Mesenchymal stem cell infusion for enhancing hematopoietic recovery and addressing cytopenias in CAR-T cell therapy.

BACKGROUND: Chimeric antigen receptor (CAR)-T therapy has emerged as a promising treatment for hematologic malignancies. However, cytopenia remains one of the most frequent and challenging adverse effects of this therapy. METHODS: We conducted a retrospective analysis of 26 patients with relapsed/refractory aggressive B-cell lymphoma who received CAR-T therapy at our center. Subsequently, to investigate measures to address cytopenias following CAR-T therapy, we isolated and generated murine CAR-T cells and bone marrow-derived mesenchymal stem cells (MSCs), establishing a murine syngeneic CAR-T therapy model. We assessed the impact of MSC infusion on hematopoietic recovery post-CAR-T therapy by evaluating complete blood count, bone marrow hematopoietic stem cells and their subpopulations, bone marrow histomorphology, and hematopoiesis-related genes. RESULTS: All patients experienced cytopenias to varying degrees, with complete lineage involvement in half of the patients. Grade ≥ 3 cytopenias were observed in 88.46% of the patients. CAR-T therapy was associated with a higher incidence of biphasic, late-onset, or prolonged cytopenias. Survival analysis indicated that neutropenia and lymphopenia tended to be associated with better prognosis, whereas thrombocytopenia tended to be related to poorer outcomes. Through animal experiments, we discovered that MSCs infusion boosted HSCs and their long-term subpopulations, enhancing hematopoietic recovery, particularly in the megakaryocyte lineage, and mitigating bone marrow damage. Importantly, both in vitro and in vivo experiments demonstrated that MSCs did not compromise the activity or antitumor efficacy of CAR-T cells. CONCLUSIONS: Our findings propose MSCs infusion as a promising strategy to address cytopenias, particularly thrombocytopenia, after CAR-T therapy. This approach could help overcome certain limitations of cellular immunotherapy by enhancing hematopoietic recovery without compromising the efficacy of CAR-T cells. HIGHLIGHTS: 1 Cytopenia is a frequently observed adverse effect following CAR-T therapy, and it is often characterized by biphasic and prolonged patterns. 2 MSCs play a critical role in promoting hematopoietic recovery and mitigating bone marrow damage in a murine model of CAR-T therapy 3 The activity and antitumor efficacy of CAR-T cells were not impaired by MSCs.

Moxibustion combined with guasha therapy for recurrent neutropenia following multiple cycles of chemotherapy of ovarian cancer: A case report.

Neutropenia, a common side effect of chemotherapy for ovarian cancer, was observed in a 47-year-old female patient undergoing a six-cycle chemotherapy regimen. She experienced recurrent neutropenia and leukopenia but refused granulocyte colony-stimulating factor (G-CSF) due to severe bone pain and high costs. Moxibustion combined with guasha therapy (MGT) was administered each time neutropenia occurred. The treatment involved guasha therapy on the bladder meridian (BL) and the governor vessel (GV), followed by moxibustion at Zhongwan (CV 12), Guanyuan (CV 4), and Shenzhu (GV 12) points over 2-3 days. This approach led to the recovery of neutrophil and leukocyte counts, enabling the patient to complete six chemotherapy cycles without G-CSF. These findings suggest that MGT may enhance neutrophil and leukocyte counts in patients with chemotherapy-induced myelosuppression, presenting a potential alternative for those intolerant to G-CSF. However, further high-quality research is needed to confirm its efficacy.

Oral health management in children with severe congenital neutropenia with periodontitis: Case report.

RATIONALE: Severe congenital neutropenia (SCN) is a rare and heterogeneous genetic disease. By describing the diagnosis and treatment of a child with SCN and periodontitis, this case provides a reference for the oral health management of a child with SCN and periodontitis. PATIENT CONCERNS: We describe a boy with clinical manifestations of oral bleeding, neutropenia, recurrent fever, and other recurrent infections. The absolute neutrophil count (ANC) was <0.50 × 109/L most of the time. Morphological examination of bone marrow cells showed active granulocyte hyperplasia and dysmaturation. DIAGNOSES: According to the clinical manifestations, hematological examination and gene detection results, the child was diagnosed as SCN with chronic periodontitis. INTERVENTIONS: Periodontal treatment was performed after informed consent was obtained from the child guardian. These included supragingival and subgingival cleaning, hydrogen peroxide and saline irrigation, placement of iodoglycerin in the gingival sulcus, and oral hygiene instruction. Hematopoietic stem cell transplantation (HSCT) was performed later. OUTCOMES: One month after initial periodontal treatment, oral hygiene was well maintained and gingival swelling had subsided. Probing depth (PD) index on periodontal probing and bleeding was significantly reduced. However, there was no significant change in blood routine and other indicators before and after periodontal treatment. CONCLUSION: Once SCN is diagnosed, individualized treatment plans can be developed according to the characteristics of the disease and its impact on oral health, which can effectively control the interaction between SCN and periodontal disease and reduce the occurrence of serious infection.

Absence of Neutropenia in Patients With Early Exon Nonsense Mutations in ELANE : Clinical Evidence to Support Gene Therapy Approaches for Severe Congenital Neutropenia.

Severe congenital neutropenia is an inherited bone marrow failure disorder characterized by profoundly low neutrophil counts and promyelocytic maturation arrest in bone marrow. Severe congenital neutropenia is most often caused by heterozygous ELANE mutations. In vitro and mouse xenograft studies using CRISPR/Cas9 have shown that introduction of frameshift/nonsense mutations in mutant ELANE may restore neutrophil counts, providing a model for gene therapy. Here, we present 2 children with inherited nonsense mutations in ELANE analogous to those proposed for gene therapy. Their normal peripheral blood neutrophil counts provide support for this approach through human "experiments of nature."

Prolonged cytopenias after immune effector cell therapy and lymphodepletion in patients with leukemia, lymphoma and solid tumors.

BACKGROUND AIMS: The success of chimeric antigen receptor (CAR) T-cell therapy in treating B-cell malignancies has led to the evaluation of CAR T-cells targeting a variety of other malignancies. Although the efficacy of CAR T-cells is enhanced when administered post-lymphodepleting chemotherapy, this can trigger bone marrow suppression and sustained cytopenia after CD19.CAR T-cell therapy. Additionally, systemic inflammation associated with CAR T-cell activity may contribute to myelosuppression. Cytopenias, such as neutropenia and thrombocytopenia, elevate the risk of severe infections and bleeding, respectively. However, data on the incidence of prolonged cytopenias after immune effector therapy in the solid tumor context remain limited. OBJECTIVE: We compared the incidence of prolonged cytopenias after immune effector therapy including genetically modified T-cells, virus-specific T-cells (VSTs) and NKT-cells, as well non-gene-modified VSTs for leukemia, lymphoma, and solid tumors (ST) to identify associated risk factors. METHODS: A retrospective analysis was conducted of 112 pediatric and adult patients with relapsed and/or refractory cancers who received lymphodepleting chemotherapy followed by immune effector therapy. Patients treated with 13 distinct immune effector cell therapies through 11 single-center clinical trials and 2 commercial products over a 6-year period were categorized into 3 types of malignancies: leukemia, lymphoma and ST. We obtained baseline patient characteristics and adverse events data for each participant, and tracked neutrophil and platelet counts following lymphodepletion. RESULTS: Of 112 patients, 104 (92.9%) experienced cytopenias and 88 (79%) experienced severe cytopenias. Patients with leukemia experienced significantly longer durations of severe neutropenia (median duration of 14 days) compared with patients with lymphoma (7 days) or ST (11 days) (P = 0.002). Patients with leukemia also had a higher incidence of severe thrombocytopenia (74.1%), compared with lymphoma (46%, P = 0.03) and ST (14.3%, P < 0.0001). Prolonged cytopenias were significantly associated with disease type (63% of patients with leukemia, 44% of patients with lymphoma, and 22.9% of patients with ST, P = 0.006), prior hematopoietic stem cell transplant (HSCT) (66.7% with prior HSCT versus 38.3% without prior HSCT, P = 0.039), and development of immune effector cell-associated neurotoxicity syndrome (ICANS) (75% with ICANS versus 38% without ICANS, P = 0.027). There was no significant association between prolonged cytopenias and cytokine release syndrome. CONCLUSIONS: Immune effector recipients often experience significant cytopenias due to marrow suppression following lymphodepletion regardless of disease, but prolonged severe cytopenias are significantly less common after treatment of patients with lymphoma and solid tumors.

CRISPR-Cas9n-mediated ELANE promoter editing for gene therapy of severe congenital neutropenia.

Severe congenital neutropenia (CN) is an inherited pre-leukemia bone marrow failure syndrome commonly caused by autosomal-dominant ELANE mutations (ELANE-CN). ELANE-CN patients are treated with daily injections of recombinant human granulocyte colony-stimulating factor (rhG-CSF). However, some patients do not respond to rhG-CSF, and approximately 15% of ELANE-CN patients develop myelodysplasia or acute myeloid leukemia. Here, we report the development of a curative therapy for ELANE-CN through inhibition of ELANE mRNA expression by introducing two single-strand DNA breaks at the opposing DNA strands of the ELANE promoter TATA box using CRISPR-Cas9D10A nickases-termed MILESTONE. This editing effectively restored defective neutrophil differentiation of ELANE-CN CD34+ hematopoietic stem and progenitor cells (HSPCs) in vitro and in vivo, without affecting the functions of the edited neutrophils. CRISPResso analysis of the edited ELANE-CN CD34+ HSPCs revealed on-target efficiencies of over 90%. Simultaneously, GUIDE-seq, CAST-Seq, and rhAmpSeq indicated a safe off-target profile with no off-target sites or chromosomal translocations. Taken together, ex vivo gene editing of ELANE-CN HSPCs using MILESTONE in the setting of autologous stem cell transplantation could be a universal, safe, and efficient gene therapy approach for ELANE-CN patients.

Extracorporeal membrane oxygenation in long-term COVID-19 with severe neutropenia and thrombocytopenia after allogeneic hematopoietic stem cell transplantation: a case report.

BACKGROUND: Hematopoietic stem cell transplantation (HSCT) was associated with potentially life-threatening complications. Among patients supported by extracorporeal membrane oxygenation (ECMO), those who underwent HSCT had a worse prognosis than those who did not. Advances in HSCT and critical care management have improved the prognosis of ECMO-supported HSCT patients. CASE: The patient in the remission stage of lymphoma after 22 months of allogeneic hematopoietic stem cell transplantation, suffered from ARDS, severe neutropenia, thrombocytopenia, and long-term COVID-19. We evaluated the benefits and risks of ECMO for the patient, including the possibility of being free from ECMO, the status of malignancy, the interval from HSCT to ARDS, the function of the graft, the amount of organ failure, and the comorbidities. ECMO was ultimately used to save his life. CONCLUSIONS: We did not advocate for the general use of ECMO in HSCT patients and we believed that highly selected patients, with well-controlled tumors, few comorbidities, and fewer risk factors for death, tended to benefit from ECMO with well ICU management.

Clinical practice guideline-inconsistent management of fever and neutropenia in pediatric oncology: A Children's Oncology Group study.

BACKGROUND: The primary objective was to measure the proportion of episodes where care delivery was inconsistent with selected recommendations of a clinical practice guideline (CPG) on fever and neutropenia (FN) management. The influence of site size on CPG-inconsistent care delivery, and association between patient outcomes and CPG-inconsistent care were described. METHODS: This retrospective, multicenter study included patients less than 21 years old with cancer who were at high risk of poor FN outcomes and were previously enrolled to a Children's Oncology Group (COG) study at participating National Cancer Institute Community Oncology Research Program (NCORP) institutions from January 2014 through December 2015. Patients were randomly selected for chart review by participating sites from a COG-generated list. Care delivered in each episode was adjudicated (CPG-consistent or CPG-inconsistent) against each of five selected recommendations. RESULTS: A total of 107 patients from 22 sites, representing 157 FN episodes, were included. The most common CPG-inconsistent care delivered was omission of pulmonary computerized tomography in patients with persistent FN (60.3%). Of 74 episodes where assessment of four (episodes without persistent FN) or five (episodes with persistent FN) recommendations was possible, CPG-inconsistent care was delivered with respect to at least one recommendation in 63 (85%) episodes. Site size was not associated with CPG-inconsistent care delivery. No statistically significant association between CPG-inconsistent care and fever recurrence was observed. CONCLUSIONS: In this cohort of pediatric patients at high risk of poor FN outcomes, CPG-inconsistent care was common. Opportunities to optimize resource stewardship by boosting supportive care CPG implementation are highlighted.

The promise of novel treatments for severe chronic neutropenia.

INTRODUCTION: Severe chronic neutropenia, i.e. absolute neutrophil count (ANC) less than 0.5 × 109/L, is a serious health problem because it predisposes patients to recurrent bacterial infections. Management radically changed with the discovery that granulocyte colony-stimulating factor (G-CSF) could be used to effectively treat most patients; therapy required regular subcutaneous injections. In the early days of G-CSF therapy, there were concerns that it might somehow overstimulate the bone marrow and cause myelodysplasia (MDS) or acute myeloid leukemia (AML). Detailed research records from the Severe Chronic Neutropenia International Registry (SCNIR) indicate that this is a relatively low-risk event. The research records suggest that certain patient groups are primarily at risk. Presently, allogeneic hematopoietic stem cell therapy serves as an alternate form of therapy. AREAS COVERED: Due to these concerns and the desire for an easy-to-take oral alternative, several new treatments are under investigation. These treatments include neutrophil elastase inhibitors, SGLT-2 inhibitors, mavorixafor - an oral CXCR4 inhibitor, gene therapy, and gene editing. EXPERT OPINION: All of these alternatives to G-CSF are promising. The risks, relative benefits, and costs are yet to be determined.

Child and family perceptions of satisfaction with neutropenia management in pediatric acute myeloid leukemia.

PURPOSE: Chemotherapy for pediatric acute myeloid leukemia (AML) is very intensive and many, but not all centers, require extended hospitalization until neutrophil recovery. Child and family preferences, beliefs, and experiences around hospitalization have not been systematically assessed. PATIENTS AND METHODS: We recruited children with AML and their parents from nine pediatric cancer centers across the United States for a qualitative interview about their experiences of neutropenia management. Interviews were analyzed using a conventional content analysis approach. RESULTS: Of 116 eligible individuals, 86 (74.1%) agreed to participate. Interviews were conducted with 32 children and 54 parents from 57 families. Of these 57 families, 39 were cared for as inpatients and 18 were managed as outpatients. A very high proportion of respondents in both groups reported satisfaction with the discharge management strategy recommended by the treating institution: 86% (57 individuals) of respondents who experienced inpatient management and 85% (17 individuals) of respondents who experienced outpatient management expressed satisfaction. Respondent perceptions associated with satisfaction related to safety (access to emergency interventions, infection risk, close monitoring) and psychosocial concerns (family separation, low morale, social support). Respondents believed it could not be assumed that all children would have the same experience due to varied life circumstances. CONCLUSION: Children with AML and their parents express a very high degree of satisfaction with the discharge strategy recommended by their treating institution. Respondents saw a nuanced tradeoff between patient safety and psychosocial concerns that was mediated by a child's life circumstances.

USB1 is a miRNA deadenylase that regulates hematopoietic development.

Mutations in the 3' to 5' RNA exonuclease USB1 cause hematopoietic failure in poikiloderma with neutropenia (PN). Although USB1 is known to regulate U6 small nuclear RNA maturation, the molecular mechanism underlying PN remains undetermined, as pre-mRNA splicing is unaffected in patients. We generated human embryonic stem cells harboring the PN-associated mutation c.531_delA in USB1 and show that this mutation impairs human hematopoiesis. Dysregulated microRNA (miRNA) levels in USB1 mutants during blood development contribute to hematopoietic failure, because of a failure to remove 3'-end adenylated tails added by PAPD5/7. Modulation of miRNA 3'-end adenylation through genetic or chemical inhibition of PAPD5/7 rescues hematopoiesis in USB1 mutants. This work shows that USB1 acts as a miRNA deadenylase and suggests PAPD5/7 inhibition as a potential therapy for PN.

The essentials of acute oncology.

The general medical physician will often encounter patients who develop acute complications of their cancer diagnosis or anti-cancer treatment. Here we provide an overview of emergency solid tumour oncology to guide the initial management of these patients.

Diagnosis and management of isolated neutropenia: A survey of pediatric hematologist oncologists.

BACKGROUND: Isolated neutropenia is a common referral to pediatric hematology oncology (PHO) physicians. There are no established consensus guidelines in the diagnosis and management of patients with isolated, asymptomatic, and incidentally discovered neutropenia. METHODS: A survey was distributed to PHO physicians on the American Society of Pediatric Hematology Oncology member discussion page to determine the common diagnostic and management decisions regarding patients with isolated neutropenia and to explore beliefs regarding the term "benign ethnic neutropenia." RESULTS: One hundred twenty-six PHO attending physicians completed the survey. The most common tests reportedly ordered for this patient population included complete blood cell count (CBC) (98%), peripheral smear (75%), antineutrophil antibody testing (29%), and immunoglobulins (24%). Providers were more likely to order an antineutrophil antibody in toddlers (p = .0085), and antinuclear antibody (ANA) panels in adolescents (p < .001). Half of providers do not request additional CBCs prior to their initial consultation, and most suggest referring patients with mild neutropenia after confirming a declining absolute neutrophil count (ANC) (51%). The three most important factors influencing ongoing follow-up included: history of recurrent/severe infections (98%), family history of blood disorders (98%), and more severe/progressively worsening neutropenia (97%). Seventy percent of respondents have diagnosed patients with "benign ethnic neutropenia," and 75% support replacement of the term to "typical neutrophil count with Fy(a-/b-) status," if confirmed with red cell phenotyping. CONCLUSION: We identified practice patterns of PHO physicians for the diagnosis and management of patients referred for asymptomatic and isolated neutropenia. These data provide the framework to conduct cost-effectiveness studies.

How are granulocytes for transfusion best used? The past, the present and the future.

Granulocyte transfusions continue to be used in clinical practice, predominantly for treatment of refractory infection in the setting of severe neutropenia. There is biological plausibility for effectiveness in these patients with deficiencies of neutrophils, either as a consequence of disease or treatment. However, there is a chequered history of conducting and completing interventional trials to define optimal use, and many uncertainties remain regarding schedule and dose. Practice and clinical studies are severely limited by the short shelf life and viability of current products, which often restricts the timely access to granulocyte transfusions. In the future, methods are needed to optimise donor-derived granulocyte products. Options include use of manufactured neutrophils, expanded and engineered from stem cells. Further possibilities include manipulation of neutrophils to enhance their function and/or longevity. Granulocyte transfusions contain a heterogeneous mix of cells, and there is additional interest in how these transfusions may have immunomodulatory effects, including for potential uses as adjuncts for anti-cancer effects.

Neutropenia in Preterm Infants.

BACKGROUND: The non-specific and antigen-specific components of host defense mechanisms are subject to the adaptation process in the neonate; however, the neutrophil quantitative and qualitative deficiency is one of the most significant causative factors of neonatal-increased vulnerability to infection. OBJECTIVE: To review the incidence and outcome of neutropenia of unknown cause in preterm infants. RESULTS: The incidence of early and late-onset idiopathic neutropenia of prematurity is significant. CONCLUSION: The low neutrophil counts respond quickly to G-CSF treatment; however, due to the low probability of septic complications, particularly in the late-onset neutropenia, a deep diagnostic approach and the potential hematopoietic growth factor treatment should be limited to the severe cases, such as a neutrophil count <500/µL, lasting for more than 2 days.

How I manage inpatient consultations for quantitative neutrophil abnormalities in adults.

Neutrophilia and neutropenia commonly lead to inpatient hematology consultation. Quantitative neutrophil abnormalities have a broad differential and include diagnoses that are important to recognize because they may be associated with increased mortality. Neutrophilia can reflect etiologies such as infection, medications, inflammation, splenectomy, and congenital disorders. Neutropenia can arise from infection, medications, autoimmune destruction, sequestration, nutritional deficiency, malignancy, and congenital neutropenia syndromes. In the evaluation of all abnormalities of neutrophil number, the timing of the change, and the patient's historical neutrophil count are crucial.

Genetics of severe congenital neutropenia as a gateway to personalized therapy.

Severe congenital neutropenias (SCNs) are rare diseases, and to date about 30 subtypes have been described according to their genetic causes. Standard care aims to prevent infections and limit the risk of leukemic transformation; however, several subtypes may have additional organ dysfunction(s), requiring specialized care. Granulocyte colony-stimulating factor and hematopoietic stem cell transplantation are now the bedrock of standard care. Better understanding of SCN mechanisms now offers the possibility of adapted therapy for some entities. An inhibitor of sodium glucose cotransporter, an antidiabetic drug, may attenuate glycogen storage disease type Ib and glucose-6-phosphatase catalytic subunit 3 neutropenias by clearing 1,5-anhydroglucitol, the precursor of the phosphate ester responsible for these SCNs. Chemokine receptor CXCR4 inhibitors contribute to reversing the leukocyte defect in warts, hypoglobulinemia, infections, and myelokathexis syndrome. All these new approaches use oral drugs, which notably improve quality of life. Additionally, improved research into clonal evolution has highlighted some ways to potentially prevent leukemia, such as stimulating somatic genetic rescue, a physiological process that might limit the risk of leukemic transformation.

Invasive Pulmonary Aspergillosis Successfully Treated with Granulocyte Transfusions Followed by Hematopoietic Stem Cell Transplantation in a Patient with Severe Childhood Aplastic Anemia.

Granulocyte transfusions (GTX) have been used in patients with neutropenia or neutropenia associated with invasive fungal infection. An 11-year-old girl with severe aplastic anemia (SAA) received immunosuppressive therapy (IST) with rabbit antithymocyte globulin, cyclosporine, and granulocyte colony-stimulating factor. However, IST was not effective and her condition became complicated with life-threatening invasive pulmonary aspergillosis. Owing to the necessity for early neutrophil recovery to resolve the infection, GTX were performed, followed by bone marrow transplantation (BMT) from her mother with human leukocyte antigen-B locus mismatch. Her dyspnea improved and she eventually became afebrile after the initiation of GTX. Despite engraftment failure following BMT, successful engraftment was achieved by salvage therapy with peripheral blood stem cell transplantation. Chest computed tomography scan obtained 4 months after BMT revealed marked improvement in pneumonia. The current case illustrates that GTX may be useful in controlling invasive fungal infections before hematopoietic stem cell transplantation in patients with SAA.

New approaches to management of fever and neutropenia in high-risk patients.

PURPOSE OF REVIEW: Patients receiving treatment for acute leukaemia and haematopoietic cell transplantation (HCT) have prolonged neutropenia and are at high risk of neutropenic fever, with bacterial and particularly invasive fungal infections as feared complications, possessing potentially serious consequences including intensive care admission and mortality. Concerns for these serious complications often lead to long durations of broad-spectrum antimicrobial therapy and escalation to even broader therapy if fever persists. Further, the default approach is to continue neutropenic fever therapy until count recovery, leaving many patients who have long defervesced on prolonged antibiotics. RECENT FINDINGS: This article details recent progress in this field with particular emphasis on early discontinuation studies in resolved neutropenic fever and improved imaging techniques for the investigation of those with persistent neutropenic fever. Recent randomized controlled trials have shown that early cessation of empiric neutropenic fever therapy is well tolerated in acute leukaemia and autologous HCT patients who are clinically stable and afebrile for 72 h. Delineation of the best approach to cessation (timing and/or use of fluoroquinolone prophylaxis) and whether this approach is well tolerated in the higher risk allogeneic HCT setting is still required. Recent RCT data demonstrate utility of FDG-PET/CT to guide management and rationalize antimicrobial therapy in high-risk patient groups with persistent neutropenic fever. SUMMARY: Acute leukaemic and autologous HCT patients with resolved neutropenic fever prior to count recovery can have empiric therapy safely discontinued or de-escalated. There is an emerging role of FDG-PET/CT to support decision-making about antibiotic and antifungal use in high-risk persistent/recurrent neutropenic fever patients.

Adult Evans' Syndrome.

Evans syndrome (ES) is a rare immune disorder defined as the simultaneous or sequential occurrence in a single patient of immune thrombocytopenia (ITP) and warm autoimmune hemolytic anemia (wAIHA) ± autoimmune neutropenia (AIN). ES represents approximately 5% to 10% of all wAIHA and 2%-5% of all ITP cases in adults and its mortality rate is high. When ITP and wAIHA occurred concomitantly, other differential diagnoses must be ruled out. ES can be primary or secondary and isolated or associated with another underlying disorder and secondary ES. The management of ES is mostly empirical with a low level of evidence. This review reports some new insights on this rare disease and provides some practical tools for the diagnosis and management of adult ES.