HLA-haploidentical T-cell receptor αßT/B-cell-depleted stem cell transplantation for Fanconi anemia.

HLA-haploidentical stem cell transplantation (haplo-SCT) using post-transplant high-dose cyclophosphamide (PT-CY) is an alternative choice when a suitable donors is unavailable. However, PT-CY is difficult in patients with Fanconi anemia (FA) due to their high vulnerability to alkylating agents. For FA, we prefer haplo-SCT by T-cell receptor αßT-cell and B-cell depletion (αßT/B-depleted haplo-SCT), which can reduce the risks of PT-CY-related complications and graft-versus-host disease (GVHD). An 11-year-old boy with diagnosed FA (FANCG mutation) and bone marrow failure was to receive αßT/B-depleted haplo-SCT from his father (HLA 4/8 allele matched) due to absence of an HLA-matched donors. αßT/B-depleted peripheral blood stem cells (CD34 + cell count, 1.17 × 107/kg; αß + T-cell count, 1.3 × 105/kg) were infused following conditioning consisting of fludarabine (150 mg/m2), cyclophosphamide (40 mg/kg), anti-thymocyte globulin (5 mg/kg), rituximab (375 mg/m2), and thoraco-abdominal irradiation (3 Gy). Tacrolimus was used for GVHD prophylaxis until day + 30. Neutrophil engraftment was achieved on day + 9, and complete chimerism was confirmed on days + 28 and + 96. At 12-month post-SCT, the patient was well without GVHD or any other complications. αßT/B-depleted haplo-SCT is a good choice not only for patients unsuitable for PT-CY, but also for all pediatric recipients to reduce SCT-related complications.

A murine model to evaluate immunotherapy effectiveness for human Fanconi anemia-mutated acute myeloid leukemia.

Fanconi anemia (FA)-mutated acute myeloid leukemia (AML) is a secondary AML with very poor prognosis and limited therapeutic options due to increased sensitivity to DNA-damaging agents. PD-1 immune checkpoint inhibitors upregulate T-cell killing of cancer cells and is a class of promising treatment for FA-AML. Here, we developed a novel FA-AML murine model that allows the study of human AML with a humanized immune system in order to investigate immunotherapeutic treatments in vivo. FA-AML1 cells and non-FA-mutated Kasumi-1 cells were injected into 8-10 week old NSG mice. Once leukemic engraftment was confirmed by HLA-DR expression in the peripheral blood, human peripheral blood mononuclear cells (hPBMCs) were injected into the mice. One week post-hPBMCs injection, Nivolumab (PD-1 inhibitor) or PBS vehicle control was administered to the mice bi-weekly. In our Nivolumab treated mice, FA-AML1, but not Kasumi-1-engrafted mice, had significantly prolonged overall survival. Both FA-AML1 and Kasumi-1 engrafted mice had decreased spleen weights. Higher leukemic infiltration into vital organs was observed in FA-AML1 engrafted mice compared to Kasumi-1 engrafted mice. In conclusion, our novel humanized murine model of FA-mutated AML is an attractive tool for supporting further studies and clinical trials using PD-1 inhibitors to treat FA-mutated AML.

Liver abnormalities are frequent and persistent in patients with Fanconi anemia.

ABSTRACT: Liver disease has not been well described in patients with Fanconi anemia (FA). Improvements in outcomes of transplant mean that more individuals with FA are reaching adulthood and new features of the FA phenotype are being discovered. We performed a retrospective review of liver function in a cohort of 97 patients with FA followed-up for at least 10 years at a single center. We identified a high frequency of transaminitis (n = 31, 32%) without elevation of bilirubin and with no evidence of structural hepatic abnormality in patients with FA. Transaminitis was persistent in many cases, sometimes lasting more than a decade without clinical manifestation, although 2 patients with prolonged transaminitis are deceased from liver failure, indicating important long-term clinical consequences. Transaminitis was found in patients who had and had not received transplant but was more frequent in recipients of transplant. Exposure to total body irradiation increased risk (odds ratio, 15.5 [95% confidence interval, 2.44-304.54]; P = .01), whereas treatment with androgens did not. Review of limited numbers of liver biopsies and autopsy material showed a cholestatic pattern of liver injury, with progressive fibrosis, in the majority of patients. Occurrence in cases without transplant as well as cases with transplant argues against a potential diagnosis of atypical liver graft-versus-host disease. Limited data regarding therapy suggest no benefit from treatment with steroids or other immune suppressive medications or ursodeoxycholic acid. Our data show that liver disease is common in patients with FA, and because most children with FA now reach adulthood, end-stage liver disease in young adulthood means systematic testing of potential therapies is urgently needed.

Predictors of outcomes in hematopoietic cell transplantation for Fanconi anemia.

Allogeneic hematopoietic cell transplantation (HCT) remains the only cure for the hematologic manifestations of Fanconi anemia (FA). We performed retrospective predictor analyses for HCT outcomes in FA for pediatric and young adult patients transplanted between 2007 and 2020 across three large referral institutions. Eighty-nine patients, 70 with bone marrow failure +/- cytogenetic abnormalities, 19 with MDS/AML, were included. Five-year overall survival (OS) was 83.2% and event-free survival (EFS) was 74%. Age ≥19, HLA mismatch and year of HCT were multivariable predictors (MVPs) for OS, EFS and treatment-related mortality (TRM). In the pediatric group, TCD was a borderline MVP (P = 0.059) with 5-year OS of 73.0% in TCD vs. 100% for T-replete HCT. The cumulative incidence of day 100 grade II-IV aGvHD and 5-year cGvHD were 5.6% and 4.6%, respectively. Relapse in the MDS/AML subgroup occurred in 4 patients (16%). Graft failure was seen in 9 patients (TCD 6/37 [16%]; T-replete 3/52 [5.7%]). Six patients developed malignancy after HCT. Survival chances after HCT for FA are excellent and associated with high engrafted survival and low toxicity. Age ≥19, HLA mismatch, year of transplant and 'TCD in the <19 years group' (although borderline) were found to be negative predictors for survival.

T-cell depleted allogeneic hematopoietic stem cell transplant for the treatment of Fanconi anemia and MDS/AML.

The only curative approach for myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML) arising in patients with Fanconi anemia (FA) is allogeneic hematopoietic stem cell transplantation (HCT); however, HCT approaches are inconsistent and limited data on outcomes exist. We retrospectively evaluated outcomes of thirty patients with FA and MDS/AML who underwent first allogeneic HCT with a T-cell depleted (TCD) graft at our institution. Patients were transplanted on successive protocols with stepwise changes in cytoreduction and GVHD prophylaxis. All but two patients (93%) experienced durable hematopoietic engraftment. With median follow-up of 8.7 years, 5-year OS was 66.8% and DFS 53.8%. No significant differences in survival were found in patients with high-risk prognostic features (age ≥20 years, AML diagnosis, alternative donor graft) or when stratified by conditioning regimen. The 5-year cumulative incidences of relapse and NRM were 24.3% and 21.9%, respectively. NRM was higher in patients ≥20 years at HCT but did not otherwise differ. We herein demonstrate promising outcomes following allogeneic HCT for patients with FA and MDS/AML using TCD grafts, particularly in a cohort of high-risk patients with 50% ≥20 years and a majority receiving mismatched grafts. Future prospective studies are needed to compare this approach with other HCT platforms.

Management of Fanconi anemia beyond childhood.

Fanconi anemia (FA) has long been considered a severe inherited bone marrow failure (BMF) disorder of early childhood. Thus, management of this multisystem disorder has previously been unfamiliar to many hematologists specializing in the care of adolescents and young adults (AYA). The increased diagnosis of FA in AYA patients, facilitated by widely available germline genomic testing, improved long-term survival of children with FA following matched sibling and alternative donor hematopoietic stem cell transplantation (HSCT) performed for BMF, and expanding need in the near future for long-term monitoring in patients achieving hematologic stabilization following ex vivo gene therapy are all reasons why management of FA in AYA populations deserves specific consideration. In this review, we address the unique challenges and evidence-based practice recommendations for the management of AYA patients with FA. Specific topics addressed include hematologic monitoring in AYA patients yet to undergo HSCT, management of myeloid malignancies occurring in FA, diagnosis and management of nonhematologic malignances and organ dysfunction in AYA patients with FA, and evolving considerations for the long-term monitoring of patients with FA undergoing gene therapy.

Graft-versus-host disease and other cutaneous manifestations in pediatric patients transplanted for Fanconi anemia.

OBJECTIVE: The aim of this study was to elaborate a specific protocol for the assessment and early identification of skin lesions in pediatric patients with Fanconi anemia undergoing hematopoietic stem cell transplantation. METHODS: This is a longitudinal, retrospective, and descriptive study. The medical records of 136 pediatric patients with Fanconi anemia who underwent hematopoietic stem cell transplantation between 2008 and 2018 at the Clinical Hospital of the Federal University of Paraná were reviewed. A specific protocol was created for data collection, which included age, sex, skin color, age at diagnosis of Fanconi anemia, transplantation data, family history of consanguinity, and pre- and post-transplant complications. In addition, the data included the presence of graft-versus-host disease of the skin and other organs, its classification, type of lesion, location, and also skin lesions not related to graft-versus-host disease. RESULTS: Among the skin manifestations in pre-transplant period, café-au-lait spots stood out (32.4%). At least one organ was affected by graft-versus-host disease in 55.1% of patients; the most common involvement being the mouth, followed by the skin. Rash and erythema were the most frequently observed cutaneous manifestations of graft-versus-host disease. CONCLUSION: A high prevalence of cutaneous manifestations of the disease was observed, as well as cutaneous manifestations of graft-versus-host disease. The protocol developed gathers relevant and standardized information for the follow-up of patients with Fanconi anemia undergoing hematopoietic stem cell transplantation, ensuring greater reliability of the information, and its implementation will allow the prospective evaluation of patients.

RUNX1 mutations mitigate quiescence to promote transformation of hematopoietic progenitors in Fanconi anemia.

Many inherited bone marrow failure syndromes (IBMFSs) present a high risk of transformation to myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). During transformation of IBMFSs, hematopoietic stem and progenitor cells (HSPCs) with poor fitness gain ectopic, dysregulated self-renewal secondary to somatic mutations via undefined mechanisms. Here, in the context of the prototypical IBMFS Fanconi anemia (FA), we performed multiplexed gene editing of mutational hotspots in MDS-associated genes in human induced pluripotent stem cells (iPSCs) followed by hematopoietic differentiation. We observed aberrant self-renewal and impaired differentiation of HSPCs with enrichment of RUNX1 insertions and deletions (indels), generating a model of IBMFS-associated MDS. We observed that compared to the failure state, FA MDS cells show mutant RUNX1-mediated blunting of the G1/S cell cycle checkpoint that is normally activated in FA in response to DNA damage. RUNX1 indels also lead to activation of innate immune signaling, which stabilizes the homologous recombination (HR) effector BRCA1, and this pathway can be targeted to abrogate viability and restore sensitivity to genotoxins in FA MDS. Together, these studies develop a paradigm for modeling clonal evolution in IBMFSs, provide basic understanding of the pathogenesis of MDS, and uncover a therapeutic target in FA-associated MDS.

Beyond current treatment of Fanconi Anemia: What do advances in cell and gene-based approaches offer?

Fanconi anemia (FA) is a rare inherited disorder that mainly affects the bone marrow. This condition causes decreased production of all types of blood cells. FA is caused by a defective repair of DNA interstrand crosslinks and to date, mutations in over 20 genes have been linked to the disease. Advances in science and molecular biology have provided new insight between FA gene mutations and the severity of clinical manifestations. Here, we will highlight the current and promising therapeutic options for this rare disease. The current standard treatment for FA patients is hematopoietic stem cell transplantation, a treatment associated to exposure to radiation or chemotherapy, immunological complications, plus opportunistic infections from prolonged immune incompetence or increased risk of morbidity. New arising treatments include gene addition therapy, genome editing using CRISPR-Cas9 nuclease, and hematopoietic stem cell generation from induced pluripotent stem cells. Finally, we will also discuss the revolutionary developments in mRNA therapeutics as an opportunity for this disease.

Gene therapy restores the transcriptional program of hematopoietic stem cells in Fanconi anemia.

Clinical trials have shown that lentiviral-mediated gene therapy can ameliorate bone marrow failure (BMF) in nonconditioned Fanconi anemia (FA) patients resulting from the proliferative advantage of corrected FA hematopoietic stem and progenitor cells (HSPC). However, it is not yet known if gene therapy can revert affected molecular pathways in diseased HSPC. Single-cell RNA sequencing was performed in chimeric populations of corrected and uncorrected HSPC co-existing in the BM of gene therapy-treated FA patients. Our study demonstrates that gene therapy reverts the transcriptional signature of FA HSPC, which then resemble the transcriptional program of healthy donor HSPC. This includes a down-regulated expression of TGF-ß and p21, typically up-regulated in FA HSPC, and upregulation of DNA damage response and telomere maintenance pathways. Our results show for the first time the potential of gene therapy to rescue defects in the HSPC transcriptional program from patients with inherited diseases; in this case, in FA characterized by BMF and cancer predisposition.

Gonadal function in pediatric Fanconi anemia patients treated with hematopoietic stem cell transplant.

Gonadal dysfunction and reduced fertility are clinical manifestations well described in patients with Fanconi anemia (FA) and following hematopoietic stem cell transplantation (HSCT). It is difficult to differentiate gonadal dysfunction from the primary disease itself or from HSCT procedures. Therefore, it is important to manage expectations about gonadal failure and infertility for all patients with FA, regardless of the HSCT status. We performed a retrospective analysis of 98 pediatric patients with FA who were transplanted between July 1990 and June 2020 to evaluate the incidence of gonadal dysfunction in female and male patients with FA. New-onset premature ovarian insufficiency (POI) was diagnosed in a total of 30 (52.6%) patients. Follicle-stimulating hormone and luteinizing hormone levels were increased in patients diagnosed with POI. Anti- Mullerian hormone levels declined in POI patients after HSCT (r2=0.21; P=0.001). Twenty (48.8%) male patients were diagnosed with testicular failure. Follicle-stimulating hormone levels increased after HSCT even in patients without testicular failure (r2=0.17; P=0.005). Inhibin B levels decreased over time after HSCT in patients with testicular failure (r2=0.14; P=0.001). These data indicate brisk decline in already impaired gonadal function in transplanted children with FA.

Successful sequential liver and hematopoietic stem cell transplantation in a patient with Fanconi anemia.

BACKGROUND: In Fanconi anemia bone marrow failure is the major cause of morbidity and mortality and hematopoietic stem cell transplantation represents the only curative treatment. Liver disease, in terms of elevated liver function tests, as well as benign and malignant liver tumors, occurs especially in case of androgen treatment. We report a unique case of a child with Fanconi anemia with FANCD2 mutation who developed neonatal cryptogenic liver cirrhosis and bone marrow failure. The child successfully underwent sequential liver transplantation and hematopoietic stem cell transplantation in the first 2 years of life. Nineteen months after hematopoietic stem cell transplantation and 30 months after liver transplantation, the patient is clinically well with normal hematopoietic function and excellent liver function. CONCLUSION: This is the first FA patient who successfully received sequential LT and HSCT highlighting that successful sequential transplantation is feasible in Fanconi anemia patients.

[Chinese expert consensus on the diagnosis and treatment of Fanconi anemia (version 2022)].

Fanconi anemia (FA) is an autosomal recessive or X-linked hereditary bone marrow failure disease, in which mutations or deletions of FA-related genes lead to abnormalities in DNA repairment after damage and DNA cross-linking repair. The most common mutation genes include FANCA, FANCC, FANCG, FANCE and FANCF. FA is a disorder with high phenotypic and genotypic heterogeneity and mainly manifests as congenital somatic dysplasia, progressive cytopenia and increased risk of malignant tumors. In recent years, the survival of FA patients has greatly improved with the progress of FA management strategy and treatment. In order to better guide the clinical practice of doctors in China, the Red Blood Cell Disease (Anemia) Group of Chinese Society of Hematology of the Chinese Medical Association reached the"Chinese expert consensus on the diagnosis and treatment of Fanconi anemia (version 2022)"by widely collecting experts' suggestions and referring to the latest literature of FA, aiming to further standardize the diagnosis and treatment of FA in China.

[STEM CELL TRANSPLANTATIONS FOR PATIENTS WITH FANCONI ANEMIA: AN ISRAELI TERTIARY CENTER EXPERIENCE].

INTRODUCTION: Fanconi anemia (FA) is a rare genetic syndrome characterized by increased chromosomal breakage, congenital anomalies, bone marrow failure and an increased tendency to develop malignancies. Allogeneic hematopoietic stem cell transplantation (HSCT) is the only curative treatment for bone marrow failure and the hematologic malignancies these patients develop. Given the sensitivity of FA patients to chemotherapy and radiation, as to the clinical symptoms of graft versus host disease (GvHD), HSCT in these patients is challenging. Since the mid-nineties, HSCT for FA patients is performed in our center by using the fludarabine based reduced-intensity protocol. AIMS: To summarize the results of HSCT for patients with FA using a fludarabine based reduced-intensity conditioning regimen at the Hadassah Medical Center. METHODS: This retrospective research is based on the collection and analysis of clinical and laboratory data from the medical records of patients. RESULTS: Since June 1996 up till February 2020, 39 patients with FA underwent 43 HSCTs with a fludarabine based protocol at the Hadassah Medical Center. Four patients required a second transplant due to primary engraftment failure. Nine patients (23%) suffered from acute GvHD, four of them severe. Eight patients (20%) developed chronic GvHD, two with an extensive and debilitating disease. Thirty-three (85%) of the patients survived and six died, five shortly after the transplant, and one twenty years later from malignancy. CONCLUSIONS: Our results show high survival rates with low rates of engraftment failure and reasonable rates of GvHD. DISCUSSION: As of today, there is an effective and safe treatment for patients with FA who require HSCT by using a fludarabine-based reduced-intensity conditioning regimen, with high survival rates and few complications.

Genotype-phenotype and outcome associations in patients with Fanconi anemia: the National Cancer Institute cohort.

Fanconi anemia (FA) is caused by pathogenic variants in the FA/BRCA DNA repair pathway genes, and is characterized by congenital abnormalities, bone marrow failure (BMF) and increased cancer risk. We conducted a genotype-phenotype and outcomes study of 203 patients with FA in our cohort. We compared across the genes, FA/BRCA DNA repair pathways (upstream, ID complex and downstream), and type of pathogenic variants (hypomorphic or null). We explored differences between the patients evaluated in our clinic (clinic cohort) and those who provided data remotely (field cohort). Patients with variants in upstream complex pathway had less severe phenotype [lacked VACTERL-H (Vertebral, Anal, Cardiac, Trachea-esophageal fistula, Esophageal/duodenal atresia, Renal, Limb, Hydrocephalus) association and/or PHENOS (Pigmentation, small-Head, small-Eyes, Neurologic, Otologic, Short stature) features]. ID complex was associated with VACTERL-H. The clinic cohort had more PHENOS features than the field cohort. PHENOS was associated with increased risk of BMF, and VACTERL-H with hypothyroidism. The cumulative incidence of severe BMF was 70%, solid tumors (ST) 20% and leukemia 6.5% as the first event. Head and neck and gynecological cancers were the most common ST, with further increased risk after hematopoietic cell transplantation. Among patients with FANCA, variants in exons 27-30 were associated with higher frequency of ST. Overall median survival was 37 years; patients with leukemia or FANCD1/BRCA2 variants had poorest survival. Patients with variants in the upstream complex had better survival than ID or downstream complex (p=0.001 and 0.016, respectively). FA is phenotypically and genotypically heterogeneous; detailed characterization provides new insights towards understanding this complex syndrome and guiding clinical management.