Lentiviral Vector Gene Therapy Protects XCGD Mice From Acute Staphylococcus aureus Pneumonia and Inflammatory Response

Abstract

Chronic granulomatous disease (CGD) is a primary immunodeficiency due to a deficiency in one of the subunits of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex. CGD patients are characterized by an increased susceptibility to bacterial and fungal infections, and to granuloma formation due to the excessive inflammatory responses. Several gene therapy approaches with lentiviral vectors have been proposed but there is a lack of in vivo data on the ability to control infections and inflammation. We set up a mouse model of acute infection that closely mimic the airway infection in CGD patients. It involved an intratracheal injection of a methicillin-sensitive reference strain of S. aureus. Gene therapy, with hematopoietic stem cells transduced with regulated lentiviral vectors, restored the functional activity of NADPH oxidase complex (with 20-98% of dihydrorhodamine positive granulocytes and monocytes) and saved mice from death caused by S. aureus, significantly reducing the bacterial load and lung damage, similarly to WT mice even at low vector copy number. When challenged, gene therapytreated XCGD mice showed correction of proinflammatory cytokines and chemokine imbalance at levels that were comparable to WT. Examined together, our results support the clinical development of gene therapy protocols using lentiviral vectors for the protection against infections and inflammation.

High efficiency gene correction in hematopoietic cells by donor-template-free CRISPR/Cas9 genome editing

“…Gene-replacement therapy in hematopoietic stem cells requires integrating vectors to ensure long term gene expression. Although more recently non-viral transposon vectors have been used [39][40][41] , the vast majority of clinical trials employed γ-retroviral 53,54 and lentiviral vectors [55][56][57] . All integrating vectors insert semirandomly throughout the genome, each having a specific integration pattern dictated by distinct features of different genomic regions 39,58,59 .…”

Staphylococcus aureus Strain Newman D2C Contains Mutations in Major Regulatory Pathways That Cripple Its Pathogenesis

“…In addition, Newman has served as a valuable measuring stick for assessing the therapeutic efficacy of agents designed to protect the host from S. aureus infection. The importance of Newman has been demonstrated by multiple studies in animals that have shown reduced S. aureus-caused lethality in cohorts treated with a number of different therapies (12,(14)(15)(16)(17)(18)(19)(20). In our own attempts to develop a therapeutic agent to treat S. aureus infection, we discovered significant inconsistencies in the virulence profiles of Newman strains and a derivative thereof, Newman D2C.…”

“…Concomitant with the demand to develop an effective antistaphylococcal therapy is the need to select the most applicable model(s) for studying disease caused by S. aureus. S. aureus strain Newman (here referred to as Newman) is a commonly used model strain for studying S. aureus pathogenesis (3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13) and in turn has been extensively used to examine the therapeutic efficacy of agents designed to treat S. aureus infection (12,(14)(15)(16)(17)(18)(19)(20). Newman was originally isolated from a human patient in 1952 (21,22) and has since been a fixture in the investigation of S. aureus pathogenesis due in large part to its strong virulence in both animal models and human ex vivo systems.…”

Hematopoietic Tumors in a Mouse Model of X-linked Chronic Granulomatous Disease after Lentiviral Vector-Mediated Gene Therapy

“…26 GT mediated by MSP.Gp91.2T-transduced HSPCs was effective in protecting XCGD mice from S. aureus pulmonary infection. 27 Despite the extensive investigation of the efficacy of HSPC GT for XCGD in cells and mouse model systems, there is little information about long-term (>5 months) safety and efficacy in non-clinical studies with LV-mediated GT for XCGD. 24,25 Also, insertional genotoxicity has been conducted by transplanting LV-transduced Lin À cells isolated from CD45.2 C57BL/6 mice into CD45.1 recipients followed up to 26 weeks.…”

“… 22 , 23 Different LVs encoding human Gp91 phox under the control of a myeloid-specific promoter (MSP) have been developed, and their efficacy was tested in preclinical models. 24 , 25 , 26 , 27 Transduction of mouse BM-derived XCGD lineage-negative (Lin − ) cells and human XCGD CD34 + HSPCs with an LV encoding Gp91 phox under a synthetic chimeric myeloid promoter (pCCLchimGp91 s ) drove higher Gp91 phox expression and NADPH activity in myelomonocytic cells (i.e., neutrophils) and myeloid cell progenitors compared with lymphoid cells in vitro and in vivo . 24 Therapeutic efficacy and no evidence of toxicity were reported in a preclinical study performed according to Good Laboratory Practice (GLP) using a clinical-grade G1XCGD LV to transduce human CD34 + XCGD cells and mouse XCGD Lin − cells.…”