I graduated from Wuerzburg University Medical School (Germany) in 2010. My MD research and thesis was evaluating the effect of reactive oxygen species on circulating angiogenic cells and endothelial function in patients with coronary artery disease, which proved that certain drugs that are believed to improve blood flow to the heart are creating oxidative stress which worsens endothelial dysfunction and impairs the function of angiogenic cells.
I underwent training in Pediatrics in the residency program at the Children's Hospital of the Ludwig Maximilian University (LMU) in Munich and started specialization in Pediatric Hematology, Oncology and Stem Cell Transplantation. Since 2015 I am board certified in Pediatrics.
Honors & Awards
Research fellowship, Care-For-Rare Foundation (2016)
Poster Prize, Stanford Institute for Stem Cell Biology and Regenerative Medicine (2017)
Research fellowship, German Scientific Society (DFG) (2017-2018)
Meritorious Abstract Award, ASGCT Annual Meeting 2019, American Society of Gene & Cell Therapy (ASGCT) (2019)
Boards, Advisory Committees, Professional Organizations
Member, German Society for Pediatrics and Adolecent Medicine (DGKJ) (2012 - Present)
Member, American Society for Gene and Cell Therapy (ASGCT) (2017 - Present)
State Examination Medicine, Julius Maximilians Universitaet Wuerzburg (2010)
Doctor of Medicine (Dr. med.), Hanover Medical School (2011)
Pediatrics Board Registration, Ludwig Maximilians University, Munich, Pediatrics (2015)
Matthew Porteus, Postdoctoral Faculty Sponsor
Current Research and Scholarly Interests
I am interested in using state-of-the-art genome editing tools in primary human cells to develop novel treatment approaches for diseases with unmet needs. The Porteus lab has established efficient methods for targeted gene correction in human hematopoietic stem cells which we are using to develop treatments for patients with immunodeficiencies and hematologic diseases. In addition, I am using gene editing of immune cells to develop the next generation of CAR T cells for cancer immunotherapy.
Metabolic engineering generates a transgene-free safety switch for cell therapy.
Safeguard mechanisms can ameliorate the potential risks associated with cell therapies but currently rely on the introduction of transgenes. This limits their application owing to immunogenicity or transgene silencing. We aimed to create a control mechanism for human cells that is not mediated by a transgene. Using genome editing methods, we disrupt uridine monophosphate synthetase (UMPS) in the pyrimidine de novo synthesis pathway in cell lines, pluripotent cells and primary human T cells. We show that this makes proliferation dependent on external uridine and enables us to control cell growth by modulating the uridine supply, both in vitro and in vivo after transplantation in xenograft models. Additionally, disrupting this pathway creates resistance to 5-fluoroorotic acid, which enables positive selection of UMPS-knockout cells. We envision that this approach will add an additional level of safety to cell therapies and therefore enable the development of approaches with higher risks, especially those that are intended for limited treatment durations.
View details for DOI 10.1038/s41587-020-0580-6
View details for PubMedID 32661439
Tuning the Antigen Density Requirement for CAR T Cell Activity.
Insufficient reactivity against cells with low antigen density has emerged as an important cause of CAR resistance. Little is known about factors that modulate the threshold for antigen recognition. We demonstrate that CD19 CAR activity is dependent upon antigen density and the CAR construct in axicabtagene-ciloleucel (CD19-CD28z) outperforms that in tisagenlecleucel (CD19-4-1BBz) against antigen low tumors. Enhancing signal strength by including additional ITAMs in the CAR enables recognition of low antigen density cells, while ITAM deletions blunt signal and increase the antigen density threshold. Further, replacement of the CD8 hinge-transmembrane (H/T) region of a 4-1BBz CAR with a CD28-H/T lowers the threshold for CAR reactivity despite identical signaling molecules. CARs incorporating a CD28-H/T demonstrate a more stable and efficient immunological synapse. Precise design of CARs can tune the threshold for antigen recognition and endow 4-1BBz-CARs with enhanced capacity to recognize antigen low targets while retaining a superior capacity for persistence.
View details for DOI 10.1158/2159-8290.CD-19-0945
View details for PubMedID 32193224
Genome editing of donor-derived T-cells to generate allogenic chimeric antigen receptor-modified T cells: Optimizing αβ T cell-depleted haploidentical hematopoietic stem cell transplantation.
Allogeneic hematopoietic stem cell transplantation is an effective therapy for high-risk leukemias. In children, graft manipulation based on the selective removal of αβ T cells and B cells has been shown to reduce the risk of acute and chronic graft-versus-host disease, thus allowing the use of haploidentical donors which expands the population that allogeneic hematopoietic stem cell transplantation can be used in. Leukemic relapse, however, remains a problem. T cells expressing chimeric antigen receptors can potently eliminate leukemia, including in the central nervous system. We hypothesized that by modifying donor αβ T cells to simultaneously express a CD19-specific chimeric antigen receptors and inactivating the T cell receptor by genome editing, we could create a therapy that enhances the anti-leukemic efficacy of the stem cell transplant without increasing the risk of graft-versus-host disease. Using genome editing with Cas9 ribonucleoprotein and adeno-associated virus serotype 6, we integrate a CD19-specific chimeric antigen receptor in-frame into the TRAC locus. Greater than 90% of cells lost TCR expression, while >75% expressed the CAR. The product was further purified to ultimately have less than 0.05% residual TCR+ cells. In vitro, the CAR T cells efficiently eliminated target cells and produced high cytokine levels when challenged with CD19+ leukemia cells. In vivo, the gene modified T cells eliminated leukemia without causing xenogeneic graft-versus-host disease in a xenograft model. Gene editing was highly specific with no evidence of off-target effects. These data support the concept that the addition of αβ T cell-derived, genome edited T cells expressing CD19-specific chimeric antigen receptors could enhance the anti-leukemic efficacy of αβ T cell-depleted haploidentical hematopoietic stem cell transplantation without increasing the risk of graft-versus-host disease.
View details for DOI 10.3324/haematol.2019.233882
View details for PubMedID 32241852
- T-cell replete haploidentical bone marrow transplantation and post-transplant cyclophosphamide for patients with inborn errors HAEMATOLOGICA 2019; 104 (10): E478–E482
genome editing of graft-derived T cells for post-transplant immunotherapy in combination with TCR alpha beta(+)/CD19(+)-depleted haploidentical HSCT
NATURE PUBLISHING GROUP. 2019: 183–84
View details for Web of Science ID 000487707800198
- Highly Efficient and Marker-free Genome Editing of Human Pluripotent Stem Cells by CRISPR-Cas9 RNP and AAV6 Donor-Mediated Homologous Recombination CELL STEM CELL 2019; 24 (5): 821-+
Can't Live without "U": Genetic Engineering of UMPS to Create Auxotrophy in Human Cells
CELL PRESS. 2019: 454
View details for Web of Science ID 000464381005072
- Gene correction for SCID-X1 in long-term hematopoietic stem cells NATURE COMMUNICATIONS 2019; 10
Genome Editing Using CRISPR/Cas9 and rAAV6 to Functionally Correct Wiskott-Aldrich Syndrome in Human HSPCs
CELL PRESS. 2018: 376–77
View details for Web of Science ID 000435342205038
A high-fidelity Cas9 mutant delivered as a ribonucleoprotein complex enables efficient gene editing in human hematopoietic stem and progenitor cells.
2018; 24 (8): 1216–24
Translation of the CRISPR-Cas9 system to human therapeutics holds high promise. However, specificity remains a concern especially when modifying stem cell populations. We show that existing rationally engineered Cas9 high-fidelity variants have reduced on-target activity when using the therapeutically relevant ribonucleoprotein (RNP) delivery method. Therefore, we devised an unbiased bacterial screen to isolate variants that retain activity in the RNP format. Introduction of a single point mutation, p.R691A, in Cas9 (high-fidelity (HiFi) Cas9) retained the high on-target activity of Cas9 while reducing off-target editing. HiFi Cas9 induces robust AAV6-mediated gene targeting at five therapeutically relevant loci (HBB, IL2RG, CCR5, HEXB, and TRAC) in human CD34+ hematopoietic stem and progenitor cells (HSPCs) as well as primary T cells. We also show that HiFi Cas9 mediates high-level correction of the sickle cell disease (SCD)-causing p.E6V mutation in HSPCs derived from patients with SCD. We anticipate that HiFi Cas9 will have wide utility for both basic science and therapeutic genome-editing applications.
View details for PubMedID 30082871
IPEX due to an exon 7 skipping FOXP3 mutation with autoimmune diabetes mellitus cured by selective TReg cell engraftment.
Clinical immunology (Orlando, Fla.)
2018; 191: 52–58
Immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome is a rare inherited disorder leading to severe organ-specific autoimmunity. IPEX is caused by hemizygous mutations in FOXP3, which codes for a master transcription factor of regulatory T (TReg) cell development and function. We describe a four-year-old boy with typical but slightly delayed-onset of IPEX with autoimmune diabetes mellitus, enteropathy, hepatitis and skin disease. We found the unreported FOXP3 splice site mutation c.816+2T>A that leads to the loss of leucine-zipper coding exon 7. RNA-Seq revealed that FOXP3Δ7 leads to differential expression of FOXP3 regulated genes. After myeloablative conditioning the patient underwent allogeneic HSCT from a matched unrelated donor. HSCT led to the resolution of all IPEX symptoms including insulin requirement despite persisting autoantibody levels. After initial full donor engraftment nearly complete autologous reconstitution was documented, but donor-derived TReg cells persisted with a lineage-specific chimerism of >70% and the patient remained in clinical remission.
View details for PubMedID 29567430
- Allogeneic stem cell transplantation in adolescents and young adults with primary immunodeficiencies. The journal of allergy and clinical immunology. In practice 2017; 6 (1): 298–301.e2
- MRD response in a refractory paediatric T-ALL patient through anti-programmed cell death 1 (PD-1) Ab treatment associated with induction of fatal GvHD. Bone marrow transplantation 2017; 52 (8): 1221–24
Reduced toxicity, myeloablative HLA-haploidentical hematopoietic stem cell transplantation with post-transplantation cyclophosphamide for sickle cell disease.
Annals of hematology
2017; 96 (8): 1373–77
Allogeneic hematopoietic stem cell transplantation (HSCT) offers the possibility of cure for sickle cell disease (SCD) patients. Unfortunately, the probability of finding an HLA-matched donor for SCD patients is low. HSCT from HLA-haploidentical donors using reduced intensity conditioning, unmanipulated bone marrow and post-transplantation cyclophosphamide (ptCy) has resulted in negligible toxicity but high rates of graft rejection. We hypothesized that combining ptCy with a myeloablative reduced toxicity conditioning including serotherapy to increase immune ablation would allow for better engraftment. In a pilot approach, we treated three patients with SCD (5, 8, and 20 years old) lacking a matched donor. All patients had severe disease-related complications despite standard treatment. They received unmanipulated bone marrow from parental HLA-haploidentical donors. Conditioning consisted of alemtuzumab 0.2 mg/kg/day on days -9 and -8, fludarabine 30 mg/m2/day on days -7 to -3, treosulfan 14 g/m2/day on days -7 to -5, thiotepa 2 × 5 mg/kg/day on day -4, and cyclophosphamide 14.5 mg/kg/day on days -3 and -2. GVHD prophylaxis was performed using cyclophosphamide 2 × 50 mg/kg on days +3 and +4 and mycophenolate mofetil, tacrolimus from day +5. After a follow-up of 11, 14, and 30 months, all three patients are alive and well, off immunosuppression, and without symptoms of SCD. One patient experienced mild skin GVHD grade I, none showed chronic GVHD. Asymptomatic CMV reactivation was seen in two patients. HLA-haploidentical HSCT can extend the donor pool for patients with SCD. Whether intensification of the conditioning regimen and intensive immunosuppression leads to improvement in engraftment rates while still allowing a favorable toxicity profile deserves further investigation.
View details for PubMedID 28573314
Maturation of Platelet Function During Murine Fetal Development In Vivo.
Arteriosclerosis, thrombosis, and vascular biology
2017; 37 (6): 1076–86
Platelet function has been intensively studied in the adult organism. However, little is known about the function and hemostatic capacity of platelets in the developing fetus as suitable in vivo models are lacking.To examine fetal platelet function in vivo, we generated a fetal thrombosis model and investigated light/dye-induced thrombus formation by intravital microscopy throughout gestation. We observed that significantly less and unstable thrombi were formed at embryonic day (E) 13.5 compared with E17.5. Flow cytometry revealed significantly lower platelet counts in E13.5 versus E17.5 fetuses versus adult controls. In addition, fetal platelets demonstrated changed activation responses of surface adhesion molecules and reduced P-selectin content and mobilization. Interestingly, we also measured reduced levels of the integrin-activating proteins Kindlin-3, Talin-1, and Rap1 during fetal development. Consistently, fetal platelets demonstrated diminished spreading capacity compared with adults. Transfusion of adult platelets into the fetal circulation led to rapid platelet aggregate formation even in young fetuses. Yet, retrospective data analysis of a neonatal cohort demonstrated no correlation of platelet transfusion with closure of a persistent ductus arteriosus, a process reported to be platelet dependent.Taken together, we demonstrate an ontogenetic regulation of platelet function in vivo with physiologically low platelet numbers and hyporeactivity early during fetal development shedding new light on hemostatic function during fetal life.
View details for PubMedID 28428216
Impairment of endothelial progenitor cell function and vascularization capacity by aldosterone in mice and humans.
European heart journal
2011; 32 (10): 1275–86
Hyperaldosteronism is associated with vascular injury and increased cardiovascular events. Bone marrow-derived endothelial progenitor cells (EPCs) play an important role in endothelial repair and vascular homeostasis. We hypothesized that hyperaldosteronism impairs EPC function and vascularization capacity in mice and humans.We characterized the effects of aldosterone and mineralocorticoid receptor (MR) blockade on EPC number and function as well as vascularization capacity and endothelial function. Treatment of human EPC with aldosterone induced translocation of the MR and impaired multiple cellular functions of EPC, such as differentiation, migration, and proliferation in vitro. Impaired EPC function was rescued by pharmacological blockade or genetic ablation of the MR. Aldosterone protein kinase A (PKA) dependently increased reactive oxygen species formation in EPC. Aldosterone infusion in mice impaired EPC function, EPC homing to vascular structures and vascularization capacity in a MR-dependent but blood pressure-independent manner. Endothelial progenitor cells from patients with primary hyperaldosteronism compared with controls of similar age displayed reduced migratory potential. Impaired EPC function was associated with endothelial dysfunction. MR blockade in patients with hyperaldosteronism improved EPC function and arterial stiffness.Endothelial progenitor cells express a MR that mediates functional impairment by PKA-dependent increase of reactive oxygen species. Normalization of EPC function may represent a novel mechanism contributing to the beneficial effects of MR blockade in cardiovascular disease prevention and treatment.
View details for DOI 10.1093/eurheartj/ehq254
View details for PubMedID 20926363
View details for PubMedCentralID PMC3094546
Organic nitrates differentially modulate circulating endothelial progenitor cells and endothelial function in patients with symptomatic coronary artery disease.
Antioxidants & redox signaling
2011; 15 (4): 925–31
Symptomatic coronary artery disease (CAD) is usually treated with organic nitrates. Endothelial progenitor cells (EPCs) are a circulating cell population participating in vascular homeostasis in a nitric oxide-dependent manner. We investigated the effects of the nitric oxide donors isosorbide dinitrate (ISDN) and pentaerythritol tetranitrate (PETN) on EPC and endothelial function in patients with symptomatic CAD. We randomized 36 patients with angiographically proven CAD to treatment with either ISDN (40 mg retarded release orally two times per day; n = 18) or PETN (80 mg orally two times per day; n = 18) for 14 days (clinical trial number: NCT01030367). PETN treatment substantially increased numbers of circulating CD34(+)/KDR(+) EPCs (p = 0.02), whereas no effects were observed in patients treated with ISDN. EPC function assessed by formation of endothelial colonies was enhanced by twofold (p = 0.04) in patients treated with PETN. No changes were observed after ISDN treatment. Endothelial function, assessed by peripheral arterial tonometry, remained unchanged during PETN treatment, but was significantly impaired in patients treated with ISDN. Treatment of symptomatic CAD patients with PETN for 14 days significantly increased levels of circulating EPC and improved markers for EPC function, whereas ISDN was without effects on EPCs and worsened endothelial function.
View details for DOI 10.1089/ars.2010.3503
View details for PubMedID 20812862