Complete Donor Research Articles

Liver Transplantation in Situs Inversus and Portal Vein Thrombosis (First Case Report in Russia)

Situs inversus is a rare mirror transposition of internal organs relative to the median plane. Liver transplantation in situs inversus is a serious medical challenge imposed by an abnormal arrangement of the recipient's organs. Aim . Clinical description of a successful cadaveric liver transplantation in an adult patient with situs inversus and portal vein thrombosis. Key points . A 32 years-old patient with situs inversus , chronic hepatitis B/D-induced liver cirrhosis and portal vein thrombosis had cadaveric liver transplantation, with the transplant reoriented in abdominal cavity 90° clockwise relative to the median plane. End-to-side cavocaval anastomosis was formed between the donor's retrohepatic and the recipient's main inferior vena cava, with successive portal and arterial anastomoses and complete donor liver reperfusion. A biliary reconstruction was performed with an end-to-end biliobiliary anastomosis. The surgery duration comprised 5 h 20 min, intraoperative blood loss — 2000 mL, cold ischemia time — 4 h. The patient was discharged on the 10 th day after surgery in satisfactory condition. Conclusion . Situs inversus is not a contraindication to liver transplantation.

DONOR HLA GENOTYPING BY NEXT GENERATION SEQUENCING OF DNA FROM URINE-DERIVED DONOR CELLS OF RENAL ALLOGRAFT RECIPIENTS

Introduction: Antibody-mediated rejection (AMR), dominantly induced by anti-HLA antibody, remains the most important cause of long-term graft loss after kidney transplantation. Donor-specific anti-HLA antibody (DSA) can’t often be determined at a specific allele level since historical donor DNA sample is not available and the existing donor HLA typing information is incomplete or even missing. Here, we developed a novel non-invasive technique by selectively expanding donor cells from urine of renal transplant recipients, and thus generating unlimited donor DNA to perform reliable allele-level HLA typing with next generation sequencing method for all HLA loci. Materials and Methods: This study enrolled 28 renal transplant recipients. The fresh midstream urine from each recipient was collected. After urine sample was centrifuged, the supernatant was discarded and washed twice with PBS. The resulting sediment was resuspended in the culture medium. The cells were plated and non-adherent cells were removed. DNA was extracted from the urine-derived donor cells (UDDCs). NGS sequencing is carried out to determine the HLA loci. Results: UDDCs from 26 patients were successfully isolated. High-resolution HLA results were unambiguously determined in 25 out of 26 DNA samples (25/26, 96%) and they are 100% concordant with donor HLA. We showed five HLA sequencing cases of UDDCs (Fig 1). The HLA typing of UDDCs is completely matched with HLA of donor peripheral blood. We further isolated UDDCs from a patient whose donor HLA information is missing at 10 years after kidney transplantation. Based on the UDDCs HLA typing and LSA results, DQA1*05:01 DQB1*02:01 were established as the culprit DSA loci of this patient (Fig 2). Discussion: DNA could be extracted from fresh kidney graft tissues for HLA genotyping; however, this approach is invasive and may aggravate graft injury. The amount of tissues obtained may be minimal and cannot yield sufficient quantity of DNA for HLA typing[1]. Other investigators extract DNA from paraffin embedded tissues or urinary sediments of KTx patients, and mismatched donor antigens could be deduced by subtracting the recipient’s HLA from the allograft HLA[2,3]. However, robust high-resolution HLA typing data can not be obtained by these methods. With an ex vivo expandable source of exfoliated cells, we provide a novel noninvasive urine-based molecular diagnostic method for donor HLA genotyping. Conclusion: The UDDCs-based HLA genotyping strategy successfully establishes complete donor DNA genotypes and provides a feasible and clinically relevant molecular diagnostic method that help to determine DSA loci for kidney transplant patients with historically insufficient donor HLA information.

Abstract 1516: Deep profiling of T-cell repertoire and tumor heterogeneity in chronic lymphocytic leukemia patients following allogeneic T-cell therapy

Abstract Chemotherapy/ targeted therapy are both known to trigger evolution of treatment resistant clones that can lead to relapse. Allogeneic stem cell transplant (alloSCT) for refractory Chronic Lymphocytic Leukemia (CLL) patients is associated with better outcomes. We hypothesized that allogeneic T-cell immunotherapies, including alloSCT and donor lymphocyte infusion (DLI) would impact tumor evolution through the application of selective immunologic pressure with reciprocal changes in the T-cell compartment. Here, we tested a cohort of 24 heavily pre-treated CLL patients treated. Treatments consisted of alloSCT alone, or with follow-up DLI, which are two established mediators of effective Graft versus Leukemia (GVL). Our cohort included 11 patients who relapsed (denoted as non-responder, NR) after alloSCT and 13 patients who had complete response (CR) after alloSCT, with 11/13 patients showing durable CR with a median post-transplant overall survival (OS) of 9.8 years. We mapped the evolutionary trajectories of tumor cells by whole exome sequencing (WES) of sort purified CLL in post-transplant relapsed patients. To investigate changes in immune repertoire and gene expression post-transplant, CD3 positive T-cells from peripheral blood and bone marrows of CLL patients at complete donor chimerism were analyzed both at bulk and at the single cell level. We found evidence of subclonal leukemic evolution in the majority of our CLL patient cohort after nonmyeloablative HLA-matched alloSCT. Different patterns of CLL evolution were observed, and these changes included putative CLL drivers in every case. In all of the 11 patients with longitudinal post-alloSCT samples available, we observed branched CLL evolution in 4 patients, linear evolution in 4 patients, and no evolution in 3 patients. These data suggest that differential sensitivity of leukemic subclones to allogeneic T cell killing may underlie the branched and linear evolution that we observed, and therefore can shape leukemic subclonal architecture after transplant. Of note, we found that clonal CLL was more responsive to alloSCT in comparison to CLL with subclonal disease architecture.To identify T-cells with GVL potential, we first cataloged potential neoantigens by screening mutated regions in CLL with in silico HLA binding prediction models. Neoantigen specific T-cells were then sorted from longitudinal peripheral blood samples using tetramers, followed by identification of GVL specific TCR in both bulk and single cell setting. We were able to identify T-cells that coevolved with specific tumorigenic lesions in a subset of CLL patients. Taken together, our results suggest that donor-derived antigen-specific T-cells mediate clonal selection of CLL with concurrent changes in allogeneic T-cells, and that these changes can be monitored in longitudinal patient samples. Citation Format: Celine Kerros, John P. Miller, Xizeng Mao, Haven R. Garber, Hannah C. Beird, Jianhua Zhang, Jason Roszik, Paul Leonard, Li Zhao, Sahil Seth, Pei Lin, Huandong Sun, William G. Wierda, Issa F. Khouri, Karen Clise-Dwyer, Andrew Futreal, Shoudan Liang, Koppikar Priya, Jeffrey Molldrem. Deep profiling of T-cell repertoire and tumor heterogeneity in chronic lymphocytic leukemia patients following allogeneic T-cell therapy [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1516.

HIV and lymphoma: from oncological futility to treatment

Cancer and HIV have been inextricably linked since the first report in 1982 of biopsy-confirmed Kaposi's sarcoma among previously healthy homosexual men living in Los Angeles and Orange counties, California, USA. 1 Centers for Disease ControlA cluster of Kaposi's sarcoma and pneumocystis carinii pneumonia among homosexual male residents of Los Angeles and Orange counties, California. MMWR Morb Mortal Wkly Rep. 1982; 31: 305-307 PubMed Google Scholar The cases established concerns for what later would be defined as AIDS. The report speculated: “One hypothesis consistent with the observations reported here is that infectious agents are being sexually transmitted among homosexually active males. Infectious agents not yet identified may cause the acquired cellular immunodeficiency that appears to underlie KS [Kaposi's sarcoma] and/or PCP [Pneumocystis carcinii pneumonia] among homosexual males.” The discovery of HIV proved it to be the aetiological agent. Given the known association between immunodeficiency and cancer and oncogenic virus and cancer, the discovery of human herpesvirus 8 (also known as Kaposi's sarcoma-associated herpesvirus) completed the puzzle on the mysterious outbreak of Kaposi's sarcoma. 2 Chang Y Cesarman E Pessin MS et al. Identification of herpesvirus-like DNA sequences in AIDS-associated Kaposi's sarcoma. Science. 1994; 266: 1865-1869 Crossref PubMed Scopus (4973) Google Scholar Epidemiology of haematological malignancies in people living with HIVPeople living with HIV or AIDS are at increased risk of Hodgkin and non-Hodgkin lymphoma compared with HIV-negative individuals. Data on the risk of multiple myeloma or leukaemia are inconsistent and of low quality but the risk does not seem to be increased. Specific haematological malignancies occur in different contexts of age, CD4 cell count, HIV control, viral co-infections, or chronic inflammation, and the expansion of combination antiretroviral therapy has led to varied demographic and epidemiological shifts among people with HIV. Full-Text PDF CAR T-cell therapy for cancer and HIV through novel approaches to HIV-associated haematological malignanciesPeople living with HIV are a global population with increased cancer risk but their access to modern immunotherapies for cancer treatment has been limited by socioeconomic factors and inadequate research to support safety and efficacy in this population. These immunotherapies include immune checkpoint inhibitors and advances in cellular immunotherapy, particularly chimeric antigen receptor (CAR) T-cell therapy. Despite the field of cancer immunotherapy rapidly expanding with ongoing clinical trials, people with HIV are often excluded from such trials. Full-Text PDF Haemopoietic cell transplantation in patients living with HIVHaemopoietic cell transplantation is established as a standard treatment approach for people living with HIV who have haematological malignancies with poor prognosis. Studies with autologous and allogeneic haemopoietic cell transplantation suggest that HIV status does not adversely affect outcomes, provided that there is adequate infection prophylaxis. Attention to possible drug–drug interactions is important. Allogeneic haemopoietic cell transplantation substantially reduces the long-term HIV reservoir when complete donor chimerism is established. Full-Text PDF Treatment management of haematological malignancies in people living with HIVAlthough the incidence of HIV-associated lymphomas decreased after the introduction of effective combination antiretroviral therapy, they became the most common AIDS-related cancer in high-income countries. Moreover, as people living with HIV live longer, a wide range of non-AIDS-related cancer has emerged, including other haematological malignancies. Nonetheless, combination antiretroviral therapy has offered people with HIV the opportunity to receive the same therapies as those provided to the general population, and intensive curative therapies have become the standard. Full-Text PDF

A Case Series of Post-Transplantation Cyclophosphamide in Unrelated Donor Hematopoietic Cell Transplantation for Aplastic Anemia.

Patients with severe aplastic anemia (SAA) who fail immunosuppressive therapy have a dismal prognosis. Hematopoietic stem cell transplantation (HSCT) from an unrelated donor (URD) is one of the most effective treatment options. Two institutions have independently adopted a post-transplantation cyclophosphamide (PTCy) approach for patients with SAA undergoing HSCT from a URD. Thirteen patients were included, 11 of whom had been treated with immunosuppressive therapy. Eight patients had a mismatched URD. All patients were conditioned with fludarabine, cyclophosphamide, and total body irradiation, in various dosage combinations. PTCy was given at a dose of 100 mg/kg. Two patients died, and overall survival was 85% at 2 years. All patients engrafted, but 1 patient developed secondary graft failure. Of the 11 patients alive after 2 years, 9 had complete donor chimerism. All surviving patients were transfusion-independent. Ten patients (77%) had cytomegalovirus reactivation, and 2 patients had more than 1 reactivation. No Epstein-Barr virus reactivation or post-transplantation lymphoproliferative disease was observed. Four patients had mild hemorrhagic cystitis. In summary, our findings show that PTCy is a promising treatment for patients with SAA undergoing URD HSCT.

Large-scale electron microscopy database for human type 1 diabetes

Autoimmune β-cell destruction leads to type 1 diabetes, but the pathophysiological mechanisms remain unclear. To help address this void, we created an open-access online repository, unprecedented in its size, composed of large-scale electron microscopy images (‘nanotomy’) of human pancreas tissue obtained from the Network for Pancreatic Organ donors with Diabetes (nPOD; www.nanotomy.org). Nanotomy allows analyses of complete donor islets with up to macromolecular resolution. Anomalies we found in type 1 diabetes included (i) an increase of ‘intermediate cells’ containing granules resembling those of exocrine zymogen and endocrine hormone secreting cells; and (ii) elevated presence of innate immune cells. These are our first results of mining the database and support recent findings that suggest that type 1 diabetes includes abnormalities in the exocrine pancreas that may induce endocrine cellular stress as a trigger for autoimmunity.

Prevention of acute graft-versus-host-disease by Withaferin a via suppression of AKT/mTOR pathway

Acute Graft versus Host Disease (aGVHD) is a frequent and serious complication in patients receiving allogeneic bone marrow transplantation (allo-BMT) and often requires rigorous prophylaxis. The current treatment regimens for aGVHD are associated with several side effects which necessitates the development of novel interventions that prevent aGVHD without precluding graft-versus-tumor effects. In the present study, we show that treatment of donor graft with plant steroidal lactone Withaferin A (WA) prior to transplantation markedly reduced aGVHD mediated damage in target organs without compromising the graft-versus.-tumor activity of the transplanted lymphocytes. WA abrogated post-transplant cytokine storm associated with allo-activation of donor lymphocytes. This was attributed to the ability of WA to inhibit early signaling events in T-cell activation including lymphoblast formation and activation of AKT/mTOR pathway. Mortality and morbidity related to allo-transplantation was significantly reduced in recipients of WA treated donor splenocytes compared to recipient of vehicle treated donor splenocytes. Further, WA treatment did not have any effect on reconstitution of lymphoid and myeloid lineages in recipients, resulting in stable and complete donor chimerism. In agreement with previous reports showing the effectiveness of WA in a mouse model of partial chimerism, our data further establishes that WA is able to attenuate aGVHD in an MHC-mismatched high dose chemo-conditioned murine model without compromising engraftment. This study provides compelling scientific basis for possible application of WA for prevention and treatment of aGVHD in patients receiving allo-BMT.

Outcome of donor lymphocyte infusion for fall in chimerism after hematopoietic stem cell transplant

Achievement of complete donor chimerism after an allogeneic hematopoietic stem cell transplant is necessary for elimination of underlying malignant disease. A decline in donor chimerism may herald an impending relapse and therefore, early recognition and intervention plays an important role in such cases. A 32 year old male patient diagnosed as a case of Philadelphia positive mixed phenotypic acute leukaemia underwent peripheral blood hematopoietic stem cell transplant (HSCT) with his sibling as donor. During follow-up, a fall in donor chimerism was observed from 91.86% on day +37 to 88.83% on day +57 and 85.34% on day +77. Donor Lymphocyte Infusion (DLI) was harvested via apheresis. A dose of 1 × 106 per kg was infused and the rest was cryopreserved in aliquots of escalating doses. On day +102, he presented with biopsy proven acute mucocutaneous GVHD grade 2 which was managed conservatively and donor chimerism of 57.99%. On day +126, a repeat donor chimerism was performed which showed 100% chimerism. He continues to do well at day +161. Timely use of DLI can improve donor chimerism in patients with Philadelphia positive acute leukaemia who tend to relapse after HSCT.

Innominate variant artery in the first web space

Numerous variations are characteristic for hand anatomy. Although a lot of work has been done in the field, a detailed description of the branches of the radial artery is still missing. The aim of this study is to determine the incidence and diameter of the accessory artery, which can be found running on the dorsal surface of the interosseus dorsalis primus muscle, to deliver the detailed description of this arterial variation and based on that to suggest a systematic name of the artery which would be in line with Terminologia Anatomica. We used 133 complete donor bodies and 237 cadaverous hands in our study, giving us a total sample size of 503 samples. When possible, we determined the age of the donors which was between 62 and 90 years. We performed detailed anatomical dissection to determine the individual branching. We also measured the diameter of selected arteries. When relevant we performed statistical comparisons. To do that we first applied Shapiro-Wilk test to determine the normality of distribution and after that we used Mann-Whitney U test and One-way ANOVA. The variation of interest was found in 11.93% of cases. Based on the anatomical differences we determined four types of branching, type 1-3 forming the anastomosis with superficial palmar arch (being considered as positive findings) and type 4 which did not form this anastomosis (thus considered to be a negative finding). We successfully determined the incidence of this arterial variation on a sufficient sample size. We also described the anatomy of this branching in detail and were able to determine four types of this branching. After careful consideration of these findings we proposed the new name for this artery and suggest to use the name superficial dorsal branch of radial artery. This could contribute to a better understanding of this branching and potential use in clinical practice.

Failure to Prevent Severe Graft-Versus-Host Disease in Haploidentical Hematopoietic Cell Transplantation with Post-Transplant Cyclophosphamide in Chronic Granulomatous Disease.

Haploidentical related donor (HRD) transplantation was performed in 7 recipients with chronic granulomatous disease (CGD) who had no matched-related or unrelated donor. Peripheral blood cell (PBC) products were used with a conditioning regimen consisting of low-dose cyclophosphamide, fludarabine, total body irradiation, and busulfan. Graft-versus-host disease (GVHD) prophylaxis consisted of high-dose post-transplant cyclophosphamide and sirolimus. Recipients were ages 14-26years, and 3 had severe infections active at transplant. All 7 recipients achieved full engraftment with complete donor chimerism early in the post-transplant period. Acute GVHD occurred in all cases and was grade 3 or steroid refractory in 3. Two patients with steroid-refractory GVHD died. Three patients with severe infectious complications active at transplant, 1 Nocardia pneumonia and 2 extensive invasive fungal infections), survived and were cured of their infection at last follow-up. Bacterial disease occurred post-transplant in all recipients, and viral infections/reactivation were common, including 4 cases of BK virus-associated hemorrhagic cystitis. Seven patients with CGD achieved rapid and full-donor engraftment from HRDs utilizing PBCs and a conditioning regimen with PTCy and sirolimus GVHD prophylaxis. However, the incidence of grade 3 and steroid-refractory GVHD was high and led to 2 deaths. Patients with active infections at transplant had successful transplant courses and were cured of their disease. Although there was an initial success with this regimen, the cumulative experience does not support its use in CGD due to an unacceptable rate of severe GVHD.

Treosulfan\u2013fludarabine\u2013thiotepa-based conditioning treatment before allogeneic hematopoietic stem cell transplantation for pediatric patients with hematological malignancies

Treosulfan-based conditioning prior to allogeneic transplantation has been shown to have myeloablative, immunosuppressive, and antineoplastic effects associated with reduced non-relapse mortality (NRM) in adults. Therefore, we prospectively evaluated the safety and efficacy of treosulfan-based conditioning in children with hematological malignancies in this phase II trial. Overall, 65 children with acute lymphoblastic leukemia (35.4%), acute myeloid leukemia (44.6%), myelodysplastic syndrome (15.4%), or juvenile myelomonocytic leukemia (4.6%) received treosulfan intravenously at a dose of 10 mg/m2/day (7.7%), 12 g/m2/day (35.4%), or 14 g/m2/day (56.9%) according to their individual body surface area in combination with fludarabine and thiotepa. The incidence of complete donor chimerism at day +28 was 98.4% with no primary and only one secondary graft failure. At 36 months, NRM was only 3.1%, while relapse incidence was 21.7%, and overall survival was 83.0%. The cumulative incidence of acute graft-vs.-host disease was 45.3% for grades I–IV and 26.6% for grades II–IV. At 36 months, 25.8% overall and 19.4% moderate/severe chronic graft-vs.-host disease were reported. These data confirm the safe and effective use of treosulfan-based conditioning in pediatric patients with hematological malignancies. Therefore, treosulfan/fludarabine/thiotepa can be recommended for myeloablative conditioning in children with hematological malignancies.

Race Does Not Predict Pancreas Graft Failure after Pancreas Transplantation in Modern Era

Background: Race has been proposed to be associated with graft survival after pancreas transplantation (PTx). African-American (AA) recipients, as well as those with AA donors, always experience worse graft survival than other racial groups. We hypothesize that the racial disparity may disappear in modern times. Methods: We analyzed data from the Scientific Registry of Transplant Recipients (January 1989 - August 2018). Using Kaplan-Meier estimation and Cox proportional hazards regression, we examined the impact of race on pancreatic graft survival after PTx. Findings: A total of 28797 pancreatic recipients were included in final analysis, including 22430 Whites, 3828 AAs, 2235 Hispanics, and 304 Asians, and 71.6% of donors came from Whites, 15.1% from AAs, 11.8% from Hispanics, and 1.6% from Asians. After 3 years of follow-up evaluation, there were 6903 pancreatic grafts failed (PGF). Before 2009, AA recipients (hazard ratio [HR], 1.34; 95% confidence interval [CI], 1.10-1.64) were associated with PGF after adjusting confounding factors, and risk of PGF for White and Asian recipients were both comparable with Hispanic counterparts. However, the risk of PGF in AA recipients decreased to 2% and was no longer significant since 2009 (HR, 1.02; 95% CI, 0.77-1.34). Interestingly, donor race had the similar results as recipient race. Furthermore, the concordance statistic of the complete pancreas donor risk index (PDRI) (including donor race) was 0.582, whereas the concordance did not change when the donor race was eliminated from the PDRI model (0.580). Interpretation: With the potent immunosuppressive regimen and improved surgical techniques in recent era, AA and other racial recipients had similar pancreatic graft outcomes. Furthermore, racial disparity of donor also could be neutralized, so donor race should not be considered as an indicator of donor quality. Funding: Natural Science Foundation of Zhejiang, National Natural Science Foundation, Scientific Research Project of Education of Zhejiang. Ethical Approval Statement: The whole study was reviewed and approved by the ethics committee at Zhejiang University, China. Competing Interest Declaration: The authors of this manuscript have no conflicts of interest to disclose.

Successful umbilical cord blood transplantation in children with leukocyte adhesion deficiency type I.

This study aims to investigate the efficacy and safety of umbilical cord blood transplantation (UCBT) without serotherapy for treating children with leukocyte adhesion deficiency type I (LAD-I). Clinical characteristics and data of five children with LAD-I who underwent UCBT at our hospital between September 2016 and September 2018 were retrospectively analyzed. Five (two boys and three girls) patients with LAD-I were included. The median age at UCBT was 9 months (range, 8 to 32 months). The same myeloablative conditioning regimen was administered for each patient and included busulfan, fludarabine, and cyclophosphamide. HLA matching of patients and umbilical cord blood was 8/10 to 10/10. The median dose of total nucleated cells (TNC) infused was 10.2×107/kg (range, 4.5×107 to 20.6×107/kg) and the median dose of CD34+ cells was 3.2×105/kg (range, 1.9×105 to 5.7×105/kg). The median time of neutrophil engraftment was 20 days (range, 13 to 28 days). The median time of platelet engraftment was 36 days (range, 32 to 56 days). All patients received complete donor chimerism (CDC). Four of the five patients developed grade II-IV acute graft-versus-host disease (GvHD). The median follow-up time after transplantation was 19 months (range, 8 to 38 months). Four of the patients survived and achieved complete clinical remission. The other patient died of bronchiolitis obliterans 8 months after UCBT. UCBT is an effective treatment method for LAD-I patients. Also, severe LAD-I patients should undergo stem cell transplantation as early as possible.

Predictors of Relapse after Haploidentical Hematopoietic Progenitor Cell Transplantation (Haplo-HCT); A Single-Institution Experience

<h3>Background</h3> Haplo-HCT emerged as a reliable option for patients with high risk hematological malignancies with no HLA-matched related or unrelated donors available. Retrospective registry data suggest comparable outcomes to HLA-matched donors; however, relatively little is known regarding predictors of disease relapse following haplo-HCT. We hypothesize that attainment of full-donor CD3 and CD33 chimerism at day 30 (D30) and 100 (D100) is associated with lower incidence of relapse after haplo-HCT. <h3>Methods</h3> We undertook a retrospective analysis of 78 patients who underwent haplo-HCT at the Ohio State University James Cancer Hospital between January 2013 and December 2017. The associations between patient characteristics including CD3/CD33 donor chimerisms and the cumulative incidence rate (CIR) of relapse were evaluated using proportional sub-distribution hazards model, treating death without relapse as the competing risk. The models were built adopting a landmark analysis approach at D30 and D100. Progression-free survival (PFS) was estimated using the method of Kaplan-Meier. <h3>Results</h3> Median age at haplo-HCT was 56 (20-74) years and 68% were males. 50% had acute myeloid leukemia/myelodysplastic syndrome, and 54% had intermediate disease risk index (DRI). Most patients (85%) received reduced-intensity conditioning and 50% received bone marrow (BM) grafts. Complete (100%) CD3 donor chimerism was observed in 63 (84%) patients at D30, and subsequently 59 (95%) patients at D100. Complete CD33 donor chimerism was observed in 67 (89%) patients at D30, and subsequently 56 (88%) patients at D100. In univariable D30 landmark analysis, the CIR of relapse was significantly lower in patients who achieved complete CD3 (and CD33) chimerism compared with patients with less than complete chimerism (Figure 1A; p<0.01 and p<0.01, respectively). Attainment of complete CD3 chimerism at D30 was also associated with PFS (Figure 1B; p<0.01). However, in multivariable analysis, complete CD3 chimerism at D30 was no longer independently associated with the CIR of relapse due to its high correlation with DRI (p=0.01). Receipt of a BM graft was associated with significantly higher CIR of relapse both in univariable and multivariable analyses (HR 2.43, 3.34; p=0.05, 0.01, respectively). D100 complete CD33 chimerism conferred a significant CIR advantage in both univariable (p<0.01) and multivariable (p<0.01) models, although CD3 chimerism did not retain significance (p=0.35). <h3>Conclusions</h3> Complete CD3 and CD33 donor chimerisms at D30 and D100 predict a patient's risk for relapse following haplo-HCT. We observed a strong association between CD3 chimeric status at D30 with DRI, as well suggestive that high risk disease may be associated with impaired immune reconstitution after transplant and perhaps a less robust graft vs disease effect.

A Novel Targeted Approach to Achieve Immune System Reset: CD45-Targeted Antibody Drug Conjugates Enable Autologous HSCT and Ameliorate Disease in Preclinical Autoimmune Disease Models

Resetting the immune system through autologous hematopoietic stem cell transplant (autoHSCT) is a highly effective treatment in patients with autoimmune diseases (AID). AutoHSCT achieved long-term remission in patients with relapsed refractory and secondary progressive multiple sclerosis (Muraro 2017), superior to their previous standard of care (Burt 2019). AutoHSCT in scleroderma patients achieved superior outcomes in two randomized studies (Tyndall 2014, Sullivan 2018). These impressive results are achieved by both eradication of autoreactive immune effector cells and re-establishment of a self-tolerant immune system, i.e., immune system reset. However, only a fraction of eligible patients undergo autoHSCT, in part due to toxicity associated with current conditioning regimens that remove the disease-causing cells. To enable more patients to benefit from immune reset without debilitating chemotherapy, we generated novel anti-human CD45 ADCs that cross react with NHP and evaluated these for the ability to deplete hematopoietic cells in vitro and in vivo (Fig.1). In vitro the CD45-ADC efficiently killed human CD34+ progenitors and peripheral CD3+ T cells from both healthy donors and MS patients. In vivo in humanized NSG mice, single doses of the CD45-ADCs were well-tolerated and led to depletion of human hematopoietic cells in BM (Fig. 1A) and periphery. In NHPs, single doses of CD45-ADCs were well tolerated and depleted BM HSCs and peripheral lymphocytes (Fig.1B). CD45-ADC treatment of hNSGs with sclerodermatous xenoGVHD resulted in resolution of symptoms (Fig. 1C). To model the complete immune reset approach in mouse models of AID, we generated a tool anti-mouse CD45 ADC. A single-dose enabled full myeloablation (>99% depletion of LT-HSCs) and complete donor chimerism with congenic HSCT (>90% chimerism at 16 weeks). In an adoptive transfer model of type I diabetes, treatment with a single dose of CD45-ADC and congenic HSCT led to disease prevention. In a murine model of MS EAE, a single dose of the CD45-ADC followed by congenic HSCT enabled full donor chimerism; treatment prior to disease onset significantly delayed disease onset and reduced disease severity; treatment after disease onset also halted progression of symptoms. These data demonstrate that CD45-ADC conditioning followed by congenic HSCT is sufficient for full myeloablation and immune reset. Experiments are ongoing, and evaluation of this ADC in murine models of diabetes and arthritis will be presented. These results indicate that targeted immune depletion with a single dose of CD45-ADC may be sufficient to enable auto-HSCT and immune reset in multiple AID indications. Targeted conditioning with CD45-ADC may enable more patients to benefit from immune reset through removal of pathogenic cells and autoHSCT without the morbidity and mortality associated with current chemotherapeutic conditioning.

T-Cell Replete Haploidentical Bone Marrow Transplantation and Post-Transplant Cyclophosphamide for Patients with Inborn Errors

Introduction Matched donor hematopoietic stem cell transplantation (HSCT) cures most patients with inborn hematopoietic disorders, but haploidentical HSCT was historically associated with graft rejection, GVHD and infections. In leukemia, in vivo depletion of alloreactive T-cells with post-transplant cyclophosphamide (pTCy) results in outcomes comparable to those with matched related or unrelated donors. Objectives To assess safety and efficacy of haploidentical HSCT with pTCy in patients with inborn errors of hematopoiesis who lack a suitable matched donor or have failed a previous matched donor HSCT. Methods Nineteen patients (10 primary immunodeficiencies, 7 hemoglobinopathies, 1 congenital erythropoietic porphyria, 1 bone marrow failure) with a median age of 9 years (0-22) were treated prospectively on an institutional protocol with a conditioning regimen of alemtuzumab 0.4mg/kg, rituximab 375mg/m², fludarabine 150mg/m², treosulfan 42g/m², thiotepa 10mg/kg and cyclophosphamide 29mg/kg followed by haploidentical bone marrow transplantation, pTCy and tacrolimus/MMF. In four patients who were in bone marrow aplasia after non-engraftment of a first HSCT, thiotepa was omitted and treosulfan reduced to 24g/m². Results After a median follow-up of 42 months (3-87), overall survival is 95% (18/19). One patient died on day +7 from suspected fludarabine neurotoxicity. Two patients experienced autologous reconstitution, one of whom had a successful second HSCT. Two patients have stable mixed chimerism predominantly of donor origin and remain disease free. All others exhibit complete donor chimerism. This results in event free survival of 84% (16/19) and disease free survival of 89% (17/19). Engraftment occurred after 17days (6-61) for neutrophils and 28 days (14-191) for platelets. The incidences of acute GVHD II-IV° and of chronic GVHD were 0%, respectively. Two patients (11%) developed GVHD I° of the skin.Viral reactivations were observed in 53% (10/19) of patients (8 CMV, 1ADV, 1 VZV), but only one patient developed overt disease (ADV). Immune reconstitution was swift with mean CD3+ and CD19+ counts of 793/µl and 271/µl on day +100. Other complications were bacterial bloodstream infections in 5 patients (26%) and moderate severity VOD in one (5%). No renal or hepatic toxicities greater than CTCAE grade 2 were observed. No patient developed hemorrhagic cystitis. Four patients had anti-donor HLA antibodies pre-HSCT, three of whom were treated with plasmapheresis; all engrafted. Conclusion Chemotherapy based conditioning and haploidentical HSCT with pTCy was safe and resulted in good disease free survival in this cohort of patients with inborn errors in the absence of relevant GHVD. These results compare well with outcomes achieved with matched donors in similar cohorts of patients, but larger multi-center studies are warranted to further evaluate this approach.

Haploidentical Transplant for Myelofibrosis: A Single Institution Experience

Background Myelofibrosis (MF) is a chronic hematologic malignancy with poor prognosis for which the only curative therapy is allogeneic stem cell transplant (alloHSCT). It is suggested that eligible patients with Intermediate-2 and high-risk disease proceed to transplant. Frequency of transplant for MF is increasing, given the advent of reduced intensity conditioning regimens and earlier identification of high-risk disease with mutation testing. Patients with MF are at high risk of regimen related toxicities and poor transplant outcomes due to increased risk of prolonged engraftment and graft failure. These concerns may contribute to a greater reluctance to embrace alternative donor source allografts such as haploidentical (haplo) donors. Herein, we report a case series of patients with MF undergoing alloHSCT. Methods A retrospective analysis was performed on all patients having received a haploidentical HSCT at our institution. Results Three patients were identified as having received haplo HSCT for MF. All received a bone marrow graft. Median age was 57 (range 46-61). One patient had primary MF, one post-PV MF, and one post-ET MF. Risk category at time of transplant by DIPSS was Intermediate -2 in two patients, and high in one. Two of the three patients had a major ABO mismatch. All patients and donors were positive for CMV IgG. Mean cell dose was 2.07 × 10^6 CD34+/kg recipient body weight (range 1.65 × 10^6 to 3.19 × 10^6). All patients received busulfan/fludarabine/melphalan conditioning (one myeloablative dosing, two reduced intensity). Post-transplant cyclophosphamide on days +3 and +4, tacrolimus, and mycophenolate mofetil were used for GvHD prophylaxis in all cases. Median time to neutrophil engraftment was Day +22 (range 20-24), median time to platelet engraftment was 67 days (range 37 - > 70). All patients had complete donor chimerisms at Day +30. Recovery of marrow cellularity was delayed in all patients. All patients had CMV reactivation by PCR, one patient had CMV esophagitis. Infectious complications were common, including BK virus, C. difficile colitis, invasive pulmonary aspergillosis, and klebsiella bacteremia Two patients developed chronic GvHD; one with cGVH of eyes & mouth not requiring systemic therapy, one with cGVH of upper GI tract/mouth. All patients are currently off immunosuppression. All patients are alive at a median follow up of 1.5 years post-transplant (range 456-656 days). Discussion Our case series using a haploidentical donor source and busulfan/fludarabine/melphalan conditioning along with post-transplant cyclophosphamide for patients with MF demonstrates outcomes at least comparable to conventional donor source transplants for MF. While infectious complications and slightly prolonged engraftment were noted in the early transplant period, these can be overcome with aggressive supportive care.

Evaluation of y-str specific quantitative pcr after sex mismatch hsct

Evaluation of ideal sample for identification and essential to evaluate successful engraftment after hematopoietic stem cell transplantation needs a sensitive and accurate method. A successful HSCT results in complete donor derived cells in patient's hematopoiesis. The main aim of chimerism analysis is to check the concentration of donor cells which is considered as useful method for the success of treatment. The present study was carried out to determine the chimeric status of patients who underwent hematopoietic stem cell transplantation involving female patient and male donor using commercially available Y-DNA based quantification kit. This study enrolled fourteen female patients of HSCT. Samples from patient and donor were collected before transplant as reference sample. Post-transplant samples were collected from female patients after 30 days, 60 days, and 90 days of time interval. Chimeric status of the patients was analyzed using Y- DNA quantitation.

Organization and development of a liver transplant program in Mar del Plata. Initial Results (2010-2018)

Background: Over the past 30 years, new liver transplant programs have emerged as a response to the increasing use of this therapy to treat irreversible liver failure. Objective: the aim of this presentation is to describe the organization, development and initial results of a Liver Transplant Program in Mar del Plata. Material and methods: A team of professionals trained in liver transplantation was created, with specialists in surgery, anesthesiology, intensive care, hepatology, hemotherapy, hematology, and registered nurses and nurse scrubs granted by regulatory agencies. Building alterations, and technical and adaptations were implemented. Thirty-two transplantations were performed with complete cadaveric donor in patients from neighboring localities (55%) and from Mar del Plata (45%). The most common etiologies of cirrhosis were chronic hepatitis C virus infection and alcohol abuse. Results: Operative mortality was 12.5%. Actuarial survival at 55 months of the 32 recipients was 75%. Six bile drug strictures late postoperative period which were treated successfully trated with a conservative approach Conclusions: the Liver Transplant Program in Mar del Plata provides care to the geographic area of the southeast of the province of Buenos Aires. The initial results are consistent with those expected for the current standards in liver transplantation.

Simultaneous Monitoring of Mutation and Chimerism Using Next-Generation Sequencing in Myelodysplastic Syndrome.

Monitoring minimal residual disease (MRD) provides important information during treatment of hematologic malignancies. Chimerism analysis also provides key information after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Recent advances in next-generation sequencing (NGS) have enabled identification of various mutations and quantification of mutant allele burden. In this study, we developed a new analytic algorithm to monitor chimerism applicable to NGS multi-gene panel in use to identify mutations of myelodysplastic syndrome (MDS). We enrolled patients who were diagnosed with MDS and received allo-HSCT and their corresponding donors. Monitoring MRD by NGS assay was performed using 53 DNA samples by calculating mutant allele burden after treatment. For monitoring chimerism by NGS, we selected 121 single nucleotide polymorphisms (SNPs) after careful stepwise evaluation and calculated average donor allele burden. Data obtained from NGS were compared with bone marrow findings, chromosome analysis and short tandem repeat (STR)-based chimerism. SNP-based NGS chimerism analysis was accurate and even superior to conventional STR method by overcoming the various technical limitations of STR. In addition, simultaneous monitoring of mutation and chimerism using NGS could implement comprehensive pre- and post-HSCT monitoring of various clinical conditions such as complete donor chimerism, persistent mixed chimerism, early relapse, and even donor cell-derived diseases.