplanted CD34+ and CD3+ T-cells were 8 ´ 106 cells/kg and 2 ´ 107 cells/kg, respectively. After HCT transplants, all patients had neutrophil recovery with a median 14 (10-32) days. Except for 2/35 (6%) patients having late graft failure, all other 33 (94%) patients achieved sustained engraftment. Platelet recovery was reached in 29 (83%) patients at a median 19 (13-321) days. A total of 97% of patients achieved >95% donor myeloid chimerism (median 14 days), and 94% of patients achieved >95% donor T-cell chimerism (median 30 days). Reactivation of CMV occurred in 21/29 (72%) patients at risk with none developing CMV disease. Full immune reconstitution occurred for most patients by 1-year post-transplant. With a median follow-up of 5 years, the estimated 200-day survival and overall survival were 89% and 85%, respectively. The cumulative incidence of acute grade II-IV GVHD was 23%, and 1-year chronic GVHD rate was 11%, which was significantly lower than our hypothesis that this regimen would lower the chronic GVHD rate to 50% (P < .001). The subgroups of patients with MRD and MUD were compared (Table 1): patients with MUD showed a slower time to achieve neutrophil recovery (P = .001) and platelet recovery (P = .03), and a slightly high graft failure incidence (2 cases), but most other outcomes were similar, compared to patients with MRD. Conclusion For patients with SAA and other BMF syndromes, transplanting high doses of mobilized CD34+ selected cells from MRD and MUD donors combined with a bone marrow transplant equivalent of non-mobilized donor T-cells (a partially T-cell depleted PBSC transplant) achieves rapid and durable donor engraftment and excellent survival. The cumulative incidence of both acute and chronic GVHD with this regimen was significantly lower than that historically reported with conventional PBSC transplants. Notably, low acute and chronic GVHD rates were also achieved in SAA patients with disease refractory to conventional IST who received a graft from a 10/10 HLA-matched MUD donor, although hematopoietic recovery was slightly delayed compared to recipients of a partially T-cell depleted graft from a MRD donor. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Conditioning Regimens are Associated with Distinct Patterns of Microbiota Injury in Allogeneic Hematopoietic Cell Transplantation.

The gut microbiota is subject to multiple insults in allogeneic hematopoietic cell transplantation (allo-HCT) recipients. We hypothesized that preparative conditioning regimens contribute to microbiota perturbation in allo-HCT. This was a retrospective study that evaluated the relationship between conditioning regimens exposure in 1,188 allo-HCT recipients and the gut microbiome. Stool samples collected from 20 days before transplantation up to 30 days after were profiled using 16S rRNA sequencing. Microbiota injury was quantified by changes in α-diversity. We identified distinct patterns of microbiota injury that varied by conditioning regimen. Diversity loss was graded into three levels of conditioning-associated microbiota injury (CMBI) in a multivariable model that included antibiotic exposures. High-intensity regimens, such as total body irradiation (TBI)-thiotepa-cyclophosphamide, were associated with the greatest injury (CMBI III). In contrast, the nonmyeloablative regimen fludarabine-cyclophosphamide with low-dose TBI (Flu/Cy/TBI200) had a low-grade injury (CMBI I). The risk of acute GVHD correlated with CMBI degree. Pretransplant microbial compositions were best preserved with Flu/Cy/TBI200, whereas other regimens were associated with loss of commensal bacteria and expansion of Enterococcus. Our findings support an interaction between conditioning at the regimen level and the extent of microbiota injury.

Low-Dose Total Body Irradiation Added to Fludarabine and Busulfan Reduced-Intensity Conditioning Reduces Graft Failure in Patients with Myelofibrosis

Allogeneic hematopoietic cell transplantation (alloHCT) is indicated for patients with intermediate-risk or high-risk myelofibrosis (MF) and remains the sole potential cure. Reduced-intensity conditioning (RIC) is commonly used because of older patient age, comorbidities, and a high incidence of transplantation-related mortality. Patients with MF are at increased risk of graft failure (GF), which is more common with RIC regimens, and is associated with shortened overall survival (OS). Owing to the high rate of GF with conventional fludarabine (Flu) and busulfan (Bu) RIC, we added low-dose total body irradiation (TBI; 200 cGy) for patients with MF. We retrospectively compared alloHCT outcomes in adult patients with MF who received RIC with Flu/Bu/TBI and those who received RIC with Flu/Bu. The primary endpoint was the incidence of GF. Secondary endpoints included time to engraftment, acute and chronic graft-versus-host disease (GVHD), hepatic sinusoidal obstruction syndrome (SOS), nonrelapse mortality, overall response rate, progression-free survival, and OS. Of 33 patients who underwent alloHCT, 8 received Flu/Bu RIC and 25 received Flu/Bu/TBI RIC. GF occurred in 50% of the Flu/Bu recipients (all secondary GF) and in 4% of the Flu/Bu/TBI recipients (1 case of primary GF;relative risk, .08; 95% confidence interval [CI], .01 to .62; P=.0016). GF incidence was similar with related or unrelated donors and in patients who did and did not receive Janus-associated kinase inhibitors prior to alloHCT. Molecular remission and donor chimerism ≥99% were significantly more common with Flu/Bu/TBI. No significant differences in acute GVHD, chronic GVHD, or time to engraftment were observed. SOS occurred in none of the 8 patients who received Flu/Bu and in 6 of the 25 patients who received Flu/Bu/TBI, but this difference did not reach statistical significance. Progression or relapse at 1 year was less common with Flu/Bu/TBI (0% versus 63%; P < .001). The median OS was 49 months for Flu/Bu/TBI recipients and 30.8 months for Flu/Bu recipients (hazard ratio, .98; 95% CI, .33 to 2.88; P=.97). Flu/Bu/TBI resulted in a significant reduction in GF and a significant improvement in the frequency of molecular remission and full donor chimerism compared with Flu/Bu. The addition of low-dose TBI to Flu/Bu successfully mitigates against GF in patients with MF without increased rates of complications.

Brain toxoplasmosis after HLA haploidentical transplantation for acute myeloid leukemia

A 66-year-old man with acute myeloid leukemia with a complex karyotype achieved complete remission with idarubicin + cytarabine (Arac) (3 + 7) followed by high-dose Arac for three cycles which were followed by haploidentical stem-cell transplantation with fludarabine, intravenous busulfan, and low-dose total body irradiation. Post-transplant cyclophosphamide was used for the prevention of graft-versus-host disease (GVHD). Neutrophil engraftment was observed on day 21. On day 32, the patient developed acute gastrointestinal GVHD and was started on prednisolone (1 mg/kg). GVHD gradually improved, and prednisolone was reduced, but on day 72, the patient developed pulmonary GVHD and was treated with a high-dose corticosteroid. Around the same time, he began to experience disorientation, which worsened rapidly. His respiratory condition gradually improved with immunosuppressive drugs and artificial respiration, but his disorientation persisted. Computed tomography (CT) of the brain showed no abnormalities, and magnetic resonance imaging showed multiple high-signal findings with myelography. Examination of the cerebrospinal fluid was negative for both bacteria and viruses. Because serum toxoplasma immunoglobulin M (IgM) was elevated, that is 1.4 IU/ml (range: 0.0–0.8 IU/ml), we considered the possibility of cerebral toxoplasmosis and administered sulfamethoxazole/trimethoprim; however, there was no improvement in consciousness, and the patient died on day 99. A brain CT scan taken after his death showed multiple small calcified lesions in his brain (Figure 1A), and autopsy findings showed infarct-like necrosis (Figure 1B) and Toxoplasma cysts (bradyzoites) in the bilateral basal ganglia, cerebral cortex, and subcortical areas (Figure 1C). Immunostaining by anti-Toxoplasma gondii antibody was positive (Figure 1D). Multiple cerebral calcified lesions are a typical picture of congenital cerebral toxoplasmosis, but they are rare in cases of acquired cerebral toxoplasmosis in immunosuppressed adults. Moreover, calcification in a short period, as in this patient, has rarely been reported before. The authors received no specific funding for this work. We also thank Editage (www.editage.jp) for English language editing. The authors declare they have no conflicts of interest. Ethical approval was not sought for the present study because this study is only a single case report with complete removal of all identifying information. The patient provided consent for the publication of this case with the removal of all identifying information to remain anonymous and retain his privacy. Takafumi Tsushima designed the study, collected data, wrote the manuscript, and provided patient care. Rikako Tabata and Kentaro Narita provided patient care. Koichi Honma and Kengo Takeuchi performed pathology. Kosei Matsue supervised this paper. All authors reviewed and approved the manuscript.

PO-1746 Semi-automated treatment planning for low-dose total body irradiation (TBI)

PO-1746 Semi-automated treatment planning for low-dose total body irradiation (TBI)

308 - Switching to IV Melphalan with or without Low Dose Total Body Irradiation Reduces Primary Graft Failure Rates in Children with Malignancies When Busulfan Monitorization Is Unavailable

308 - Switching to IV Melphalan with or without Low Dose Total Body Irradiation Reduces Primary Graft Failure Rates in Children with Malignancies When Busulfan Monitorization Is Unavailable

A CD45-targeted antibody-drug conjugate successfully conditions for allogeneic hematopoietic stem cell transplantation in mice

AbstractAllogeneic hematopoietic stem cell transplantation (allo-HSCT) is a potentially curative treatment of patients with nonmalignant or malignant blood disorders. Its success has been limited by graft-versus-host disease (GVHD). Current systemic nontargeted conditioning regimens mediate tissue injury and potentially incite and amplify GVHD, limiting the use of this potentially curative treatment beyond malignant disorders. Minimizing systemic nontargeted conditioning while achieving alloengraftment without global immune suppression is highly desirable. Antibody-drug-conjugates (ADCs) targeting hematopoietic cells can specifically deplete host stem and immune cells and enable alloengraftment. We report an anti-mouse CD45-targeted-ADC (CD45-ADC) that facilitates stable murine multilineage donor cell engraftment. Conditioning with CD45-ADC (3 mg/kg) was effective as a single agent in both congenic and minor-mismatch transplant models resulting in full donor chimerism comparable to lethal total body irradiation (TBI). In an MHC-disparate allo-HSCT model, pretransplant CD45-ADC (3 mg/kg) combined with low-dose TBI (150 cGy) and a short course of costimulatory blockade with anti-CD40 ligand antibody enabled 89% of recipients to achieve stable alloengraftment (mean value: 72%). When CD45-ADC was combined with pretransplant TBI (50 cGy) and posttransplant rapamycin, cyclophosphamide (Cytoxan), or a JAK inhibitor, 90% to 100% of recipients achieved stable chimerism (mean: 77%, 59%, 78%, respectively). At a higher dose (5 mg/kg), CD45-ADC as a single agent was sufficient for rapid, high-level multilineage chimerism sustained through the 22 weeks observation period. Therefore, CD45-ADC has the potential utility to confer the benefit of fully myeloablative conditioning but with substantially reduced toxicity when given as a single agent or at lower doses in conjunction with reduced-intensity conditioning.

Abstract LBA049: Possibility of total body irradiations for lymph node metastasis and lung metastasis

Abstract Metastatic lymph nodes (MLNs) are one of the most common causes of distant metastasis, and their presence signals a serious deterioration in the life prognosis of an affected patient. The treatment of MLNs at an early stage is necessary to inhibit metastases to distant LNs and organs. Radiation therapy is an effective therapeutic modality for local tumor control in many cancers, and its effect is enhanced by fractionated radiation therapy or in combination with other therapeutic regimens. In addition, recent studies have reported that total body irradiation (TBI) increases the activation of immune cells such as NK cells and infiltration of CD4+ T cells to tumor tissue. However, the effects of TBI on LN metastasis remains unclear. This study assessed the effect of TBI by changing the radiation dose and number of irradiations in an MLN mouse model. To induce metastasis into the proper axillary lymph node (PALN) and lung, mouse breast cancer FM3A-Luc cells were inoculated into the subiliac lymph node (SiLN) of MXH10/Mo/lpr mice. Mice were assigned to 4 groups thus: received no TBI; received a single dose of 0.2 Gy as low-dose TBI (single L-TBI); received a single dose of 1.0 Gy as middle-dose TBI (single M-TBI); or received two fractions of 2.0 Gy (fractionated M-TBI). The irradiation was carried out on day 4 post-inoculation in the single L-TBI and M-TBI groups. In the fractionated M-TBI group the irradiation was carried out on days 4 and 7 post-inoculation. Tumor growth was evaluated using an in vivo bioluminescence imaging system. In the non-irradiated group, tumor activity in the SiLN and PALN increased over time and lung metastasis was confirmed on day 28 post-inoculation. In addition, the single L-TBI dose did not delay tumor growth in the SiLN and PALN or inhibit lung metastasis. In other words, the tumor activity was not significantly changed compared to the non-irradiated group. Single and fractionated M-TBI doses significantly delayed tumor growth in the SiLN and PALN and inhibited lung metastasis. Furthermore, the fractionated M-TBI group exhibited the greatest delay in tumor growth in the PALN compared to the other three groups. These results suggest that the TBI at a dose &amp;gt; 1.0 Gy can delay tumor growth of MLNs in the early stage and distant metastasis. Moreover, the radiation dose and number of irradiations may be essential factors for enhancing the anti-tumor effects of TBI. We anticipate that TBI will contribute to the prevention and treatment of cancer metastasis via LNs. Citation Format: Shouta Sora, Radhika Mishra, Ariunbuyan Sukhbaatar, Maya Sakamoto, Shiro Mori, Tetsuya Kodama. Possibility of total body irradiations for lymph node metastasis and lung metastasis [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr LBA049.

Reduced Intensity Hematopoietic Cell Transplantation Improves Cerebral Hemodynamics in Children with Sickle Cell Disease

IntroductionAllogeneic hematopoietic cell transplantation (HCT) is increasingly being utilized in patients with sickle cell disease (SCD) to prevent progressive organ dysfunction, including for primary or secondary stroke prevention. However, outcomes after HCT have mostly been evaluated using broad measures such as overall survival and event free survival with limited direct physiologic evidence of improved cerebral hemodynamics. Cerebral blood flow (CBF; mL blood/100 g tissue/min) is increased in SCD to compensate for chronic anemia, reduced oxygen carrying capacity and to maintain adequate oxygen delivery to the brain tissue. In SCD patients, CBF is inversely associated with intelligence quotient (Strouse JJ, et al. Blood. 2006;108(1):379-381.) and working memory (Prussien KV, et al. Stroke. 2021;52(5):1830-1834.) and has been shown to improve with blood transfusions (Guilliams KP, et al. Blood. 2018;131(9):1012-1021.). Assessment of CBF may help predict long-term neurocognitive outcomes in patients with SCD and assess therapeutic responses to therapies. Additionally, the high metabolic demand of the brain potentially makes assessment of CBF a sensitive predictor of future multiorgan damage in patients with SCD and hence may be useful to help determine eligibility of patients for curative therapies.MethodsWe performed anatomical and hemodynamic magnetic resonance imaging (MRI) of the brain prior to, and at 6 months after HCT in children with SCD undergoing a reduced intensity conditioning based HCT on a clinical study (NCT04362293). All patients received pretransplant conditioning with hydroxyurea, azathioprine, alemtuzumab, thiotepa and low dose total body irradiation (200-400 cGy). All patients received an unmanipulated mobilized peripheral blood derived hematopoietic stem and progenitor cell graft. One patient who received the graft from a haploidentical (HAPLO) donor also received post-transplant cyclophosphamide. Graft versus host disease prophylaxis comprised of sirolimus. A 3-dimensional (3D) T1 sequence was used for gray/white matter segmentation. 3D-pulsed arterial spin labeling (ASL) perfusion imaging (3 mm isotropic) was acquired to measure resting CBF. Mean CBF values were calculated from gray matter. Mean ± standard deviation and median values for various variables are presented.ResultsFour consecutive patients had 2 serial MRIs performed (one before HCT and another 6 months after HCT). Median age at HCT was 14.5 years. Indications for HCT, donor type and whether patients were receiving chronic transfusion therapy (CTT) prior to HCT are indicated in the Table. No patient had prior evidence of overt stroke. All patients engrafted and had >99% donor myeloid chimerism at the time of the post-HCT imaging. Mean pre-HCT hemoglobin was 8.7 ± 0.8 g/dL (median 8.6 g/dL) and it increased to 12.5 ± 2.1 g/dL (median 12.4 g/dL) at 6 months after HCT. All patients exhibited elevated resting CBF values prior to HCT (123.8 ± 32.2 mL blood/100 g tissue/min, median 112.8 mL blood/100 g tissue/min), that decreased following HCT (71.1 ± 32.1 mL blood/100 g tissue/min, median 72.8 mL blood/100 g tissue/min) (Table and Figure). None of the patients had any clinical neurological complications or imaging evidence of new infarcts or anatomical abnormalities in the follow up period.ConclusionOur preliminary results indicate that it is feasible to quantify CBF in patients with SCD before and after HCT using MRI. We demonstrate that cerebral hemodynamics improve after HCT in children with SCD, indicated by a decrease in CBF to near normal values (normal CBF ~80 mL blood/100 g tissue/min). The more substantial changes (-50% and -78%) are seen in children who were not receiving CTT, suggesting that CTT might partially mitigate CBF elevation prior to HCT. Nevertheless, 2 patients receiving CTT prior to HCT also had a further modest decrease in CBF after HCT (-5% and -18%). We plan to follow these changes at serial time points annually after HCT and correlate them with changes in neurocognitive functioning and other functional MRI measures. Improved CBF is expected to reduce stroke risk and may also preserve or improve neurocognitive functioning, and prospective monitoring of these outcomes are underway.Data on all the study participants who have undergone an HCT and had 2 serial MRIs performed by the ASH annual meeting will be presented. [Display omitted] DisclosuresSharma: Vertex Pharmaceuticals/CRISPR Therapeutics: Other: Salary support paid to institution; Medexus Inc: Consultancy; CRISPR Therapeutics: Other, Research Funding; Novartis: Other: Salary support paid to institution; Spotlight Therapeutics: Consultancy; Vindico Medical Education: Honoraria. Triplett: Miltenyi: Other: Travel, meeting registration. Hankins: Vindico Medical Education: Consultancy; UpToDate: Consultancy; Global Blood Therapeutics: Consultancy; Bluebird Bio: Consultancy.