Hematopoietic Stem Cell Recipients Research Articles

The Impact of Foundation of the National Turkish Donor Registry, Turkok on Donor Matching Kinetics

Purpose: Turkish Stem Cell Coordination Center (TURKOK) was established by the Turkish Ministry of Health in April 2015 in order to facilitate unrelated hematopoietic stem cell (HSC) transplantation in Turkey. The donor recruitment task has been linked to the Turkish Crescent's (National Blood Product Provider) nationwide distributed branches. As of July 2017, more than 300.000 volunteer donors are registered into the database. As of July 2017 they have facilitated more than 300 HSCTs. Transplant Registry Istanbul (TRIS) and Transplant Registry Ankara (TRAN) are two University affiliated donor search - co-ordination centers, which are officially responsible for the search, matching and organization of the delivery for unrelated donor HSC in Turkey. The foundation of TURKOK should have theoretically decreased our time spent to find a suitable matched donor. The aim of this study is to compare the time frames between admission, confirmation, work-up, and infusion of HSC between TURKOK and other WMDA or NMDP facilitated International Volunteer Donor Registries.Subjects & Methods: In 2015 1000 allogeneic HSCTs were peformed in Turkey and only 18% were from MUDs. This is a single center analysis of a dedicated active HSCT center, which is performing >70 allogeneic HSCTs per year and utilizing alternative donor sources >50%. Patient charts were reviewed retrospectively and 51 patients who admitted to our center within 2015-2017 for various high risk hematological malignancies (mostşy acute leukemia, in which allogeneic HSCT was indicated, were identified for this study. Patients were divided into two groups: (1) Patients who had received HSC from matched unrelated donors (MUD-T) (10/10-9/10) facilitated by TURKOK (N=36) ; (2) 15 patients who had received HSC from International Registries facilitated by TRIS or TRAN (MUD-I) . The duration between application and finding a suitable match (tAM), duration between matching and confirmation (tMC) and time from application to HSC transplantation (tATX) were calculated for each of these two groups. The time duration between admission and donor identification and admission to HSCT were not distributed normally. The mean values were compared between two groups by Mann-Whitney U test. p < 0.05 was considered statistically significant.Results: In 51 HSC recipients the overall median durations for tAM, tMC and tATX were 71 (range, 1-172), 33 (range, 1-181) and 97 (range, 13-172), respectively. In MUD-I overall median durations for tAM, tMC and tATX were 90 (range, 1-170), 40.5 (range, 1-181) and 98 (range, 13-172), whereas in MUD-T the timelines for tAM, tMC and tATX were 37 (range, 4-172), 17 (range, 6-151) and 96 (range, 73-128), respectively. The p values for comparative analysis betwen MUD-I and MUD-T for tAM, tMC and tATX were p=0.005, p=0.007 and p>0.05, respectively.Conclusion: In this relative limited size of HSC recipients from MUDs we observed accelerated kinetics for tAM and tMC, which means having a National Registry have a positive impact on finding an available match and confirmation process. On the other hand this improvement did not translate into rapid HSCT. The tATX between MUD-I and MUD-T are not significant. TURKOK has to establish and improve the organization of confirmed donor for collection of HSCs. The decreased timeline from application to confirmation is encouraging, but the time spent from confirmation to work/up and collection are still high. As TURKOK is a recently established donor bank, there is room for improvement. DisclosuresNo relevant conflicts of interest to declare.

The Application of ‘Drug-Reversible’ CAR-T Cells Directed Against Recipient Hematopoietic Cells As a Selective Conditioning Strategy for Stem Cell Transplantation

The transplant of autologous or allogeneic hematopoietic stem cells (HSCs) has the proven ability to treat a wide array of malignant and non-malignant hematological diseases. The preparative regimen, however, routinely entails aggressive and genotoxic treatment with total body irradiation and/or chemotherapy, which brings severe and even life-threatening complications that limit its broader application. Previous experimental studies have established that depletion of recipient HSCs is an essential requirement of these conditioning regimens in allowing successful engraftment of the composite donor HSCs. Animal and clinical studies have also indicated that alloreactive anti-HSC donor T cells additionally facilitate stem cell engraftment, but this is often accompanied by the risks of GvHD. This has prompted the consideration of alternative conditioning methods for the depletion of HSCs with less toxic side-effects, such as anti-c-kit and anti-CD45 antibody-directed treatments. In this way, more precise HSC targeting may also be achieved by the application of short-lived, genetically engineered chimeric antigen receptor (CAR)-T cells for stem cell transplantation conditioning.We developed a novel and controllable CAR-T approach for recipient HSC targeting via genetic modification using the non-viral piggyBac™ (PB) transposon system. As opposed to viral vector delivery systems, the relatively large carrying capacity of PB allows the stable introduction of at least three separate genes encoded within the same tri-cistronic transgene cassette. This includes a second-generation CAR that targets either human c-kit (CD117) or prominin-1 (CD133), markers known to be antigenically expressed on the surface of HSCs. In addition, a drug resistance element serves as a selection gene that, in combination with a non-genotoxic drug, provides an effective method of CAR-T cell purification during manufacture. Importantly, a small molecule drug-inducible safety switch gene is also included to facilitate rapid in vivo clearance of the CAR-T cells after depletion of recipient HSCs and prior to donor HSC transplant. Lastly, as a result of the manufacturing process, the majority of the CAR-T cells express chemokine receptors such as CXCR4 that can allow more selective trafficking to the bone marrow (BM) for eradication of resident HSCs.To select a lead candidate from a panel of anti-HSC CAR constructs, CD3/CD28 stimulated T cells from human peripheral blood were first electroporated with mRNA encoding each of the CAR candidates directed against either c-kit or CD133. CAR surface expression was confirmed in transfected T cells by flow cytometry. In vitro functional assays were performed by co-culturing mRNA-transfected CAR-T cells with mouse or human cell lines (EML-C1, TF-1 and K562), expressing either c-kit or CD133, as well as mouse and human primary BM cells. Lead CAR candidates were identified from their specific activation of the CAR-T cells through degranulation according to CD107a expression and secretion of IFNγ. Furthermore, those CARs were also capable of selectively depleting c-kit or CD133 positive cells. Interestingly, some mRNA-transfected CAR-T cells retained effector activity against target c-kit+ TF-1 cells even in the presence of its soluble ligand, stem cell factor. Next, lead CAR candidates were co-expressed with the selection and drug-inducible safety switch genes in the same tri-cistronic transgene and then stably delivered to T cells using PB. The manufacturing process yielded CAR-T cells that were mainly of the T memory stem cell (Tscm) phenotype, as determined by positive expression of CD62L and CD45RA, and also expressed high levels of the CXCR4 chemokine receptor. Similar to the mRNA-transfected CAR-T cells, these stably-transposed cells were capable of extensive effector capabilities including specific depletion of c-kit or CD133 expressing target cells.Future studies will evaluate PB-produced lead anti-HSC CAR-T cells in immune-deficient NSG mice with pre-established xenogeneic human hematopoietic chimerism, along with standard busulfan or radiation conditioning controls. This approach constitutes a novel targeted biological therapy that is envisaged to lead the way towards minimally toxic transplant regimens for depletion of endogenous HSCs in the BM and to procure their replacement with engrafted allogeneic or gene-corrected stem cells. DisclosuresTimberlake:Poseida Therapeutics, Inc.: Employment. Richter:Poseida Therapeutics, Inc.: Employment. Wang:Poseida Therapeutics, Inc.: Employment. Cam:Poseida Therapeutics, Inc.: Employment. Tan:Poseida Therapeutics, Inc.: Employment. Ostertag:Poseida Therapeutics, Inc.: Employment, Equity Ownership. Shedlock:Poseida Therapeutics, Inc.: Employment. Down:Poseida Therapeutics, Inc.: Employment.

Differences of Human Cytomegalovirus Strains in Recipients of Allogeneic Hematopoietic Stem Cells before and after Transplantation

Background. Human cytomegalovirus (HCMV) is a ubiquitous viral infection of worldwide distribution. This virus is the most significant infectious cause of congenital disease, an important opportunist in the immunocompromised hosts, such as allogeneic hematopoietic stem cells transplantation (allo-HSCT) recipients (Mokarski E.S.,2013). If seropositive recipient develops HCMV infection after allo-HSCT, it is usually unknown whether it is a reactivation of endogenous HCMV or a reinfection by a new exogenous HCMV strain. The impact of genetic diversity of HCMV on its pathogenesis has not been well evaluated by now. However, the emerging data indicate a possible correlation: the viral load and the course of the infection were different in AIDS patients with different HCMV genotypes (Renzette, 2015). The estimation of a possible link between viral genotype and probability of HCMV infection occurrence is relevant. Very little data on difference in HCMV genotypes within one patient before and after allo-HSCT is available.The aim. The aim of this study was the identification of difference in HCMV UL139 gene sequences before and after the allo-HSCT.Materials and Methods. Patients: 13 allo-HSCT recipients (5 ALL, 3 AML, 3 CML, 1 AA, 1 lymphoma of the mantle) were enrolled in the study, male/female 6/7; median age was 27 (22-49) years. Patients' blood samples were collected within 6 month before and 6 month after transplantation and tested for the HCMV DNA by PCR. Positive samples were sequenced by Sanger inside the UL139 (putative membrane gp gene) region for genotyping. Sequences within one patient before and after transplantation were compared by BLAST (Basic Local Alignment Search Tool) software.Results and discussion. Sequencing was successfully completed only in samples of 7 patients (pts), probably due to low concentration of the viral DNA in the samples or other technical issues. The comparison of sequences showed (see table) that 3 out of 7 pts had reactivation of endogenous virus, because sequences exhibited 100% match. Another 2 out of 7 patients' sequences showed about 88% match. This indicates change in genotype to phylogenetically closest one. The rest 2 pts did not show any similarity in sequences at all which definitely points to reinfection after allo-HSCT. Data obtained are similar to a recent study by Zawilinska (Zawilinska B, 2016), which appeals that reinfection occurs slightly more often than reactivation. Data analys showed that reactivation was detected in patients with AML, reinfection in patients with ALL. Average period before reactivation occurrence was 27 days and before reinfection occurrence - 111 days. Statistical reliability cannot be obtained due to patients' number paucity. The genotyping of patients' HCMV by comparison of sequences with known genotypes of the UL139 gene from GenBank® revealed that before transplantation HCMV genotype in 3 patients had a strong resemblance to genotype 2, another 2 patients had genotype close to the genotype 5, and 1 patient had genotype 1. After transplantation genotype 2 became dominant and was identified in 6 out of 7 patients (see table).Conclusions. The conducted study shows that HCMV infection after allo-HSCT may be the result of both reactivation of endogenous virus and reinfection by exogenous strain. Reactivation was detected in patients with AML only, reinfection - in patients with ALL only. Reactivation developed earlier than reinfection (27 days versus 111 days). The chosen method allowed us to detect several mononucleotide substitutions in sequences, what can be useful in cases when no evident change of genotype happened after allo-HSCT. There are several possible explanations of the occurrence of reinfection: possible transmission from the outer environment or reactivation of a minor genotype if 2 or more HCMV genotypes originally had taken place. Further studies are required for more reliable conclusions. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Vaccination against SARS-COV‑2 in oncology

The objective of the study a detailed, systematic review of the world literature data, which includes all aspects of recommendations for vaccination against SARS-COV‑2 in cancer patients.Materials and methods. Information search was carried out in PubMed, MedLine, Scopus, Web of Science, RSCI.The work included data from literature and information sources that were published before February 2021.Results. The data of retrospective and prospective clinical studies are analyzed. This paper reflects considerations and recommendations for the vaccination of cancer patients by Russian and foreign specialists in the context of COVID‑19 pandemic. The review presents current recommendations for vaccination against SARS-COV‑2 in patients with solid tumors, hematological malignancies, recipients of hematopoietic stem cells and cell therapy.Conclusion. To date, groups at increased risk of infection with the new coronavirus have been identified. These groups include patients with cancer. The presence of tumor does not allow a delay in start of therapy, and requires careful monitoring and observation. In this regard, despite the pandemic, the treatment of cancer patients must be continued regardless of the circumstances. Cancer patients should not be deprived of the opportunity to be vaccinated against SARS-COV‑2. Every patient should be decided individually. At the moment, there are no officially approved recommendations for vaccination against SARS-COV‑2 for cancer patients. Before the creation and approval of final recommendations for cancer patients, it is necessary to focus on compliance with sanitary and anti-epidemic measures and the prevention of COVID‑19 infection. The global cancer community continues to actively develop recommendations for the optimal vaccination against SARS-COV‑2 in cancer patients.The most relevant ones are outlined in this article.

Using mitochondrial activity to select for potent human hematopoietic stem cells

Hematopoietic cell transplantation is a critical curative approach for many blood disorders. However, obtaining grafts with sufficient numbers of hematopoietic stem cells (HSCs) that maintain long-term engraftment remains challenging; this is due partly to metabolic modulations that restrict the potency of HSCs outside of their native environment. To address this, we focused on mitochondria. We found that human HSCs are heterogeneous in their mitochondrial activity as measured by mitochondrial membrane potential (MMP) even within the highly purified CD34+CD38−CD45RA−CD90+CD49f+ HSC population. We further found that the most potent HSCs exhibit the lowest mitochondrial activity in the population. We showed that the frequency of long-term culture initiating cells in MMP-low is significantly greater than in MMP-high CD34+CD38−CD45RA−CD90+ (CD90+) HSCs. Notably, these 2 populations were distinct in their long-term repopulating capacity when transplanted into immunodeficient mice. The level of chimerism 7 months posttransplantation was >50-fold higher in the blood of MMP-low relative to MMP-high CD90+ HSC recipients. Although more than 90% of both HSC subsets were in G0, MMP-low CD90+ HSCs exhibited delayed cell-cycle priming profile relative to MMP-high HSCs. These functional differences were associated with distinct mitochondrial morphology; MMP-low in contrast to MMP-high HSCs contained fragmented mitochondria. Our findings suggest that the lowest MMP level selects for the most potent, likely dormant, stem cells within the highly purified HSC population. These results identify a new approach for isolating highly potent human HSCs for further clinical applications. They also implicate mitochondria in the intrinsic regulation of human HSC quiescence and potency.

Prevention and treatment of anticipatory chemotherapy-induced nausea and vomiting in pediatric cancer patients and hematopoietic stem cell recipients: Clinical practice guideline update.

This 2021 clinical practice guideline update provides recommendations for preventing anticipatory chemotherapy-induced nausea and vomiting (CINV) in pediatric patients. Recommendations are based on systematic reviews that identified (1) if a history of acute or delayed CINV is a risk factor for anticipatory CINV, and (2) interventions for anticipatory CINV prevention and treatment. A strong recommendation to optimize acute and delayed CINV control in order to prevent anticipatory CINV is made. Conditional recommendations are made for hypnosis, systematic desensitization, relaxation techniques, and lorazepam for the secondary prevention of anticipatory CINV. No recommendation for the treatment of anticipatory CINV can be made.

Clinical and epidemiological characteristics of catheter-associated bloodstream infections in patients with hematological profile

Patients with hemoblastosis, especially recipients of hematopoietic stem cells, are the most immunocompromised group of patients, especially during prolonged episodes of drug-associated granulocytopenia. The aim of the study . To characterize catheter-associated bloodstream infections in patients with hematological profile. Materials and methods . During June to October 2017, catheter-associated bloodstream infections were detected in six patients in the oncogematology department. Results and discussion . All patients came from different regions of the Russian Federation. The diagnosis of infectious complication: catheter-associated bloodstream infection was established only in 33.3 %. When determining laboratory markers of sepsis, high numbers of procalcitonin were observed in 50.0 % of patients, presepsin in 100.0 The growth of the bacterium Burholderia cepacia was observed in both central venous catheters (CVCs) and blood taken from CVCs. Phentotypically, all isolated cultures of Burholderia cepacia can be divided into three types: pan-resistant; polyresistant; polyresistant, but keeping sensitivity to co-trimoxazole. Multilocus sequencing of their DNA was carried out. The same sequence type was found in four patients. Given that the patients came from different regions, had not previously been in the same medical organization, care for CVC in the department of oncogematology was carried out by the same medical staff it can be assumed that four cases were caused by the same hospital strain Burkholderia cepacia.

Cytomegalovirus management after allogeneic hematopoietic stem cell transplantation: A mini-review

Because of the high incidence of cytomegalovirus (CMV) seropositivity in the population, CMV infection is a common and severe complication of allogeneic hematopoietic stem cell transplantation (allo-HSCT) in Taiwan. Here we propose a CMV management strategy for patients undergoing allo-HSCT from the Taiwanese perspective, which focuses on the epidemiology, diagnosis, monitoring, prophylaxis, and treatment of CMV infection after allo-HSCT. In terms of CMV monitoring, weekly CMV monitoring with the COBAS® AmpliPrep system is the standard approach because the pp65 CMV antigenemia assay has a lower sensitivity than CMV monitoring with the COBAS® AmpliPrep system. However, pp65 CMV antigenemia assay has a better correlation with clinical symptoms in immunocompromised patients. A 14-week prophylactic course of letermovir is recommended for allo-HSCT recipients in Taiwan, especially for recipients of hematopoietic stem cells from mismatched unrelated and haploidentical donors. Preemptive ganciclovir therapy should be initiated when the CMV viral load exceeds 1000 copies/mL, and should not be discontinued until CMV DNA is no longer detected in the blood. For allo-HSCT recipients who have CMV-related diseases, ganciclovir with or without CMV-specific intravenous immunoglobulin is the standard of care. The limited availability of foscarnet, an alternative for patients who are not responsive to or cannot tolerate ganciclovir, is a crucial issue in Taiwan. For pediatric allo-HSCT recipients, more data are needed to propose a CMV management recommendation.

Tuberculosis in Pediatric Solid Organ and Hematopoietic Stem Cell Recipients.

Children undergoing solid organ and hematopoietic stem cell transplantation are at high risk of morbidity and mortality from tuberculosis (TB) disease in the post-transplant period. Treatment of TB infection and disease in the post-transplant setting is complicated by immunosuppression and drug interactions. There are limited data that address the unique challenges for the management of TB in the pediatric transplant population. This review presents the current understanding of the epidemiology, clinical presentation, diagnosis, management, and prevention for pediatric transplant recipients with TB infection and disease. Further studies are needed to improve diagnosis of TB and optimize treatment outcomes for these patients.

Cytomegalovirus after allogeneic hematopoietic stem cell transplantation: reactivation or reinfection?

Objective. To determine type of cytomegalovirus (CMV) infection (reactivation of a virus strain that was present before transplantation or re-infection with another virus strain) in allogeneic hematopoietic stem cell recipients by sequencing. Materials and Methods. Clinical and laboratory data of 179 recipients of allogeneic hematopoietic stem cells collected from 2014 to 2018 were analyzed for CMV DNA in clinical specimens before and after transplantation. A total of 14 patients (28 samples) were included in the study. The CMV UL139 gene encoding viral glycoprotein was chosen for virus genotyping and sequence alignment. The primers complementary to its conservative sites were used. The resulting DNA sequence was analyzed using nucleotide BLAST software (https://blast.ncbi.nlm.nih.gov/Blast.cgi) and Genome compiler (https://designer.genomecompiler.com). The type of infection was determined by comparing DNA sequences before and after transplantation. Results. All enrolled patients were anti-CMV-positive prior to transplantation, which indicates the presence of CMV infection. Therefore, none of the patients had a primary infection as a result of transplantation. Of 14 patients, high percentage of sequence alignment (~100%) was observed in 8 patients. For the other 6 patients, substantial differences in sequences which indicate the different genotypes and the different type of infection were found. However, there was no statistically significant difference in the time to viral DNA appearance after transplantation in patients with re-infection and reactivation (p &gt; 0.05), nor was there a statistically significant correlation between the type of infection (reactivation/re-infection) and the main diagnosis or transfusion load. Conclusions. Reactivation of the previously registered viral strain and re-infection with another viral strain were equally probable (8 vs. 6 cases). There were no associations between the main diagnosis and the type of infection (reactivation/re-infection) possibly due to a small sample size. Time to post-transplantation CMV DNA detection in blood was longer for re-infection (median of 69.5 days) compared to reactivation (median of 27 days), but this difference was also non-statistically significant. In addition, there was no significant contribution of blood transfusion burden to the type of CMV infection, which may suggest the donor blood is not a source of the different strains.

1102. Reconstitution of CMV-specific cell-mediated immunity during letermovir prophylaxis in hematopoietic stem cell recipients

BackgroundPatients who are cytomegalovirus (CMV) seropositive (R+) prior to hematopoietic cell transplant (HCT), have 30% incidence of clinically significant CMV reactivation in the absence of prophylaxis. At our institution, letermovir prophylaxis through Day 100 is used in CMV R+ high-risk (HR) (cord blood, haplocord, haploidentical) HCT recipients. We hypothesized that clinically nonsignificant CMV reactivation during letermovir prophylaxis may lead to reconstitution of CMV specific cell mediated immunity (CMV CMI), which may protect the host against CMV disease after letermovir discontinuation.MethodsBlood samples from CMV R+ HR HCT recipients on letermovir were tested by dual color CMV specific IL2/IFNg FLUOROSpot pre-transplant and on Days 100, 182 and 360 post-transplant. Clinical and virologic information were obtained from medical records.ResultsAmong 35 participants enrolled to date, 19 were eligible for this analysis, which included only participants with CMV CMI defined as ≥20 spot-forming cells/106 PBMC pre-transplantation and follow up ≥180 post-transplantation. Median age was 51.5 years (range 22-75), 9 were women, 9 were white non-Hispanic, 8 were Hispanic and the most common underlying malignancy was acute myeloid leukemia (n=10). 14 participants had CMV CMI reconstitution at Day 100; including 5 with and 9 without low level CMV DNAemia, defined as <5000 international units/ml in whole blood quantitative polymerase chain reaction assay, while on letermovir prophylaxis. Among the 14 participants, 11 remained free of clinically significant CMV reactivation for a median (range) of 260 (80; 260) days post-letermovir discontinuation, while 3 developed acute graft vs. host disease (aGvHD) followed by clinically significant CMV reactivation. 5 participants did not reconstitute CMV CMI at Day 100 and none of them had DNAemia while on letermovir. 1 of 5 participants without CMV CMI reconstitution or aGvHD developed CMV disease after letermovir discontinuation.ConclusionHigh-risk patient populations can reconstitute CMV CMI while on letermovir. Ongoing investigation will help establish predictive parameters for CMV CMI that may allow risk stratification for CMV monitoring and letermovir usage.Disclosures Maheen Abidi, MD, Merck (Research Grant or Support) Jonathan Gutman, MD, Merck (Research Grant or Support) Adriana Weinberg, MD, GSK (Grant/Research Support)merck (Grant/Research Support)

Impaired host response and the presence of Acinetobacter baumannii in the serum microbiome of type-II diabetic patients.

SummaryType II diabetes (T2D) affects over 10% of the US population and is a growing disease worldwide that manifests with numerous comorbidities and defects in inflammation. This dysbiotic host response allows for infection of the host by numerous microorganisms. In the course of T2D disease, individuals can develop chronic infections including foot ulcers and periodontitis, which lead to further complications and opportunistic infections in multiple body sites. In this study, we investigated the serum of healthy subjects and patients with T2D with (T2DP) or without periodontitis for both microbiome signatures in addition to cytokine profiles. Surprisingly, we detected the presence of Acinetobacter baumanii in the serum of 23% individuals with T2D/T2DP tested. In T2DP, IL-1β, TNF-α, MCP-1, IL-6, IL-8, and IFN-γ were significantly elevated in ABC-positive subjects. As an emerging pathogen, A. baumanii infection represents a risk for impaired inflammation and the development of comorbidities in subjects with T2D.

High Dose Valaciclovir As CMV Prophylaxis in Allogeneic Hematopoietic Stem Cell Recipients at High Risk of CMV Reactivation

Background Cytomegalovirus (CMV) is a common, potentially devastating complication of allogeneic haematopoietic stem cell transplantation (alloHSCT). Universal antiviral prophylaxis strategies including letermovir are effective, but unsubsidised in Australia. Prophylactic ganciclovir or valganciclovir are challenging due to myelotoxicity. Valaciclovir demonstrates anti-CMV activity in high doses, but little current data explore prophylaxis in the alloHSCT setting, particularly in haploidentical transplantation. We aimed to evaluate the clinical efficacy and tolerability of high dose valaciclovir (high dose VALA) as CMV prophylaxis in high risk patients undergoing alloHSCT. Methods This study was completed at the Royal Melbourne Hospital, Melbourne Australia. We performed a retrospective analysis of alloHSCT recipients at high risk of CMV reactivation (defined as recipient and/or donor CMV seropositivity, and undergoing T-cell depletion, haploidentical or umbilical cord stem cell transplantation). Patients transplanted between July 2018 - June 2019, treated with high dose VALA (2g TDS) from day +7 to +100 and beyond were compared to a historical cohort (transplanted between July 2017 - June 2018) on standard dose valaciclovir (std dose VALA) (500mg BD until engraftment then 500mg daily). We compared the rates and time to reach a CMV threshold of 400 IU/ml, at which point pre-emptive CMV therapy was commenced. Tolerability was also evaluated. Results Patient demographics are described in Table 1. Of the standard dose VALA cohort, (median follow-up 259 days), 23/31 (74%) developed a viral load &amp;gt;400 IU/mL, requiring pre-emptive CMV therapy. None had CMV disease. Median time to viral load &amp;gt;400 IU/mL was 39 days (range 13 - 68). Of the high dose VALA cohort (median follow-up 209 days), 11/25 (44%) developed a viral load &amp;gt;400 IU/mL, requiring pre-emptive CMV therapy. Of these 11 cases, 7 patients had viral load &amp;gt;400 IU/mL while on high dose VALA prior to D+100, 3 patients had ceased high dose VALA prior to D+100 due to intolerance and in 1 patient this occurred post D+100 while on high dose VALA. One patient developed CMV (gut) disease following early cessation of high dose VALA, whilst on standard dose VALA. Median time to reactivation &amp;gt;400 IU/mL was 64 days (range 26-170). Time to reactivation &amp;gt;400 IU/ml was significantly different between the standard vs high dose VALA cohorts (mean ± SEM; 37.9 ± 2.7 vs 67.8 ± 11.3 days, **p=0.0015). Median duration of high dose VALA prophylaxis was 50 days (range 11-288). Seven (28%) patients continued high dose VALA to day +100 and beyond. Intolerance led to early cessation in 10 (40%) patients (acute kidney injury, n=6; cytopenia, n=3; both, n=1). Other patients ceased due to requirement for definitive CMV therapy (n=6) and unclear reasons (n=2). Conclusions In high risk alloHSCT recipients, high dose VALA is an effective CMV prophylactic strategy resulting in lower CMV reactivation rates, and delays CMV reactivation. This may reduce requirements for myelotoxic CMV treatment during the early post-engraftment period and need for inpatient admission. CMV infection following high dose VALA cessation remains a risk, particularly when dose reductions have occurred due to toxicity, and intolerance and ongoing monitoring is required. Treatment tolerability remains a limitation. Disclosures No relevant conflicts of interest to declare. OffLabel Disclosure: Valaciclovir, for CMV prophylaxis

5-Azacytidine Depletes Hematopoietic Stem Cells and Synergizes with an Anti-CD117 Antibody to Augment Engraftment of Donor Stem Cells in Immunocompetent Mice

Monoclonal antibody (mAb)-targeting of CD117 (c-Kit) present on hematopoietic stem cells (HSC) is an emerging conditioning strategy for hematopoietic cell transplantation (HCT) to replace standard-of-care alkylator and radiation therapy used to deplete recipient HSC and permit donor HSC engraftment. In preclinical and clinical studies anti-CD117 mAbs, which inhibit stem cell factor (SCF) from binding to CD117, have shown single agent efficacy in permitting donor HSC engraftment in immunocompromised recipients. However, preclinical data suggest that anti-CD117 mAb, ACK2, used as a single agent is insufficient to enable HSC engraftment in immunocompetent mice. Hence, there is a desire to potentiate the effect of naked anti-CD117 mAbs to permit targeted elimination of host HSC for a range of immune competent bone marrow (BM) states, from monogenic disorders, e.g., hemoglobinopathies, to BM clonal malignant diseases. Here, we show that ACK2 synergizes with the widely used hypomethylating agent, 5-Azacytidine (AZA), to deplete HSC and enable robust long-term HSC engraftment in immunocompetent mice. The cell depleting effect of AZA or ACK2+AZA was first tested on mature and primitive hematopoietic cells of C57BL/6 (B6) mice. Mice received AZA at 5mg/kg i.p. for 5 days either alone or combined with ACK2 at 500 µg i.v. 5 days prior to AZA. BM and spleen were harvested days 6, 10 and 20 after the first AZA dose and analyzed by extended immunophenotype. For HCT studies both congenic [B6 (H-2b, Thy1.1, CD45.1) into B6 recipients (H-2b, Thy1.1, CD45.1/CD45.2)] and allogeneic [B6 (H-2b, Thy1.1, CD45.1) into BALB.B recipients (H-2b, Thy1.2, CD45.2)] transplantation mouse models were tested. Single agent AZA induced rapid depletion of CD117+ cells in the BM, including LT-HSC (Lin-Sca+Kit+[LSK]CD150+CD48-) and ST-HSC (LSKCD150-CD48-). In vivo HSC depletion by AZA has not been previously reported, and was surprising given that most HSC are quiescent, and non-proliferating cells are known to be insensitive to the toxicities of AZA. Examination of HSC populations in the spleen did not suggest that mobilization of LT- or ST-HSC to secondary organs caused the decrease in BM. Rather, AZA-induced myelosuppression was followed by rapid expansion of myeloid-biased MPP and increased HSC proliferation as assessed by Ki67 intracellular protein expression. We hypothesized, that the in vivo HSC-depletive effects of AZA might result from increased sensitivity of proliferating HSC to AZA. Hence, the in vitro effects of different AZA concentrations were tested on proliferating HSC. In both mouse and human HSC cultures, supplemented with SCF and TPO, we observed that AZA reduced the number of cells in a dose dependent manner and decreased cell viability at concentrations of ≥ 0.5 µg/ml. In vivo, combined ACK2+AZA led to more robust HSC depletion and prolonged time to HSC recovery compared to AZA alone. We then evaluated if blockade of CD117 with its ligand SCF by ACK2 was the cause of this prolonged HSC-depletion. 2B8 mAb, which also binds mouse CD117, but only partially inhibits CD117/SCF signaling, did not result in augmented in vivo HSC depletion (Figure 1A). We further tested if immunocompetent mice treated with ACK2+AZA could be "rescued" with exogenous recombinant SCF. SCF-treatment resulted in faster recovery of LT-HSC. Together these studies suggest that the synergy of ACK2+AZA on the depth and duration of HSC-depletion depends of the disruption of CD117/SCF binding. Finally, we tested ACK2+AZA conditioning on donor stem cell engraftment in immunocompetent mice. Transplantation of 15-20x106 WBM cells following ACK2+AZA conditioning led to significantly increased stable mean multilineage donor chimerism of &amp;gt;60% in the congenic (Figure 1B) and &amp;gt;20% in the allogeneic setting. Single agent AZA enabled sustained donor engraftment with levels of myeloid donor chimerism of 10-15%, serving as proof of principle that AZA treatment can indeed clear HSC niches and permit engraftment of true HSC. Similar to WBM grafts, 5x104 purified HSC (LSK) engrafted successfully in the ACK2+AZA group, reaching long-term (26 weeks) mean donor myeloid chimerism &amp;gt;25% in both congenic and allogeneic settings. Our studies reveal a previously unknown effect of AZA on BM HSC and provide a platform for the clinical use of a novel less toxic non-myeloablative conditioning strategy for HCT, which is aimed at the rapid translation into clinical practice. Disclosures Pang: Jasper Therapeutics, Inc: Current Employment, Current equity holder in private company. Shizuru:Jasper Therapeutics, Inc: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees.

The Epstein-Barr Virus Major Tegument Protein BNRF1 Is a Common Target of Cytotoxic CD4+ T Cells.

Cellular immunotherapy is a proven approach against Epstein-Barr virus (EBV)-driven lymphoproliferation in recipients of hematopoietic stem cells. Extending the applicability and improving the response rates of such therapy demands improving the knowledge base. We studied 23 healthy donors for specific CD4+ T cell responses against the viral tegument protein BNRF1 and found such T cells in all seropositive donors, establishing BNRF1 as an important immune target in EBV. We identified 18 novel immune epitopes from BNRF1, all of them generated by natural processing of the full-length protein from virus-transformed lymphoblastoid cell lines (LCL). BNRF1-specific CD4+ T cells were measured directly ex vivo by a cytokine-based method, thus providing a tool to study the interaction between immunity and infection in health and disease. T cells of the cytotoxic Th1 type inhibited the proliferation of autologous LCL as well as virus-driven transformation. We infer that they are important in limiting reactivations to subclinical levels during health and reducing virus propagation during disease. The information obtained from this work will feed into data sets that are indispensable in the design of patient-tailored immunotherapeutic approaches, thereby enabling the stride toward broader application of T cell therapy and improving clinical response rates.IMPORTANCE Epstein-Barr virus is carried by most humans and can cause life-threatening diseases. Virus-specific T cells have been used in different clinical settings with variable success rates. One way to improve immunotherapy is to better suit T cell generation protocols to viral targets available in different diseases. BNRF1 is present in viral particles and therefore likely available as a target for T cells in diseases with virus amplification. Here, we studied healthy Epstein-Barr virus (EBV) carriers for BNRF1 immunogenicity and report our results indicating BNRF1 to be a dominant target of the EBV-specific CD4+ T cell response. BNRF1-specific CD4+ T cells were found to be cytotoxic and capable of limiting EBV-driven B cell transformation in vitro The findings of this work contribute to forwarding our understanding of host-virus interactions during health and disease and are expected to find direct application in the generation of specific T cells for immunotherapy.

Adenovirus infection among pediatric patients with cancer and in pediatric recipients of hematopoietic stem cell: A multicenter nationwide study.

The aim was to evaluate the incidence, clinical course, and outcome of adenoviral infection (AdVI) in pediatric patients diagnosed and treated due to cancer and in pediatric recipients of hematopoietic stem cell. Over a 72-month period, all-in 5599 children with cancer: 2441 patients with hematological malignancy (HM) and 3158 with solid tumors (ST), and 971 patients after transplantation: 741 after allogeneic (allo-HSCT) and 230 after autologous (auto-HSCT) were enrolled into the study. Among cancer patients, 67 episodes of AdVI appeared in 63 (1.1%) children, including 45 (1.8%) with HM and 18 (0.6%; P < .001) with ST. Within transplanted patients, AdVIs were responsible for 88 episodes in 81 (8.3%) children (P < .001), including 78 (10.5%) patients after allo-HSCT and 3 (1.3%) after auto-HSCT. Time to develop AdVI was short, especially after allo-HSCT. The most common clinical manifestation in cancer patients was enteritis diagnosed in 63 (94.0%) cases, while among HSCT recipient asymptomatic adenoviremia was found in 36 (40.9%) cases and the most common clinical manifestation was urinary tract infection. Cancer patients with disseminated disease, as well as HSCT recipients with either asymptomatic viremia or disseminated disease, received antiviral treatment. The most commonly used first-line therapy was cidofovir. None of the cancer patients died due to AdVI, while within HSCT recipients three patients developed disseminated adenoviral disease and died despite antiviral treatment. In cancer patients, AdVIs are rare and associated with very good prognosis even without specific treatment. However, in allo-HSCT recipients, disseminated disease with fatal outcome is more likely to occur.

COVID‐19 in bone marrow transplant recipients: reflecting on a single centre experience

Covid-19 disease is caused by a novel SARS-CoV-2 virus and has been declared a pandemic on the 9th of March by WHO. Hallmark of COVID-19 management is supportive care and there is still no convincing evidence on a treatment which will reduce mortality. Severe COVID-19 associated sepsis characterized by acute respiratory distress syndrome (ARDS), secondary bacterial pneumonias thrombotic complications, myocarditis, and gastrointestinal involvement are more prevalent in those with comorbidities such as hypertension, diabetes, cardiac disease, cancer and age>70 years (Liang et al; 2020, Guan et al; 2019).

Safety of tunneled central venous catheters in pediatric hematopoietic stem cell recipients with severe primary immunodeficiency diseases.

Tunneled central venous catheters (TCVCs) provide prolonged intravenous access for pediatric patients with severe primary immunodeficiency disease (PID) undergoing hematopoietic stem cell transplantation (HSCT). However, little is known about the epidemiology and clinical significance of TCVC-related morbidity in this particular patient group. We conducted the retrospective analysis of patients with severe PID who received percutaneous landmark-guided TCVC implantation prior to HSCT. We analyzed 92 consecutive TCVC implantations in 69 patients (median [interquartile range] age 3.0 [0-11] years) with severe combined immune deficiency (n = 39, 42.4%), chronic granulomatous disease (n = 17, 18.4%), and other rare PID syndromes (n = 36, 39.2%). The median length of TCVC observation was 144.1 (85.5-194.6) days with a total of 14,040 catheter days at risk (cdr). The overall rate of adverse events during catheter insertion was 17.4% (n = 16) and 25.0% during catheter dwell period (n = 23, catheter risk [CR] per 1000 cdr = 1.64). The most common complication was TCVC-related infection with an overall prevalence of 9.8% (n = 9, CR = 0.64), followed by late dislocation (n = 6, 6.5%, CR = 0.43), early dislocation (n = 4, 4.3%) and catheter dysfunction (n = 4, 4.3%, CR = 0.28). TCVCs are safe in children with severe PID undergoing HSCT with relatively low rates of TCVC-related infection.

Clonal hematopoiesis in donors and long-term survivors of related allogeneic hematopoietic stem cell transplantation

AbstractClonal hematopoiesis (CH) is associated with age and an increased risk of myeloid malignancies, cardiovascular risk, and all-cause mortality. We tested for CH in a setting where hematopoietic stem cells (HSCs) of the same individual are exposed to different degrees of proliferative stress and environments, ie, in long-term survivors of allogeneic hematopoietic stem cell transplantation (allo-HSCT) and their respective related donors (n = 42 donor-recipient pairs). With a median follow-up time since allo-HSCT of 16 years (range, 10-32 years), we found a total of 35 mutations in 23 out of 84 (27.4%) study participants. Ten out of 42 donors (23.8%) and 13 out of 42 recipients (31%) had CH. CH was associated with older donor and recipient age. We identified 5 cases of donor-engrafted CH, with 1 case progressing into myelodysplastic syndrome in both donor and recipient. Four out of 5 cases showed increased clone size in recipients compared with donors. We further characterized the hematopoietic system in individuals with CH as follows: (1) CH was consistently present in myeloid cells but varied in penetrance in B and T cells; (2) colony-forming units (CFUs) revealed clonal evolution or multiple independent clones in individuals with multiple CH mutations; and (3) telomere shortening determined in granulocytes suggested ∼20 years of added proliferative history of HSCs in recipients compared with their donors, with telomere length in CH vs non-CH CFUs showing varying patterns. This study provides insight into the long-term behavior of the same human HSCs and respective CH development under different proliferative conditions.

Adenovirus infections

Während der Kindheit werden zumeist mehrere Adenovirusinfektionen mit jeweils wechselnden Typen durchgemacht. Das Spektrum der assoziierten Krankheitsbilder reicht von endemischen milden, selbstlimitierenden Infektionen des oberen Respirationstrakts über epidemische Gastroenteritis, Augeninfektionen und Zystitis bis hin zu atypischen Pneumonien und schweren Infektionen innerer Organe. Lebensbedrohlich sind die meisten Adenovirusinfektionen nicht, außer die seltenen Infektionen von Neugeborenen und die allerdings häufigen Infektion von Empfängern hämatopoetischer Stammzellen („disseminierte Erkrankung“).