Modified vectors for XLA gene therapy

Abstract

This disclosure relates generally to vectors, typically viral vectors, useful for the treatment of X-linked agammaglobulinemia (XLA).

Description

[0001]TITLE OF THE INVENTION MODIFIED VECTORS FOR XLA GENE THERAPY RELATED APPLICATIONS This application claims priority to US Provisional Application No. 63 / 633225 entitled “MODIFIED VECTORS FOR XLA GENE THERAPY” filed 12 April 2024, the contents of which are incorporated herein by reference in their entirety. FIELD OF THE INVENTION This disclosure relates generally to vectors, typically viral vectors, useful for the treatment of X-linked agammaglobulinemia (XLA). BACKGROUND OF THE INVENTION X-linked agammaglobulinemia (XLA) is a rare X-linked genetic disorder that affects the body's ability to fight infection. In XLA, B call maturation is compromised manifesting as a complete or near-complete lack of serum immunoglobulins and an inability to generate a protective antibody responses to pathogen challenge. Patients with XLA have normal pre-B cell populations in their bone marrow but these cells fail to mature and enter the circulation. XLA is characterized by recurrent bacterial infections, commonly otitis, conjunctivitis, sinus and pulmonary infections, diarrhea, and skin infections, in affected individuals from the first two years of life. XLA is also associated with increased risk of sepsis, including sudden death due to bacterial sepsis, pneumonia, empyema, meningitis, cellulitis, chronic and systemic enterovirus infection, chronic inflammatory bowel disease and increased risk of malignancies such as colon cancer. XLA results from mutations in the Bruton's tyrosine kinase (BTK) gene. BTK is a cytoplasmic tyrosine kinase expressed in B cells and myeloid cells, contributing to normal cellular functions in both lineages. BTK plays an essential role in B cell maturation, regulating early B cell development as well as mature B cell activation, signaling and survival. Dysregulation of BTK expression can lead to developmental blockades and autoimmunity. Mutations in the BTK gene leading to XLA are associated with BTK deficiency, directly contributing to the failure of afflicted individuals to generate mature B cells and the inability of B cells to respond to antigenic challenges. XLA therapy typically includes immunoglobulin replacement and the use of targeted antimicrobial agents. For example, current treatment options can include lifelong pooled human immunoglobulin administration (IVIg or SCIg) every 3-4 weeks. However these treatments are expensive and can themselves lead to risk for infection and sudden death. In some cases, XLA subjects have been treated with stem cell transplantation without conditioning or using reduced intensity conditioning with variable outcomes. Despite the commonly employed therapies, XLA sufferers continue to be at increased risk for a range of morbid or life-threatening complications. There is a clear need for the development of new and improved therapeutic options to treat XLA. Gene therapy of XLA, in particular retroviral-mediated expression of BTK in B cells and myeloid cells derived from hematopoietic stem cells, has gained increased attention with the development of suitable mouse models of XLA (see, e.g. Singh et al., 2015, Molecular Therapy, 23:S93; Kerns et al., 2010, Blood 115:2146-2155). Lentiviral vectors incorporating a ubiquitous chromatin opening element (UCOE), capable of inducing sustained BTK transgene expression and of rescuing B cell development, have been developed (see WO 2018 / 195297, the disclosure of which is incorporated herein in its entirety). However the UCOE described in these vectors include cryptic splice sites which can induce alternative splicing of transgene RNA, lentiviral genomic RNA, or transcripts from a gene into which the vector genome integrates, resulting in truncated, non-functional transcripts. There is a need in the art for the development of vectors with improved safety profiles for use in gene therapy applications for the treatment of XLA. SUMMARY OF THE INVENTION Provided herein are vectors that are suitable for use in gene therapy of X-linked agammaglobulinemia (XLA). The vectors contain a modified ubiquitous chromatin opening element (UCOE) in which, relative to the UCOE present in XLA gene therapy vectors described in WO 2018 / 195297, cryptic splice acceptor sites have been inactivated and single nucleotide polymorphisms (SNPs) have been corrected to revert the sequence of the UCOE to the canonical sequence. As demonstrated herein, the vectors of the present disclosure not only exhibit a desirable safety profile, but also are therapeutically effective. Despite modifications of the vector sequence to ameliorate safety concerns associated with potential alternative splicing events, the vectors of the present disclosure facilitate robust BTK expression in vivo. Indeed, the BTK expression facilitated by the vectors is sufficient to restore B cell function in XLA animal models to therapeutically effective levels. Thus, in one aspect of the invention there is provided a vector that is suitable for the treatment of X-linked agammaglobulinemia (XLA), or for inhibiting or ameliorating at least one symptom of XLA, the vector comprising: (a) a transgene expression cassette comprising a nucleotide sequence encoding Bruton's tyrosine kinase (BTK) operably linked to a promoter; (b) at least one regulatory or enhancer element; (c) a ubiquitous chromatin opening element (UCOE), wherein the UCOE comprises a nucleotide sequence at least about 90% identical to the sequence set forth in SEQ ID NO:1, and wherein the sequence of the UCOE comprises the following nucleotide substitutions relative to SEQ ID NO:1, at positions corresponding to those of SEQ ID NO:1: (i) a G residue at position 4; (ii) the sequence CGAAC at positions 347-351; (iii) the sequence CGCGC at positions 362-366; (iv) a T residue at position 385; (v) a G residue at position 496; (vi) the sequence CGCGC at positions 612-616; and (vii) a T residue at position 673. Optionally, the BTK nucleotide sequence encodes a polypeptide with the sequence of SEQ ID NO:3. The nucleotide sequence encoding BTK may be codon optimized for expression in humans. For example, the nucleotide sequence encoding BTK may comprise the sequence set forth in SEQ ID NO:4 or SEQ ID NO:5.' Optionally, the promoter in the transgene expression cassette is a B cell specific promoter. In a particular embodiment, the promoter is the BTK promoter, optionally comprising the sequence set forth in SEQ ID NO:6. In an exemplary embodiment, the transgene expression cassette comprises the sequence set forth in SEQ ID NO:7. In particular embodiments, the UCOE comprises the nucleotide sequence of SEQ ID NO:2, or a sequence at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98.5%, 99% or 99.5% identical thereto. In preferred embodiments the UCOE is at least about 0.7kb in length and comprises or consists of the nucleotide sequence of SEQ ID NO:2. In exemplary embodiments, the UCOE is in the forward orientation in the vector with respect to the transgene expression cassette. In exemplary embodiments, the UCOE is in the reverse orientation in the vector with respect to the transgene expression cassette. The UCOE and the transgene expression cassette may comprise the sequence set forth in SEQ ID NO:8. In exemplary embodiments, the at least one regulatory element may comprise an insulator, a Woodchuck Hepatitis Virus (WHV) Posttranscriptional Regulatory Element (WPRE), or a combination thereof, optionally wherein the regulatory element is located downstream of the transgene expression cassette. The vector may be, for example, a viral vector. The vector may be a retroviral vector. Preferably the vector is a lentiviral vector. In an exemplary embodiment, the vector comprises the sequence set forth in SEQ ID NO:20 or a sequence having at least about 90% sequence identity thereto, comprising the UCOE from position 2396 to 3068 of SEQ ID NO:20. In another exemplary embodiment, the vector comprises the sequence having at least about 90% sequence identity thereto, comprising the UCOE from position 2396 to 3068 of SEQ ID NO:20. Another aspect of the invention provides a cell comprising, or transduced with, a vector of the invention. By way of example, the cell may be a B cell, a myeloid cell or a hematopoietic stem cell. The hematopoietic stem cell may be, for example, a CD34+hematopoietic stem cell. Another aspect of the invention provides a method for promoting B cell survival, proliferation and / or differentiation in a subject in need thereof, the method comprising administering to the subject a vector or a cell of the invention, and optionally measuring B cell survival, proliferation and / or differentiation in the subject or in a biological sample obtained from the subject. In an embodiment, the method further comprises the step, prior to administration, of identifying the subject as one that would benefit from receiving a therapy to promote B cell survival, proliferation and / or differentiation. Optionally, the cell is administered by adoptive cell transfer. Another aspect of the invention provides a method of treating X-linked agammaglobulinemia (XLA) or inhibiting or ameliorating at least one symptom thereof, in a subject, the method comprising administering to the subject a vector or cell of the invention, and optionally measuring an improvement in the progression of XLA or at least one symptom thereof in the subject or in a biological sample obtained from the subject. In an embodiment, the method further comprises the step, prior to administration, of identifying the subject as one that would benefit from receiving a therapy to treat X-linked agammaglobulinemia (XLA) or to inhibit or ameliorate at least one symptom thereof. Optionally, the cell is administered by adoptive cell transfer. Also provided herein is the use of a vector or a host cell of the invention in the preparation of a medicament for promoting B cell survival, proliferation and / or differentiation, or the treatment of XLA or the inhibition or amelioration of at least one symptom thereof. BRIEF DESCRIPTION OF THE DRAWINGS Embodiments of the disclosure are described herein, by way of non-limiting example only, with reference to the following drawings. Figure 1 - alignment of a modified UCOE of the present invention (SEQ ID NO:2) and the unmodified ubiquitous chromatin opening element (UCOE) of SEQ ID NO:1. Figure 2 - schematic representation of exemplary lentiviral constructs described herein for expression of BTK. (A) LV XLA vector (#1419; LVXLA1) comprising UCOE.pBTK.coBTK cassette described in W02018 / 195297 (SEQ ID NO:33 of WO 2018 / 195297) comprising the unmodified UCOE sequence (SEQ ID NO:1 herein) with the CBX3 promoter in reverse direction relative to the transgene cassette. (B) Exemplary pRRL- XLA LV vector (# 3365) comprising the unmodified UCOE sequence (SEQ ID NO:1 herein) and the CBX3 promoter in forward orientation relative to the transgene cassette. (C) Exemplary pRRL-XLA vector (#3366) described herein comprising the modified UCOE of SEQ ID NO:2 herein, containing the CBX3 promoter in forward orientation relative to transgene cassette. (D) Exemplary TL20c-XLA vector (LVXLA2) comprising the modified UCOE of SEQ ID NO:2 herein, containing CBX3 promoter in forward orientation relative to the transgene cassette. In (A)-(D): RSV, Rous sarcoma virus promoter; R, HIV repeat region; U5, HIV long terminal repeat (LTR) U5 region; RRE, rev response element; cPPT, central poly purine tract; 0.7UCOE, 0.7 kb UCOE region; 0.7UCOE*, optimized (modified) 0.7 kb UCOE region; pBTK, 0.8 kb promoter region for BTK gene; coBTK, codon optimized cDNA sequence encoding BTK protein; WPRE, Woodchuck hepatitis virus post-transcriptional regulatory element; U3, partially deleted HIV LTR U3 region; SV40pA, polyadenylation signal from Simian virus 40; rbGpA, rabbit beta-globion polyadenylation signal. Figure 3 – Experimental timeline of lentiviral gene therapy study in mXLA mice Figure 4 – Comparison of percent B cell rescue using alternative LV constructs in mXLA gene therapy at 10-weeks post-transplant peripheral blood. Figure 5 – Comparison of B cell rescue using alternative LV constructs in mXLA gene therapy at primary transplant endpoint. (A) Reconstitution of B cell numbers in spleen (%B220+cells * total splenocyte count). (B) Percentage of B cells in bone marrow (BM) (%B220+of total lymphocyte gate). (C) Percentage of B cells in spleen (%B220+of total lymphocyte gate). Figure 6 – UCOE-containing vectors restore in vivo B cell responses to T- dependent immunogen and total IgG compared to mXLA mock. (A) Differences between experimental and mXLA mock cohorts for both the primary and secondary antibody responses. (B) Differences between experimental and mXLA mock cohorts. Figure 7 – Viral copy number (VCN) in input cells and in bone marrow and spleen from primary recipients. VCN determined following gDNA extraction and digital droplet PCR (ddPCR) from input cells (A), bone marrow (BM) cells (B) and spleen cells (C). Figure 8 – Methylation of BTKpBTK and lentiviral vectors with unmodified 0.7UCOE (SEQ ID NO:1) and modified 0.7UCOE (SEQ ID NO:2). Figure 9 – Schematic for secondary transplants of bone marrow cells into irradiated mXLA secondary recipient mice. Figure 10 – Lentiviral vectors with unmodified 0.7UCOE (SEQ ID NO:1) and modified 0.7UCOE (SEQ ID NO:2) mediate sustained BTK expression and B cell rescue in secondary mXLA recipients. (A and B) Percentage of B cells (B220+ of singlets) and % BTK+ B cells (B220+ BTK+) at 17 weeks post-transplant in bone marrow (BM). (C and D) Percentage of B cells (B220+ of singlets) and % BTK+ B cells (B220+ BTK+) at 17 weeks post-transplant in spleen. Figure 11 – Viral copy number (VCN) in bone marrow (BM) (A) and spleen (B) from secondary recipients as quantified by ddPCR to determine average number of vector copies per cell. Figure 12 – Experimental timeline of mouse humanization with LVXLA2- transduced human hematopoietic stem and progenitor cells (HSPCs). Figure 13 – Human CD45+ engraftment in recipient mice. Proportion of human CD45+ (hCD45+) cells within the lymphocyte gate among cells collected from the bone marrow (BM) (A) and spleen (SP) (B) of humanized mice 16-weeks post-transplant. HD, healthy donor. TDX, LVXLA2 vector. *, lower engraftment, partial dose given. **, no engraftment, unsuccessful RO transfer. Figure 14 – Lineage characterization of bone marrow cells. Proportion of hCD45 cells within the lymphocyte gate that are CD19+B cells (A), CD4 / CD8+ T cells (B) and CD33+ myeloid cells (C) in bone marrow of humanized mice 16-weeks post-transplant. HD = Healthy donor; TDX = LVXLA2 vector (as columns from left to right in graphs for each cell type: HD Mock, HD TDK, XLA Mock, XLA TDX). Figure 15 – LVXLA2 gene therapy alleviates pre-B cell developmental block in the bone marrow. (A) Proportion and (B) number of indicated B cell developmental subsets in the bone marrow of humanized mice 16-weeks post-transplant. HD = Healthy donor; TDX = LVXLA2 vector. Figure 16 – Lineage characterization of spleen cells at in vivo endpoint. Proportion of hCD45 cells within the lymphocyte gate that are (A) CD19+B cells, (B) CD4 / CD8+ T-cells, and (C) CD33+ myeloid cells in the spleens of humanized mice 16-weeks post-transplant. HD = Healthy donor; TDX = LVXLA2 vector. Figure 17 – LVXLA2 gene therapy restores mature human B cell compartments in the spleen of XLA humanized mice. (A) Proportion of Pre-B, transitional and mature naïve cells in the CD19+lymphocyte gate of the spleen cells, and (B) the absolute counts of indicated B cell subsets. Bars show the mean + / - SEM. HD = Healthy donor; TDX = LVXLA2 vector. (as columns from left to right in graphs for each cell type: HD Mock, HD TDK, XLA Mock, XLA TDX). Figure 18 – Viral copy number (VCN) of input LVXLA2-transduced human hematopoietic stem and progenitor cells (HSPC) and hematopoietic tissue at in vivo endpoint. (A) “input” VCN from LVXLA2-transduced healthy donor (HD) and XLA HSPC transplanted into NBSGW mice determined by ddPCR of gDNA 14-days post-transduction. Each point represents a single colony. Healthy donor (HD) data from input cells are from a separate, but representative experiment. (B and C) VCN from bone marrow and spleen, respectively, of humanized mice 16-weeks post-transplant determined by ddPCR. Each point represents a single humanized mouse. Average VCN for input cells, bone marrow and spleen are shown below graphs. Figure 19 - schematic of steps to generate and characterize a BTK knock-out B cell line.5 Figure 20 - Schematic of homology directed repair to knock in GFP in exon2 of BTK gene. The BTK TALEN cut site within the endogenous BTK allele is indicated with red arrow. GFP cDNA under the control of the MND promoter was inserted to replace most of BTK exon 2 by HDR as indicated. Alleles with successful HDR editing have disrupted (absent) BTK expression, but constitutive GFP expression. Figure 21 - BTK knock-out (KO) Ramos clone 1 expresses GFP but not BTK. BTK KO Ramos clone 1 cells were assessed by flow cytometry in comparison with the parental (unedited) Ramos B cell line, as indicated. Panels shown are GFP expression (column 2) and intracellular BTK (column 3) within the lymphocyte gate (column 1). The parental Ramos B cells were either unstained for BTK (top row) or stained for BTK (middle row). Figure 22 - BTK expression increases with lentiviral copy number in LVXLA2 transduced BTK KO Ramos clone-1 cells. (A and B) Flow plots show GFP and BTK expression of cells within the lymphocyte gate (columns 2, 3, and 1 for each condition, respectively). (A) Unedited Ramos B cells with (bottom row) or without (top row) BTK intracellular staining. (B) BTK-KO Ramos clone 1 cells treated with the indicated LVXLA2 and subsequently assessed for BTK expression via FACS-based BTK intracellular staining. Cells transduced with LVXLA2 are within the blue box; VCNs are indicated. (C) Histogram of BTK expression for the indicated cells (by color). Figure 23 - Intracellular phospho-PLCγ2 levels increase in response to IgM stimulation and correlate with LVXLA2 copy number in BTK KO Ramos clone1 cells. The panels on the left show overlaid histograms of GFP or phospho-PLCg2 expression of the resting (pre- stimulation) and anti-IgM stimulated flow cytometry data presented in the columns to the right. Each row corresponds to the cells indicated on the right. Figure 24 - IVIM Assay workflow. Figure 25 - Proliferation rate of samples at different days in the IVIM assay. Dotted lines mark 5% and 95% percentile values of meta-analysis data from 85 MOCK controls. Bars indicate means. Figure 26 - Replating Frequency (RF) of the control samples MOCK or RSF91 and the test vectors compared to data of a meta-analysis for control samples (Mock-MA, RSF91- MA, lv-SF-MA). The data points below the limit of detection (LOD; plates with no wells above the MTT-threshold) were manually inserted into the graph (due to the logarithmic scale of the y-axis). Above the graph, the incidence of positive (RF ≥ 3.17 x10-4) and negative plates (RF < 3.17 x10-4) according to the MTT-assay are shown. Differences in the incidence of positive and negative assays relative to Mock-MA or RSF91-MA were analyzed by Fisher's exact test with Benjamini-Hochberg correction (***P < 0.001; **P < 0.01; *P < 0.05; NS = not significant). Bars indicate mean RF. Figure 27 – Effect of HSCs transduced with LVXLA2 on B cell hematopoiesis in mXLA recipients. (A) Input VCN of LVXLA2-transduced input mHSCs were analyzed from bulk CFU colonies from two replicates per experiment; replicates were averaged. (B, C). Plots show VCN from genomic DNA (gDNA) isolated from (B) BM and (C) spleen at endpoint analysis (18- 22 weeks post-transplant of primary recipient mice). (D-G) Immunophenotyping of hematopoietic BM and splenic lineages of primary recipient mice at the scheduled endpoint. Leukocyte lineages in BM (D, E) and Spleen (F,G) graphed as a percent of all cells within the gated lymphocyte singlets. Major leukocyte markers were: monocytes (CD11b+Gr-1lo), neutrophils (CD11b+Gr-1+), B cells (B220hi), and T cells (CD4+or CD8+). Developmental bone marrow CD19+B cell markers were: mature (IgM+IgD+), immature (IgM+IgD-), and Pre+Pro (IgM- CD43+ / -). B cell developmental populations within splenic B220+B cells were defined as: marginal zone (MZ; CD21hiCD23-) and follicular mature (FM; CD21+ / -CD23+). Data represent Mean ± SD from n = 10-13 mXLA Mock, 21-23 LVXLA2, and 11-13 WT Mock primary recipients in three independent experiments. P values were determined using one-way ANOVA and Tukey’s correction for multiple comparisons; statistical significance in C and D shown in right panels; **** P ≤ 0.0001; *** P ≤ 0.001; ** P ≤ 0.01; * P ≤ 0.05. If not shown, statistical comparisons were not significant (p >0.05). For (A), (B), (C), (D), and (F), columns from left to right for each cell type / source are: mXLA Mock, LVXLA2, WT Mock. Figure 28 - Functional BTK expression in hematopoietic cells derived from LVXLA2 gene therapy treated mHSCs. All data is from cells collected at primary recipient animal sacrifice and analyzed by flow cytometry. (A) BM mHSC (Linneg) intracellular BTK expression graphed as percent of total mHSCs. (B) Cells of indicated lineages expressing intracellular BTK in bone marrow (BM) and spleen, graphed as percent of total cells in the subset. (C, D) MFI of BTK in all cells of the indicated subset, including those in the BTK+gate and those excluded from the BTK+gate; MFI for each sample normalized to average of WT Mock animals in the same cohort. (E, F) In vitro responses of splenic B cells (B220-selected) after 48- to 72-hour culture with the indicated stimulants. (E) Representative proliferation data graphed as histograms of CellTrace cell membrane dye dilution from mXLA Mock (left), LVXLA2 (center), and WT Mock (right) from live B cells in LPS, IgM, and unstimulated conditions, as indicated. (F) B cell proliferation assay showing the percent of live B cells that proliferated (CellTracelo) in response to no stimulation, IgM, or LPS stimulation, at time of collection. Panels (A-D) show the Mean ± SD from n = 13 (mXLA Mock), 21-23 (LVXLA2) and 12 (WT Mock) primary recipients in three independent experiments. In (F), bars represent mean from n= 3 (mXLA Mock No Stim), 3 (mXLA Mock IgM), 6 (mXLA Mock LPS), 17 (LVXLA2, all conditions), and 9 (WT Mock, all conditions). P values were determined using one-way ANOVA and Tukey’s correction for multiple comparisons; **** P ≤ 0.0001; ** P ≤ 0.01. If not shown, statistical comparisons were not significant (p >0.05). For (A), (B), (C), (D), and (F), columns / data points from left to right for each cell type / source are: mXLA Mock, LVXLA2, WT Mock. Figure 29 – Engraftment of human HSPCs transduced by LVXLA2 in humanized mice. (A) VCN calculated based upon droplet digital (dd)PCR of gDNA extracted from (left) single CFU colonies of input LVXLA2 transduced HSPCs, (middle) bone marrow, or (right) spleen cells collected from each mouse at the experimental endpoint (16- to 18-weeks post- transplant). Plots display combined VCN analyses of two independent experiments. For the input cells, data represents all valid colony reads from two independent DP manufacturing processes with an applied threshold of VCN ≥0.5. For the bone marrow and spleen, each dot represents data from a single animal and combined data the two experimental cohorts are displayed (Mock = columns / data on the left, and LVXA2 = columns / data on the right of each graph), with the data rep mice engrafted with Mock transduced cells on the left and . Bars represent mean ± SEM. VCN=viral copy number per cell. (B) Kaplan-Meier survival curve of recipient mice post-transplant. The % survival between the Mock (100% survival – top line) and LVXLA2 groups was not statistically significant (ns). Figure 30 - Assessment of hematopoiesis and BTK expression in bone marrow and spleen. Plots summarize immunophenotyping data of bone marrow (BM) and spleen cells at the experimental endpoint (16- to 18-weeks). (A) Absolute cell counts of human CD34+cells and CD34+CD38low(LT-HSC) cells within the BM harvested from the hind limbs of each mouse. (B) Absolute cell counts of human hematopoietic cell lineages in the bone marrow including all human leukocytes (left), or leukocyte subsets (myeloid, B cells, and T cells; right). (C) Intracellular BTK expression in splenic human B cells identified in the spleen. The mean fluorescence intensity (MFI) of BTK staining within the human B cell compartment for each mouse was normalized to the mean value for recipients of Mock transduced HSPCs within each experimental cohort. Each dot represents data from a single mouse. Bars represent mean + / - SEM for the combined two independent experimental cohorts. Mock = columns / data on the left, and LVXA2 = columns / data on the right for each cell type. P values were determined using one-way ANOVA and Tukey’s correction for multiple comparisons. ** P ≤ 0.01; ns= not significant (p >0.05). Figure 31 - VISA of LVXLA2-transduced PB-HSPCs and bone marrow of transplanted recipients. Viral integration site analysis (VISA) was determined for four experiments: (A)2203A, (B)2203B, (C)2204 and (D)2204B. VISA was performed by ProtaGene using a shearing extension primer tag selection and ligation-mediated polymerase chain reaction (S- EPTS / LM-PCR) method developed for amplification and sequencing of unknown genomic sequences flanking the integrated vector DNA. Left column shows VISA of the LVXLA2 DPs used to transplant the busulfan conditioned NBSGW-recipients (2203A_Input, 2203B_Input (A and B)) and VISA of LVXLA XLA DPs (2204_Input and 2204B_Input (C and D)). Note that the input cells were expanded in culture 14-days to dilute out non-integrated LV particles that would interfere with the assay. Right column, at the end of the in-life study, genomic DNA (gDNA) was purified from BM cells from each LVXLA2 DP recipient mouse (A and B) and LVXLA2 XLA DP recipient mouse (C and D) and used for IS analysis. Because each mouse in the experiment is engrafted by a limited pool of long-term (LT)-HSCs, each mouse is expected to present a limited, oligoclonal phenotype by VISA. To increase the breadth of LV-HSCs being sampled by the assay, gDNA from the LVXLA2 DP treated and LVXLA2 XLA DP treated mice in each cohort were pooled, labeled “BM ALL” (representing bone marrow cells harvested and pooled from all the mice used for the respective study). Sequence count of the ten most prominent IS (“Seq. count top 10”), sequence count of all remaining IS (“Seq. count other mapp. IS”) and total IS-specific sequence count from all replicates (“Total seq. count used”) are shown at the bottom for each sample. RefSeq names of genes located closest to the respective IS are given in the table (“Gene”). Relative sequence count contributions of the ten most prominent IS and all remaining mappable IS are shown (“Freq.”, Frequency [%]). The location column is composed of chromosome number, sequence orientation (plus or minus) and IS locus (based on human reference genome hg38). Figure 32 - VCN of and survival of recipients of LVXLA2 or Mock XLA or healthy human donor (HD) drug products (DPs). (A) VCN calculated based by digital droplet (dd)PCR of gDNA extracted from (left) single CFU colonies of input LVXLA2 transduced HSPCs, (middle) bone marrow, or (right) spleen cells collected from each mouse at the experimental endpoint (17- to 18-weeks post-transplant). For the input cells, data represents all valid colony reads from two independent DP manufacturing processes with an applied threshold of VCN ≥0.5. For the bone marrow and spleen, each dot represents data from a single animal. Bars represent mean ± SEM. VCN=viral copy number per cell. Columns / data points from left to right for bone marrow and spleen graphs are: HD Mock, XLA Mock, LVXLA2. (B) Kaplan-Meier survival curve of recipient mice post-transplant. The % survival was not statistically significant between any of the groups. The combined data from the two experimental cohorts (2204A and 2204B) are displayed. Figure 33 - Human cell engraftment and hematopoiesis in murine recipients of LVXLA2 or Mock transduced HSPCs from XLA patients, or Mock transduced healthy donors. Plots summarize immunophenotyping data of bone marrow (A) and spleen cells (B) at the experimental endpoint 17- to 18-weeks post-transplant. Two independent studies were performed using HSPCs from 2 independent healthy donors and 2 XLS subjects (Study 2204A, and 2204B, respectively). (A) Absolute cell counts of human hematopoietic cells within the BM harvested from the hind limbs of each mouse. Enumerated human cells were: hematopoietic cells (CD45+), HSCs (CD34+), LT-HSCs (CD34+CD38low), B cells (CD19+), myeloid cells (CD33+), and T cells (CD4 / CD8+). (B) Absolute cell counts of human hematopoietic cell lineages in the spleen including all human leukocytes (hCD45+) or the following leukocyte subsets: B cells (CD19+), myeloid (CD33+) and T cells (CD4 / CD8+). Each dot represents data from a single mouse. Bars represent mean ± SEM for the combined two independent experimental cohorts (2204A and 2204B). P values were determined using one-way ANOVA and Tukey’s correction for multiple comparisons. **= P ≤ 0.01; *=P ≤ 0.05; ns= not significant (p >0.05). Columns / data points from left to right for each graph are: HD Mock, XLA Mock, LVXLA2. Figure 34 - Human B cell developmental subsets in the bone marrow and spleen of murine recipients of LVXLA2. Plots summarizing developmental B cell immunophenotypes in the bone marrow (A) and spleen (B) at the experimental endpoint 17- to 18-weeks post- transplant. (A) Percentages of the indicated B cell developmental subset (top row) and their absolute cell counts (bottom row) within the BM harvested from the hind limbs of each mouse. Linneg=CD34+ CD19-; Pro-B cells = CD34+ CD19+; Pre-B cells = CD34- CD19+; Immature B cells = CD19+ IgM+. (B) Percentage of mature human B cells (CD19+ CD24- CD38-) in the spleen (relative to all human CD19+ B cells; left) and their absolute cell count in the spleen (right). Each dot represents data from a single mouse. Bars represent mean ± SEM for the combined two independent experimental cohorts (2204A and 2204B). P values were determined using one-way ANOVA and Tukey’s correction for multiple comparisons. ****=P≤ 0.0001; ***=P≤ 0.001; **= P ≤ 0.01; *=P ≤ 0.05; ns= not significant (p >0.05). Columns / data points from left to right for each graph are: HD Mock, XLA Mock, LVXLA2. Figure 35 - Antibody responses and reconstitution of peripheral BTK+ B cells at low chimerism of BTK-sufficient donor BM. (A) Antibody responses to T-dependent immunization (NP-CGG) in chimeric transplanted mice. Graph shows the concentration of serum antigen- specific IgG determined by ELISA for the indicated groups (from left to right for each time point: 100% KO, 1-5% WT, 10-20% WT, 50-100% WT). Serum samples were collected at three different timepoints: immediately prior to immunization (pre-immune), and ~one week following the primary and secondary immunizations. (B) Immunophenotyping data showing the percentage of BTK+ B cells at the 20-week experimental endpoint for each mouse is plotted vs. its bone marrow chimerism of WT neutrophils (a surrogate for hematopoietic stem cell chimerism). The curve was calculated using a non-linear regression asymmetric 5 parameter - curve fit. The dotted line demarcates the flattening of the curve at ~20% WT chimerism. Some figures and text contain color representations or entities. Color illustrations are available from the Applicant upon request or from an appropriate Patent Office. A fee may be imposed if obtained from a Patent Office. DETAILED DESCRIPTION OF THE INVENTION Definitions Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, preferred methods and materials are described. For the purposes of the present invention, the following terms are defined below. The articles “a” and “an” are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, “an element” means one element or more than one element. As used herein, “and / or” refers to and encompasses any and all possible combinations of one or more of the associated listed items, as well as the lack of combinations when interpreted in the alternative (or). Throughout this specification, unless the context requires otherwise, the words “comprise”, “comprises” and “comprising” will be understood to imply the inclusion of a stated step or element or group of steps or elements but not the exclusion of any other step or element or group of steps or elements. Thus, use of the term “comprising” and the like indicates that the listed elements are required or mandatory, but that other elements are optional and may or may not be present. By “consisting of” is meant including, and limited to, whatever follows the phrase “consisting of”. Thus, the phrase “consisting of” indicates that the listed elements are required or mandatory, and that no other elements may be present. By “consisting essentially of” is meant including any elements listed after the phrase, and limited to other elements that do not interfere with or contribute to the activity or action specified in the disclosure for the listed elements. Thus, the phrase “consisting essentially of” indicates that the listed elements are required or mandatory, but that other elements are optional and may or may not be present depending upon whether or not they affect the activity or action of the listed elements. The term “construct” in the context of a vector or nucleic acid construct refers to a genetic (nucleic acid) molecule including one or more polynucleotide sequences from one or more sources. Thus, constructs include recombinant or chimeric molecules in which two or more polynucleotide sequences of different origin are assembled into a single nucleic acid molecule. Vector and nucleic acid constructs may include a construct that contains polynucleotide sequences, including regulatory and coding sequences that are not found together in nature (i.e., at least one of the nucleotide sequences is heterologous with respect to at least one of its other nucleotide sequences). Representative constructs include plasmid, cosmid, virus, autonomously replicating polynucleotide molecule, phage, or linear or circular single stranded or double stranded DNA or RNA nucleic acid molecules, derived from any source, capable of genomic integration or autonomous replication, comprising a nucleic acid molecule where one or more nucleic acid molecules have been operably linked. For the practice of the methods of the present disclosure, conventional compositions and methods for preparing and using constructs and host cells are well known to one skilled in the art, see for example, Molecular Cloning: A Laboratory Manual, 3rdedition Volumes 1, 2, and 3. J. F. Sambrook, D. W. Russell, and N. Irwin, Cold Spring Harbor Laboratory Press, 2000. The term "corresponding" when used herein in the context of nucleotide positions "corresponding" to given positions in a specific sequence, refers to nucleotides or positions that occur at aligned loci. The sequences of related or variant polynucleotides are aligned by any method known to those of skill in the art. Such methods typically maximize matches (e.g. identical nucleotides or amino acids at positions), and include methods such as using manual alignments and by using the numerous alignment programs available (for example, BLASTN, BLASTP, ClustlW, ClustlW2, EMBOSS, LALIGN, KALIGN, etc) and others known to those of skill in the art. By aligning the sequences of polynucleotides, one skilled in the art can identify corresponding nucleotides. For example, by aligning the nucleotide sequence of the modified UCOE described herein and set forth in SEQ ID NO:2, with the UCOE sequence described in WO 2018 / 195297 and set forth in SEQ ID NO:1 (e.g. see Figure 1), one of skill in the art can identify nucleotides and nucleotide positions within the modified UCOE of SEQ ID NO:2 that correspond to specific nucleotides and nucleotide positions in the UCOE sequence set forth in SEQ ID NO:1. Thus, when nucleotides or positions are referred to herein with respect to the UCOE sequence of SEQ ID NO:2 it is understood that, where appropriate, the reference is also to the corresponding nucleotide or position in the UCOE sequence of SEQ ID NO:1, and vice versa. A "transgene" as used herein refers to an exogenous DNA or cDNA sequence sequence present in a polynucleotide, vector or host cell that encodes a functional Bruton's tyrosine kinase (BTK) polypeptide or protein. The transgene may be foreign to the host cell into which it is introduced, or may represent a gene whose expression is otherwise absent or reduced in the host cell in the absence of the introduction and expression of the transgene. As used herein, the term "expression cassette" refers to one or more genetic sequences within a vector which can express BTK RNA subsequently a functional BTK polypeptide or protein. The expression cassette comprises at least one promoter and at least a gene encoding BTK. In some embodiments, the expression cassette further includes at least one additional nucleic acid sequence encoding a molecule for expression. In particular embodiments, the expression cassette is positionally and sequentially oriented within the vector such that the nucleic acid in the cassette can be transcribed into RNA, and translated into a protein or a polypeptide, undergo appropriate post-translational modifications required for activity in the host cell (e.g. a transduced stem cell), and be translocated to the appropriate compartment for biological activity In some embodiments, the expression cassette has 3' and 5' ends adapted for ready insertion into a vector, e.g., it has restriction endonuclease sites at each end. As used herein, the term "operably linked" refers to functional linkage between a nucleic acid expression control sequence (such as a promoter, signal sequence, enhancer or array of transcription factor binding sites) and a second nucleic acid sequence, wherein the expression control sequence affects transcription and / or translation of the nucleic acid corresponding to the second sequence when the appropriate molecules (e.g., transcriptional activator proteins) are bound to the expression control sequence. As used herein, the term "promoter" refers to a recognition site of a polynucleotide (DNA or RNA) to which an RNA polymerase binds. An RNA polymerase initiates and transcribes polynucleotides operably linked to the promoter. In some embodiments, promoters operative in mammalian cells comprise an AT-rich region located approximately 25 to 30 bases upstream from the site where transcription is initiated and / or another sequence found about 70 to about 80 bases upstream from the start of transcription, e.g. a CNCAAT region where N may be any nucleotide. The term "enhancer" is used herein in its ordinary sense to refer to a nucleotide region comprising a sequence capable of increasing the level of transcription of a transgene from a promoter as compared to the level of transcription of the transgene from the promoter in the absence of the enhancer. Enhancers may be cis-acting or trans-acting, and may be located upstream or downstream of the transgene sequence, in either forward or reverse orientation with respect to the transgene sequence. An enhancer may be, for example, about 50, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1100, 1200, 1300, 1400, or 1500 base pairs, or any number of base pairs in a range defined by any two of the aforementioned values. As used herein, the term "host cell" refers to cells that is to be modified using the methods of the present disclosure. In some embodiments, the host cells are mammalian cells in which the lentiviral vector can be introduced. Suitable mammalian host cells include, but are not limited to, human cells, murine cells, non-human primate cells (e.g. rhesus monkey cells), human progenitor cells or stem cells, 293 cells, HeLa cells, D17 cells, MDCK cells, BHK cells, and Cf2Th cells. In certain embodiments, the host cell comprising an expression vector of the disclosure is a hematopoietic cell, such as hematopoietic progenitor / stem cell (e.g. CD34-positive hematopoietic progenitor / stem cell), a monocyte, a macrophage, a peripheral blood mononuclear cell, a CD4+ T lymphocyte, a CD8+ T lymphocyte, or a dendritic cell. The hematopoietic cells (e.g. CD4+ T lymphocytes, CD8+ T lymphocytes, and / or monocyte / macrophages) to be transduced with an expression vector of the disclosure can be allogeneic, autologous, or from a matched sibling. The hematopoietic cells are, in some embodiments, CD34-positive and can be isolated from the patient's bone marrow or peripheral blood. The isolated CD34-positive hematopoietic cells (and / or other hematopoietic cell described herein) is, in some embodiments, transduced with an expression vector as described herein. As used herein, the term "hematopoietic stem cells" or "HSCs" refer to multipotent precursor cells capable of differentiating into all the cell types of the hematopoietic system, including, but not limited to, granulocytes, monocytes, erythrocytes, megakaryocytes, lymphocytes, dendritic cells; and self-renewal activity, i.e. the ability to divide and generate at least one daughter cell with the identical (e.g., self-renewing) characteristics of the parent cell. The term "codon optimized" as used herein has its plain meaning understood by those skilled in the art, referring to a polynucleotide encoding a polypeptide in which codon usage in the polynucleotide is altered to accommodate codon bias in a host cell and thereby optimize translational efficiency in the host cell. Codon optimization can be performed using algorithms known to those skilled in the art so as to create synthetic genetic transcripts optimized for high mRNA and protein yield in humans. Programs containing algorithms for codon optimization in humans are readily available. Such programs can include, for example, OptimumGene™ or GeneGPS® algorithms. Additionally human codon optimized sequences can be obtained commercially. As used herein, the term "vector" includes reference to both polynucleotide vectors and viral vectors, each of which are capable of delivering a transgene contained within the vector into a host cell. Vectors can be episomal, i.e., do not integrate into the genome of a host cell, or can integrate into the host cell genome. The vectors may also be replication competent or replication-deficient. Exemplary polynucleotide vectors include, but are not limited to, plasmids, cosmids and transposons. Exemplary viral vectors include, for example, AAV, lentiviral, retroviral, adenoviral, herpesviral and hepatitis viral vectors. As used herein, the term "lentivirus" refers to a genus of retroviruses that are capable of infecting dividing and non-dividing cells. Several examples of lentiviruses include HIV (human immunodeficiency virus: including HIV type 1, and HIV type 2), the etiologic agent of the human acquired immunodeficiency syndrome (AIDS); visna-maedi, which causes encephalitis (visna) or pneumonia (maedi) in sheep, the caprine arthritis-encephalitis virus, which causes immune deficiency, arthritis, and encephalopathy in goats; equine infectious anemia virus, which causes autoimmune hemolytic anemia, and encephalopathy in horses; feline immunodeficiency virus (FIV), which causes immune deficiency in cats; bovine immune deficiency virus (BIV), which causes lymphadenopathy, lymphocytosis, and possibly central nervous system infection in cattle; and simian immunodeficiency virus (SIV), which causes immune deficiency and encephalopathy in sub-human primates. As used herein, "adeno-associated viral vector" or AAV vector refers to a vector derived from an adeno-associated virus, including without limitation, AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12 or AAV13, or using synthetic or modified AAV capsid proteins. An AAV vector may also be referred to herein as "recombinant AAV", "rAAV", "recombinant AAV virion", and "rAAV virion," terms which are used interchangeably and refer to a replication-defective virus that includes an AAV capsid shell encapsidating an AAV genome. The AAV genome (also referred to as vector genome, recombinant AAV genome or rAAV genome) comprises a transgene flanked on both sides by functional AAV inverted terminal repeats (ITRs). Typically, one or more of the wild-type AAV genes have been deleted from the genome in whole or part, preferably the rep and / or cap genes. Functional ITR sequences are necessary for the rescue, replication and packaging of the vector genome into the rAAV virion. As used herein, the term "lentiviral vector" is used to denote any form of a nucleic acid derived from a lentivirus and used to transfer genetic material into a cell via transduction. The term encompasses lentiviral vector nucleic acids, such as DNA and RNA, encapsulated forms of these nucleic acids, and viral particles in which the viral vector nucleic acids have been packaged. As used herein, the term "mutated" refers to a change in a sequence, such as a nucleotide or amino acid sequence, from a native, standard, or reference version of the respective sequence, i.e. the non-mutated sequence. As used herein, the terms "transduce" or "transduction" refer to the delivery of a gene(s) using a viral or retroviral vector by means of infection rather than by transfection. For example, an anti-HPRT gene carried by a retroviral vector (a modified retrovirus used as a vector for introduction of nucleic acid into cells) can be transduced into a cell through infection and provirus integration. Thus, a "transduced gene" is a gene that has been introduced into the cell via lentiviral or vector infection and provirus integration. Viral vectors (e.g., "transducing vectors") transduce genes into "target cells" or host cells. As used herein, the terms “treatment”, “treating”, and the like, refer to obtaining a desired pharmacologic and / or physiologic effect in a subject in need of treatment, that is, a subject who has a disease or disorder. By "treatment" is meant ameliorating or preventing one or more symptoms or effects (e.g. consequences) of a disease or disorder. Reference to “treatment”, “treat” or “treating” does not necessarily mean to reverse or prevent any or all symptoms or effects of a disease or disorder. For example, the subject may ultimately suffer one or more symptoms or effects, but the number and / or severity of the symptoms or effects is reduced and / or the quality of life is improved compared to prior to treatment. By “effective amount”, in the context of treating a disease or condition is meant the administration of an amount of an agent or composition to an individual in need of such treatment or prophylaxis, either in a single dose or as part of a series, that is effective for the prevention of incurring a symptom, holding in check such symptoms, and / or treating existing symptoms, of that condition. The effective amount will vary depending upon the age, health and physical condition of the individual to be treated and whether symptoms of disease are apparent, the taxonomic group of individual to be treated, the formulation of the composition, the assessment of the medical situation, and other relevant factors. Optimal dosing schedules can be calculated from measurements of drug accumulation in the body of the subject. Optimum dosages may vary depending on the relative potency in an individual subject, and can generally be estimated based on EC50 values found to be effective in in vitro and in vivo animal models. Persons of ordinary skill can easily determine optimum dosages, dosing methodologies and repetition rates. It is expected that the amount will fall in a relatively broad range that can be determined through routine trials. The terms "subject", “patient” and “individual” used interchangeably herein, refer to any subject, particularly a vertebrate subject, and even more particularly a mammalian subject, (e.g. human). Each embodiment described herein is to be applied mutatis mutandis to each and every embodiment unless specifically stated otherwise. Table 1. Brief Description of the Sequences Vectors Sequences The present disclosure provides nucleic acid constructs and vectors comprising said nucleic acid constructs facilitating the expression of Bruton's tyrosine kinase (BTK) in host cells, and useful for gene therapy applications in the treatment of X-linked agammaglobulinemia (XLA) and for inhibiting or ameliorating at least one symptom of XLA. BTK is expressed in both B cells and myeloid cells where it also contributes to normal functional responses in both lineages. Failure to express BTK leads to XLA. Conversely, overexpression of activated or wild type BTK can lead to cell transformation and / or developmental blockade. Thus, safe and successful clinical gene therapy in XLA requires restoration of BTK expression in each cell lineage that normally expresses the protein as well as tightly regulated expression that does not lead to overexpression in cells that do not normally express the protein, and where expression might promote altered cell function. Present inventors have previously developed candidate lentiviral vectors for the expression of BTK, and evaluated these vectors in in murine XLA animal models and human cell lines derived from human subjects with XLA (as described in WO 2018 / 195297, the disclosure of which is incorporated herein in its entirety). The vectors incorporated a truncated ubiquitous chromatin opening element (UCOE). UCOEs are regulatory elements derived from promoter- containing regions of ubiquitously expressed housekeeping genes, facilitating transcription through chromatin remodeling activity. Various UCOEs have been shown to confer high levels of stable expression of proximal transgenes. The vectors described in WO 2018 / 195297 include a UCOE spanning a CpG-rich region across the divergently transcribed promoter regions for the housekeeping genes CBX3 and HNRPA2B1, truncated either to a 1.5kb construct starting at exon 1 of CBX3 and extending beyond CBX3 alternate exon 1, or a 0.7 kb construct which eliminates the region downstream of CBX3 alternative exon 1. The UCOEs utilized in the vectors of WO 2018 / 195297 include cryptic splice sites which can induce alternative splicing of transgene RNA, lentiviral genomic RNA, or transcripts from a gene into which the vector genome integrates, resulting in truncated, non-functional transcripts. Vectors of the present disclosure comprise a UCOE comprising a nucleotide sequence at least about 90% identical to the sequence set forth in SEQ ID NO:1, in which cryptic splice acceptor sites have been inactivated by mutation, such that: (i) the sequence GTAAG at positions 347-351 of SEQ ID NO:1 is replaced by the sequence CGAAC; (ii) the sequence GTCGG at positions 362-366 of SEQ ID NO:1 is replaced by the sequence CGCGC; (iii) the A residue at position 385 of SEQ ID NO:1 is replaced by a T residue; and (iv) the sequence GTCGG at positions 612-616 of SEQ ID NO:1 is replaced by the sequence CGCGC. Accordingly, provided herein is a vector for the treatment of XLA, or for inhibiting or ameliorating at least one symptom of XLA, the vector comprising: (a) a transgene expression cassette comprising a nucleotide sequence encoding a functional BTK polypeptide operably linked to a promoter; (b) at least one regulatory or enhancer element; (c) a UCOE, wherein the UCOE comprises a nucleotide sequence at least about 90% identical to the sequence set forth in SEQ ID NO:1, and wherein the sequence of the UCOE comprises the following nucleotide substitutions relative to SEQ ID NO:1, at positions corresponding to those of SEQ ID NO:1: (i) the sequence CGAAC at positions 347-351; (ii) the sequence CGCGC at positions 362-366; (iii) a T residue at position 385; and (iv) the sequence CGCGC at positions 612-616. The UCOE present in vectors of the present disclosure further comprise single nucleotide mutations at positions corresponding to positions 4, 496 and 673 of SEQ ID NO:1, such that: the C residue at position 4 of SEQ ID NO:1 is replaced with a G residue; the A residue at position 496 of SEQ ID NO:1 is replaced with a G residue; and the G residue at position 673 of SEQ ID NO:1 is replaced with a T residue. Accordingly, provided herein is a vector for the treatment of XLA, or for inhibiting or ameliorating at least one symptom of XLA, the vector comprising: (a) a transgene expression cassette comprising a nucleotide sequence encoding a functional BTK polypeptide operably linked to a promoter; (b) at least one regulatory or enhancer element; (c) a ubiquitous chromatin opening element (UCOE), wherein the UCOE comprises a nucleotide sequence at least about 90% identical to the sequence set forth in SEQ ID NO:1, and wherein the sequence of the UCOE comprises the following nucleotide substitutions relative to SEQ ID NO:1, at positions corresponding to those of SEQ ID NO:1: (i) a G residue at position 4; (ii) the sequence CGAAC at positions 347-351; (iii) the sequence CGCGC at positions 362-366; (iv) a T residue at position 385; (v) a G residue at position 496; (vi) the sequence CGCGC at positions 612-616; and (vii) a T residue at position 673. The UCOE may comprise a sequence at least or about 90%, 91%, 92%, 93%, 94%, 95%, 95.5%, 96%, 96.5%, 97%97.5%, 98%, 98.5%, 99%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9% identical to the sequence of SEQ ID NO:1. In particular embodiments of the present disclosure the UCOE comprises the sequence set forth in SEQ ID NO:2. The UCOE may be at least or about 0.3kb, 0.4kb, 0.5kb, 0.6k, 0.7kb, 0.8kb, 0.9kb, 1.0kb, 1.1kb, 1.2kb, 1.3kb, 1.4kb, 1.5kb, 1.6kb, 1.7kb, 1.8kb, 1.9kb or 2kb in length. In exemplary embodiments, the UCOE may be about 0.7kb or about 1.5kb in length, optionally about 0.7kb in length. In exemplary embodiments, the UCOE consists of the sequence set forth in SEQ ID NO:2. Vectors of the present disclosure comprise a transgene expression cassette comprising a polynucleotide encoding a functional BTK polypeptide, operably linked to a promoter. Typically the BTK polypeptide is a human BTK polypeptide and the BTK polypeptide encoded by the polynucleotide is functional in that it produces a BTK enzyme that shares qualitative and / or quantitative activity in common with the wild-type BTK protein, suitable to restore or provide cellular BTK levels and activity suitable to overcome or alleviate at least one symptom characteristic of XLA. In exemplary embodiments, the BTK polypeptide encoded by the transgene expression cassette is the wild-type human BTK polypeptide comprising the amino acid sequence set forth in SEQ ID NO:3, or a variant polypeptide comprising at least or about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO:3, wherein the variant polypeptide retains activity of the wild-type BTK polypeptide. Typically, the variant polypeptide will retain at least or about 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90% or 95% of the activity of the wild-type BTK. BTK activity can be assessed by any method known in the art (e.g. as described in van Borstel et al., 2019, Rheumatology 58:2230-2239). In specific embodiments of the present disclosure, the BTK transgene is codon- optimized for expression in a suitable host cell, particularly a human codon-optimized transgene. In a particular embodiment the BTK transgene is human codon-optimized and comprises the nucleotide sequence set forth in SEQ ID NO:4 or SEQ ID NO:5, or a sequence having at least or about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto, wherein the encoded BTK polypeptide retains at least or about 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90% or 95% of the activity of the wild-type BTK enzyme. In particular exemplary embodiments of the present disclosure, the codon- optimized BTK transgene comprises the nucleotide sequence of SEQ ID NO:4. The BTK transgene is operably linked to a promoter, optionally a B cell lineage- specific promoter. Typically the promoter facilitates the expression of the BTK transgene in B cells, myeloid cells or hematopoietic stem cells. The promoter may drive constitutive, conditional or inducible expression of the transgene to which it is operably linked. In an embodiment, the promoter is a BTK promoter comprising the sequence set forth in SEQ ID NO:6, or a sequence having at least or about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto. The BTK promoter may be functional fragment of the promoter sequence set forth in SEQ ID NO:6 or sequence at least or about 80% identical thereto, wherein the functional fragment retains the ability to drive transcription of an operably linked BTK transgene. An exemplary functional fragment may comprise at least or about 300, 350, 400, 450. 500, 550, 600, 650, 700, 750 or 800 consecutive nucleotides of the sequence set forth in SEQ ID NO:6, or sequence at least or about 80% identical thereto. In exemplary embodiments, the transgene expression cassette present in vectors of the present disclosure, comprising the BTK transgene operably linked to a BTK promoter, comprise the sequence set forth in SEQ ID NO:7, or a sequence having at least or about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto. In exemplary embodiments, the UCOE is present in the vector upstream (5' to) the promoter of the transgene expression cassette. The UCOE may be located in the vector in the forward orientation with respect to the transgene expression cassette or in the reverse orientation with respect to the transgene expression cassette. In particular embodiments, the UCOE is located in the forward orientation with respect to the transgene expression cassette. Accordingly, in specific embodiments, vectors of the present disclosure comprising a UCOE comprising the sequence of SEQ ID NO:2, the BTK promoter of SEQ ID NO:6 and the codon- optimized BTK polynucleotide of SEQ ID NO:4, may comprise the sequence set forth in SEQ ID NO:8, or a sequence having at least or about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto. Vectors of the present disclosure typically comprise one more regulatory elements, including, for example, promoters, operators, enhancers, termination signals, polyadenylation signals, etc. Those skilled in the art can readily identify suitable elements for the correct processing, transcription and / or translation of nucleic acid present in and encoded by the vectors and for efficient transduction of the vectors. The vector typically comprises suitable transcription termination signals including but not limited to polyadenylation signals such as, for example, bovine growth hormone SV40 polyA, rabbit beta-globin (RBG) polyA, BGH polyA, thymidine kinase (TK) polyA sequences, and any variants thereof. The vectors typically comprise a polyadenylation (polyA) signal downstream of the transgene expression cassette. In exemplary embodiments, the polyadenylation signal is an SV40 polyA signal, for example comprising the sequence set forth in SEQ ID NO:18 or a sequence having at least or about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto. In other exemplary embodiments, the polyadenylation signal is rabbit beta-globin polyA signal, for example comprising the sequence set forth in SEQ ID NO:17 or a sequence having at least or about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto. The vector may comprise one or more post-transcriptional regulatory elements that can function to increase expression of the transgene. Such post-transcriptional regulatory elements include the woodchuck hepatitis virus post-transcriptional regulatory element (WPRE), an RNA transport element (RTE) and any variants thereof. In exemplary embodiments, the vector comprises a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE). In a particular embodiment, the WPRE is downstream of the transgene expression cassette. In some embodiments, the WPRE is a wild type MPRE sequence comprising a sequence set forth in SEQ ID NO:9 or a WPRE mut6 comprising a sequence set forth in SEQ ID NO:10, or comprises a sequence having at least or about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97% or 99% sequence identity to the sequence set forth in SEQ ID NO:9 or 10. The vector may also comprise a translational initiation enhancer, such as a Kozac sequence, optionally located between the promoter and transgene. The vector may also include one or more introns, which can serve to enhance mRNA processing and expression of the transgene, optionally located between the promoter and the transgene. The vector may comprise one or more regulatory sequences modulating transduction of the vector, such as one or more polypuyrine tracts (PPTs). For example, inclusion of a central polypurine tract (cPPT) is able to enhance transduction efficiency of lentiviral vectors. Accordingly, vectors of the present disclosure may comprise a cPPT sequence, optionally upstream of the UCOE of the transgene expression cassette, comprising the sequence of SEQ ID NO:12 or a sequence having at least or about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto. Types of vectors Vectors of the present disclosure may be viral or plasmid vectors, preferably viral vectors. The viral vector may be, for example, a retroviral vector such as a lentiviral vector, an adenovirus vector or an AAV vector. For the purposes of embodiments of the present disclosure, a viral vector is a vector which comprises nucleic acid that includes at least one component part derivable from a virus, such as, for example, a lentivirus, an adenovirus or an AAV. That component part may be involved in the biological mechanisms by which the vector infects cells, expresses genes or is replicated. Thus, viral vectors include nucleic acid molecules such as plasmids, and virus particles. In particular embodiments described herein, the vector is a lentiviral vector. Embodiments of the present disclosure are described below in the context of a lentiviral vector, however the person skilled in the art will recognise these as exemplary embodiments only and will appreciated that the scope of the present disclosure is not limited thereto. The basic structure of retrovirus and lentivirus genomes share many common features such as a 5' LTR and a 3' LTR, between or within which are located a packaging signal to enable the genome to be packaged, a primer binding site, integration sites to enable integration into a host cell genome and gag, pol and env genes encoding the packaging components, which are polypeptides required for the assembly of viral particles. Lentiviruses have additional features, such as the rev and rev response element (RRE) sequences, which enable the efficient export of RNA transcripts of the integrated provirus from the nucleus to the cytoplasm of an infected target cell. In the provirus, the viral genes are flanked at both ends by regions called long terminal repeats (LTRs). The LTRs are responsible for proviral integration, and transcription. LTRs also serve as enhancer-promoter sequences and can control the expression of the viral genes. The LTRs themselves are identical sequences that can be divided into three elements, which are called "U3," "R" and "U5." U3 is derived from the sequence unique to the 3' end of the RNA, R is derived from a sequence repeated at both ends of the RNA, and U5 is derived from the sequence unique to the 5' end of the RNA. The sizes of the three elements can vary considerably among different viruses. In one embodiment, at least part of one or more protein coding regions essential for replication may be removed from the vector, which makes the vector replication-defective. Portions of the viral genome may also be replaced by a nucleic acid in order to generate a vector comprising the nucleic acid which is capable of transducing a target non-dividing host cell and / or integrating its genome into a host genome. In one embodiment, the lentiviral vectors are non-integrating vectors as described in U.S. patent application Ser. No. 12 / 138,993 (herein incorporated by reference). The lentiviral vector may have a genome that has been manipulated to remove the non-essential elements and to retain the essential elements in order to provide the required functionality to infect, transduce and deliver a nucleotide sequence of interest to a target host cell (See, e.g., U.S. Pat. No. 6,669,936, incorporated by reference). In some embodiments, the genome is limited to sufficient lentiviral genetic information to allow packaging of an RNA genome, in the presence of packaging components, into a viral particle capable of infecting a target cell. Infection of the target cell may include reverse transcription and integration into the target cell genome. In some embodiments, the vector is incapable of independent replication to produce infectious lentiviral particles within the final target cell. In some embodiments, the lentiviral vector lacks a functional gag-pol and / or env gene and / or other genes essential for replication. In some examples, the lentiviral vector is a self-inactivating vector. Self- inactivating vectors may be constructed by deleting the transcriptional enhancers or the enhancers and promoter in the U3 region of the 3' LTR. After a round of vector reverse transcription and integration, these changes are copied into both the 5' and the 3' LTRs producing a transcriptionally inactive provirus (Yee et al., 1994, PNAS, 91:9564-68). However, any promoter(s) internal to the LTRs in such vectors will still be transcriptionally active. This strategy has been employed to eliminate effects of the enhancers and promoters in the viral LTRs on transcription from internally placed genes. Such effects include increased transcription (Jolly et al., 1983, Nucleic Acids Research, 11:1855-72) or suppression of transcription (Emerman & Temin, 1984, Cell, 39:449-67). This strategy can also be used to eliminate downstream transcription from the 3' LTR into genomic DNA (Herman & Coffin, 1987, Science, 236:845-48). A plasmid vector used to produce the viral genome within a host cell / packaging cell will also include transcriptional regulatory control sequences operably linked to the lentiviral genome to direct transcription of the genome in a host cell / packaging cell. These regulatory sequences may be the natural sequences associated with the transcribed lentiviral sequence, i.e. the 5' U3 region, or they may be a heterologous or modified promoter such as another viral promoter, for example the CMV promoter or the 7tetO promoter / operator. Some lentiviral genomes require additional sequences for efficient virus production. For example, in the case of HIV-based lentiviral vectors, the rev and RRE sequences are preferably included; however the requirement for rev and RRE may be reduced or eliminated by codon optimization (See U.S. patent application Ser. No. 12 / 587,236, incorporated by reference). Alternative sequences which perform the same function, as the rev / RRE system are also known. For example, a functional analogue of the revIRRE system is found in the Mason Pfizer monkey virus. This is known as the constitutive transport element (CTE) and comprises an RRE-type sequence in the genome which is believed to interact with a factor in the infected cell. The cellular factor can be thought of as a rev analogue. Thus, CTE may be used as an alternative to the reviRRE system. Any other functional equivalents which are known or become available may be relevant to the vectors of the present disclosure. For example, the Rex protein of HTLV-1 can functionally replace the Rev protein of HIV-1. It is also known that Rev and Rex have similar effects to IRE-BP. In some embodiments, the expression vector comprises sequences from the 5' and 3' long terminal repeats (LTRs) of a lentivirus. In some embodiments, the vector comprises the R and U5 sequences from the 5' LTR of a lentivirus and an inactivated or self- inactivating 3' LTR from a lentivirus. In some embodiments, the LTR sequences are HIV LTR sequences. In some embodiments, the lentiviral vectors contemplated herein may be integrative or non-integrating (also referred to as an integration defective lentivirus). As used herein, the term "integration defective lentivirus" or "IDLV" refers to a lentivirus having an integrase that lacks the capacity to integrate the viral genome into the genome of the host cells. In some applications, the use of by an integrating lentivirus vector may avoid potential insertional mutagenesis induced by an integrating lentivirus. Integration defective lentiviral vectors typically are generated by mutating the lentiviral integrase gene or by modifying the attachment sequences of the LTRs. Lentiviral integrase is coded for by the HIV-1 Pol region and the region cannot be deleted as it encodes other critical activities including reverse transcription, nuclear import, and viral particle assembly. Mutations in pol that alter the integrase protein fall into one of two classes: those which selectively affect only integrase activity (Class I); or those that have pleiotropic effects (Class II). Mutations throughout the N and C terminals and the catalytic core region of the integrase protein generate Class II mutations that affect multiple functions including particle formation and reverse transcription. Class I mutations limit their affect to the catalytic activities, DNA binding, linear episome processing and multimerization of integrase. The most common Class I mutation sites are a triad of residues at the catalytic core of integrase, including D64, D116, and E152. Each mutation has been shown to efficiently inhibit integration with a frequency of integration up to four logs below that of normal integrating vectors while maintaining transgene expression of the NILV. Another alternative method for inhibiting integration is to introduce mutations in the integrase DNA attachment site (LTR att sites) within a 12 base-pair region of the U3 region or within an 11 base-pair region of the U5 region at the terminal ends of the 5' and 3' LTRs, respectively. These sequences include the conserved terminal CA dinucleotide which is exposed following integrase-mediated end-processing. Single or double mutations at the conserved CA / TG dinucleotide result in up to a three to four log reduction in integration frequency; however, it retains all other necessary functions for efficient viral transduction. Production of vectors Vectors of the present disclosure can be produced using any method, and such methods are well known to those skilled in the art. In some embodiments, a modified UCOE as described herein and a transgene expression cassette are inserted into a vector, optionally a lentiviral vector, that is a plasmid, such as one selected from the group consisting of pRRL, pTL20c, pTL20d, FG, pCL20, pLKO.1 puro, pLKO.1, pLKO.3G, Tet-pLKO-puro, pSico, pLJM1- EGFP, FUGW, pLVTHM, pLVUT-tTR-KRAB, pLL3.7, pLB, pWPXL, pWPI, EF.CMV.RFP, pLenti CMV Puro DEST, pLenti-puro, pLOVE, pULTRA, pLJM1-EGFP, pLX301, pInducer20, pHIV-EGFP, Tet-pLKO-neo, pLV-mCherry, pCW57.1, pLionII, pSLIK-Hygro, and pInducer10-mir-RUP-PheS. In other embodiments, the vector, optionally the lentiviral vector, into which the modified UCOE and the transgenes expression cassette are inserted is selected from AnkT9W vector, a T9Ank2W vector, a TNS9 vector, a lentiglobin HPV569 vector, a lentiglobin BB305 vector, a BG-1 vector, a BGM-1 vector, a GLOBE vector, a G-GLOBE vector, a V5 vector, a V5m3 vector, a V5m3-400 vector, a G9 vector, and a BCL11A shmir vector. In a particular embodiment, the lentiviral expression vector is pRRL or pTL20c. For example, the UCOE and transgene expression cassette may be inserted into a pTL20c vector according to the methods described in United States Patent Publication No. 20180112233 and International Patent Publication No. WO2020 / 139796. Lentivirus particles or virions (or recombinant lentiviruses) can be produced using standard methods well known in the art. In one example, a stable producer cell line for generating virus is utilized, wherein the stable producer cell line is derived from one of a GPR, GPRG, GPRT, GPRGT, or GPRT-G packing cell line. In some embodiments, the stable producer cell line is derived from the GPRT-G cell line. In some embodiments, the stable producer cell line is generated by (a) synthesizing a vector by cloning desired nucleic acid sequences; (b) generating DNA fragments from the synthesized vector; (c) forming a concatemeric array from (i) the generated DNA fragments from the synthesized vector, and (ii) from DNA fragments derived from an antibiotic resistance cassette plasmid; (d) transfecting one of the packaging cell lines with the formed concatemeric array; and (e) isolating the stable producer cell line. Additional methods of forming a stable producer cell line are described in United States Patent Publication No. 20180112233. In some examples, lentivirus particles are produced using a stable producer cell line as described in Throm et al. 2009, Blood 113:5104–5110, International Patent Publication No. WO2016183260, and / or International Patent Publication No. WO2023187691. In other examples, transient transfection is utilized, such as described in Seymour et al. 2021, Molecular Therapy: Methods & Clinical Development 20: 635-651; Kerns et al. 2010, Blood 115(11):2146-2155; and International Patent Publication No. WO2018195297. Exemplary vectors Exemplary lentiviral vectors of the present disclosure include nucleic acid vectors (e.g. plasmids) and lentivirus virions (or virus particles) that comprise a 5’LTR (including R and U5 sequences, for example as set forth in SEQ ID NO:14) downstream of which are (in any order) a REV response element (RRE, for example as set forth in SEQ ID NO:11), a cPPT (for example as set forth in SEQ ID NO:12), a modified UCOE as described herein (for example as set forth in SEQ ID NO:2), a BTK transgene expression cassette (for example as set forth in SEQ ID NO:7), a WPRE (for example as set forth in SEQ ID NO:10), a 3' PPT, a polyA signal (for example as set forth in SEQ ID NO:17 or 18), ^U3 sequences (for example as set forth in SEQ ID NO:13) and ^R sequences. In these vectors, the modified UCOE is typically in the forward orientation with respect to the transgene expression cassette. Schematic representations of exemplary vectors of the present disclosure are shown in Figure 2C and Figure 2D. In an exemplary embodiment, a vector of the present disclosure is a lentiviral vector comprising the sequence of SEQ ID NO:20, or a sequence at least or about 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identical thereto. In another exemplary embodiment, a vector of the present disclosure is a lentiviral vector comprising the sequence of SEQ ID NO:21, or a sequence at least or about 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identical thereto. Host cells The present disclosure also provides a host cell comprising, transformed or transduced with a vector of the present disclosure. A "host cell" or "target cell" means a cell that is to be transformed or transduced using the methods and nucleic acid constructs and vectors of the present disclosure. In some embodiments, the host cells are mammalian cells in which the nucleic acid construct or vector can be expressed. Suitable mammalian host cells include, but are not limited to, human cells, murine cells, non-human primate cells (e.g. rhesus monkey cells), or human progenitor cells or stem cells. In certain embodiments, the host cell comprising a vector of the disclosure is a B cell, a myeloid cell or a hematopoietic progenitor / stem cell, such as a CD34+hematopoietic progenitor / stem cell. Hematopoietic progenitor / stem cells to be transduced with a vector of the disclosure can be allogeneic, autologous, or from a matched sibling. The cells are, in some embodiments, CD34+and can be isolated from a patient's bone marrow or peripheral blood. In some embodiments, the host cells or transduced host cells are combined with a pharmaceutically acceptable carrier. In some embodiments, the host cells or transduced host cells are formulated with PLASMA-LYTE A (e.g. a sterile, nonpyrogenic isotonic solution for intravenous administration; where one liter of PLASMA-LYTE A has an ionic concentration of 140 mEq sodium, 5 mEq potassium, 3 mEq magnesium, 98 mEq chloride, 27 mEq acetate, and 23 mEq gluconate). In other embodiments, the host cells or transduced host cells are formulated in a solution of PLASMA-LYTE A, the solution comprising between about 8% and about 10% dimethyl sulfoxide (DMSO). In some embodiments, the less than about 2x107host cells / transduced host cells are present per mL of a formulation including PLASMA-LYTE A and DMSO. In some embodiments, transduction of host cells may be increased by contacting the host cell, in vitro, ex vivo, or in vivo, with a vector of the present disclosure and one or more compounds that increase transduction efficiency. For example, in some embodiments, the one or more compounds that increase transduction efficiency are compounds that stimulate the prostaglandin EP receptor signaling pathway, i.e. one or more compounds that increase the cell signaling activity downstream of a prostaglandin EP receptor in the cell contacted with the one or more compounds compared to the cell signaling activity downstream of the prostaglandin EP receptor in the absence of the one or more compounds. In some embodiments, the one or more compounds that increase transduction efficiency are a prostaglandin EP receptor ligand including, but not limited to, prostaglandin E2 (PGE2), or an analog or derivative thereof. In other embodiments, the one or more compounds that increase transduction efficiency include but are not limited to, RetroNectin (a 63 kD fragment of recombinant human fibronectin fragment, available from Takara); Lentiboost (a membrane-sealing poloxamer), Poloxamer F127 (or Pluronic F127 or Poloxamer 407), protamine sulphate, cyclosporin H, and / or rapamycin. Pharmaceutical compositions The present disclosure also provides for compositions, including pharmaceutical compositions, comprising one or more vectors as disclosed herein. In some embodiments, pharmaceutical compositions comprise an effective amount of at least one of the vectors as described herein and a pharmaceutically acceptable carrier. For instance, in certain embodiments, the pharmaceutical composition comprises an effective amount of an vector and a pharmaceutically acceptable carrier. An effective amount can be readily determined by those skilled in the art based on factors such as body size, body weight, age, health, sex of the subject, ethnicity, and viral titers. The phrases "pharmaceutically acceptable" or "pharmacologically acceptable" refer to molecular entities and compositions that do not produce adverse, allergic, or other untoward reactions when administered to an animal or a human. For example, an expression vector may be formulated with a pharmaceutically acceptable carrier. As used herein, "pharmaceutically acceptable carrier" includes solvents, buffers, solutions, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents and the like acceptable for use in formulating pharmaceuticals, such as pharmaceuticals suitable for administration to humans. Methods for the formulation of compounds with pharmaceutical carriers are known in the art and are described in, for example, in Remington's Pharmaceutical Science, (17th ed. Mack Publishing Company, Easton, Pa. 1985); and Goodman & Gillman's: The Pharmacological Basis of Therapeutics (11th Edition, McGraw-Hill Professional, 2005); the disclosures of each of which are hereby incorporated herein by reference in their entirety. In some embodiments, the pharmaceutical compositions may comprise the vector in an amount of from about 0.1% to about 99.9% by weight. Pharmaceutically acceptable carriers suitable for inclusion within any pharmaceutical composition include water, buffered water, saline solutions such as, for example, normal saline or balanced saline solutions such as Hank's or Earle's balanced solutions), glycine, hyaluronic acid etc. The pharmaceutical composition may be formulated for parenteral administration, such as intravenous, intramuscular or subcutaneous administration. Pharmaceutical compositions for parenteral administration may comprise pharmaceutically acceptable sterile aqueous or non-aqueous solutions, dispersions, suspensions or emulsions as well as sterile powders for reconstitution into sterile injectable solutions or dispersions. Examples of suitable aqueous and non- aqueous carriers, solvents, diluents or vehicles include water, ethanol, polyols (such as glycerol, propylene glycol, polyethylene glycol, etc.), carboxymethylcellulose and mixtures thereof, vegetable oils (such as olive oil), injectable organic esters (e.g. ethyl oleate). The pharmaceutical composition may be formulated for any suitable route of administration, such as intravenous, subcutaneous, oral, intramuscular, intraperitoneal, pulmonary, intracranial, intraosseous, buccal, or nasal administration. The pharmaceutical compositions may comprise a vectors disclosed herein in an encapsulated form. For example, the vector may be encapsulated within a nanocapsule, such as a nanocapsule comprising one or more biodegradable polymers such as polylactide- polyglycolide, poly(orthoesters) and poly(anhydrides). In some embodiments, the vectors are encapsulated within polymeric nanocapsules. In other embodiments, the vectors are encapsulated within biodegradable and / or erodible polymeric nanocapsules. In some embodiments, the polymeric nanocapsules are comprised of two different positively charged monomers, at least one neutral monomer, and a crosslinker. In some embodiments, the nanocapsules further comprise at least one targeting moiety, such as an antibody. In some embodiments, the targeting moiety targets any one of a human hematopoietic stem cell CD marker including, for example, CD34. Methods of treatment By way of example, a vector of the present disclosure comprising a modified UCOE and a BTK transgene expression cassette can be administered to a subject in need thereof to restore or provide a normal level of expression of BTK in cells that normally express BTK, to thereby assist in promoting B cell survival, proliferation and / or differentiation, and / or to treat XLA or to inhibit or alleviate one or more symptoms characteristic of, or associated with XLA. In some embodiments, a population of host cells, such as hematopoietic stem cells, is transduced ex vivo with a vector of the present disclosure, said population of cells being subsequently introduced into a subject in need thereof. Accordingly, provided herein is a method for promoting B cell survival, proliferation and / or differentiation in a subject in need thereof, the method comprising administering to the subject an effective amount of a vector of or a cell of the present disclosure, and optionally measuring B cell survival, proliferation and / or differentiation in the subject or in a biological sample obtained from the subject. Also provided herein is a method for the treatment of XLA, or for inhibiting or ameliorating at least one symptom of XLA, comprising administering to a subject in need thereof an effective amount of a vector or a cell of the present disclosure, and optionally measuring an improvement in the progression of XLA or at least one symptom thereof in the subject or in a biological sample obtained from the subject. Also provided is the use of a vector or cell of the present disclosure in the manufacture of a medicament for promoting B cell survival, proliferation and / or differentiation. Also provided is the use of a vector or cell of the present disclosure in the manufacture of a medicament for the treatment of XLA, or for inhibiting or ameliorating at least one symptom of XLA. In some embodiments, treatment with the vectors or transduced host cells described herein genetically corrects or alleviates one or more of the symptoms or pathologies associated with XLA, including for example the susceptibility to infectious diseases, recurrent bacterial and viral infections of, for example, the lungs, skin, sinuses, ears, respiratory tract, urinary tract and gastrointestinal tract, and chronic diseases such as chronic lung disease. In the context of the treatment of, or alleviation of the symptoms or pathologies associated with XLA, the treatment of a subject may include identifying a subject in need of treatment thereof; transducing hematopoietic stem cells (e.g. autologous hematopoietic stem cells, allogenic hematopoietic stem cells, sibling matched hematopoietic stem cells) with a lentiviral vector of the present disclosure; and transplanting or administering the transduced hematopoietic stem cells into the subject. In some embodiments, the subject in need of treatment thereof is one suffering from the pathologies associated with XLA. In some embodiments, the method further comprises a step of myeloablative conditioning prior to the administration of the transduced hematopoietic stem cells (e.g. using a purine analog, chemotherapy, radiation therapy, treatment with one or more internalizing immunotoxins or antibody-drug conjugates, or any combination thereof). In some embodiments, the method further comprises the step of pre-conditioning, or in vivo chemoselection, utilizing a purine analog (e.g. 6TG) following administration of the transduced hematopoietic stem cells. In some embodiments, the method further comprises the step of negative selection utilizing a dihydrofolate reductase inhibitor (e.g. MTX or MPA) should side effects arise, such as graft versus host disease. In some embodiments, the method of treatment comprises the additional steps of (i) conditioning prior to hematopoietic stem cell transplantation; and / or (ii) in vivo chemoselection. One or both steps may utilize a purine analog. In some embodiments, the purine analog is selected from the group consisting of 6-thioguanine (6TG), 6-mercaptopurine (6MP) or azathiopurine (AZA). In some embodiments, following the collection of hematopoietic stem cells from a donor, the cells are transduced with a vector according to the present disclosure. In parallel, a patient to receive the hematopoietic stem cells is first treated with a myeloablative conditioning step. Following conditioning, the transduced hematopoietic stem cells are transplanted or administered to the patient. In some embodiments, the hematopoietic stem cells are administered between about 24 and about 96 hours following treatment with the conditioning regimen. In other embodiments, the patient is treated with the hematopoietic stem cell graft between about 24 and about 72 hours following treatment with the conditioning regimen. In yet other embodiments, the patient is treated with the hematopoietic stem cell graft between about 24 and about 48 hours following treatment with the conditioning regimen. In some embodiments, the hematopoietic stem cell graft comprises between about 2 x 106cells / kg to about 15 x 106cells / kg (body weight of patient). In some embodiments, the hematopoietic stem cell graft comprises a minimum of 2 x 106cells / kg, with a target of greater than 6 x 106cells / kg. In some embodiments, at least 10% of the cells administered are transduced with a lentiviral vector as described herein. In some embodiments, at least 20% of the cells administered are transduced with a lentiviral vector as described herein. In some embodiments, at least 30% of the cells administered are transduced with a lentiviral vector as described herein. In some embodiments, at least 40% of the cells administered are transduced with a lentiviral vector as described herein. In some embodiments, at least 50% of the cells administered are transduced with a lentiviral vector as described herein. The present disclosure also contemplates combination therapies whereby active agents such as antibacterial, antifungal, and / or antiviral active pharmaceutical agents are administered prior to, during, or following the administration or transplantation of transduced hematopoietic stem cells into a subject in need of treatment. Agents in combination therapies may be administered concurrently. By concurrent administration is meant that the agents are administered in a single composition or as separate compositions by the same or different routes either contemporaneously, simultaneously or sequentially within a short enough period of time that the effective result is equivalent to that obtained when all such agents are administered as a single composition. By “simultaneously” is meant that the agents are administered at substantially the same time, and desirably together in the same formulation. By “contemporaneously” it is meant that the agents are administered closely in time, e.g., one agent is administered within from about one minute to within about one day before or after another. Any contemporaneous time is useful. However, it will often be the case that when not administered simultaneously, the agents will be administered within about one minute to within about eight hours and suitably within less than about one to about four hours. When administered contemporaneously, the agents are suitably administered at the same site on the subject. The term “same site” includes the exact location, but can be within about 0.5 to about 15 centimeters, preferably from within about 0.5 to about 5 centimeters. The term “separately” as used herein means that the agents are administered at an interval, for example at an interval of about a day to several weeks or months. The agents may be administered in either order. The term “sequentially” as used herein means that the agents are administered in sequence, for example at an interval or intervals of minutes, hours, days or weeks. If appropriate the agents may be administered in a regular repeating cycle. Exemplary embodiments The following sets forth exemplary embodiments of the invention: 1. A vector suitable for the treatment of X-linked agammaglobulinemia (XLA) or for inhibiting or ameliorating at least one symptom of XLA, the vector comprising: (a) a transgene expression cassette comprising a nucleotide sequence encoding Bruton's tyrosine kinase (BTK) operably linked to a promoter; (b) at least one regulatory or enhancer element; (c) a ubiquitous chromatin opening element (UCOE), wherein the UCOE comprises a nucleotide sequence at least about 90%, 91%, 92%, 93%, 94% or 95% identical to the sequence set forth in SEQ ID NO:1, and wherein the sequence of the UCOE comprises the following nucleotide substitutions relative to SEQ ID NO:1, at positions corresponding to those of SEQ ID NO:1: (i) a G residue at position 4; (ii) the sequence CGAAC at positions 347-351; (iii) the sequence CGCGC at positions 362-366; (iv) a T residue at position 385; (v) a G residue at position 496; (vi) the sequence CGCGC at positions 612-616; and (vii) a T residue at position 673. 2. The vector of embodiment 1, wherein the nucleotide sequence encoding BTK is codon optimized for expression in humans. 3. The vector of embodiment 1 or 2, wherein the nucleotide sequence encoding BTK comprises the sequence set forth in SEQ ID NO:4 or SEQ ID NO:5. 4. The vector of any one of embodiments 1 to 3, wherein the promoter in the transgene expression cassette is a B cell specific promoter. 5. The vector of any one of embodiments 1 to 4, wherein the promoter in the transgene expression cassette is the BTK promoter, optionally comprising the sequence set forth in SEQ ID NO:6. 6. The vector of any one of embodiments 1 to 5, wherein the transgene expression cassette comprises the sequence set forth in SEQ ID NO:7. 7. The vector of any one of embodiments 1 to 6, wherein the UCOE is in the forward orientation in the vector with respect to the transgene expression cassette. 8. The vector of any one of embodiments 1 to 7, wherein the UCOE and the transgene expression cassette comprise the sequence set forth in SEQ ID NO:8. 9. The vector of any one of embodiments 1 to 8, wherein the UCOE comprises the nucleotide sequence of SEQ ID NO:2, or a sequence at least about 90% identical thereto, preferably at least about 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98.5%, 99% or 99.5% identical thereto. 10. The vector of any one of embodiments 1 to 9, wherein the UCOE is at least about 0.7kb in length. 11. The vector of any one of embodiments 1 to 10, wherein the UCOE comprises or consists of the nucleotide sequence of SEQ ID NO:2. 12. The vector of any one of embodiments 1 to 11, wherein the at least one regulatory element is an insulator, a Woodchuck Hepatitis Virus (WHV) Posttranscriptional Regulatory Element (WPRE), or a combination thereof, optionally wherein the regulatory element is located downstream of the transgene expression cassette. 13. The vector of any one of embodiments 1 to 12, wherein the vector is a viral vector. 14. The vector of embodiment 13, wherein the vector is a retroviral vector. 15. The vector of embodiment 14, wherein the vector is a lentiviral vector. 16. The vector of embodiment 15, comprising a sequence set forth in SEQ ID NO:20 or a sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98.5%, 99% or 99.5% sequence identity thereto, comprising the UCOE from position 2396 to 3068 of SEQ ID NO:20. 17. The vector of embodiment 15, comprising a sequence set forth in SEQ ID NO:21 or a sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98.5%, 99% or 99.5% sequence identity thereto, comprising the UCOE from position 2390 to 3062 of SEQ ID NO:21. 18. A cell comprising, or transduced with, the vector of any one of embodiments 1 to 17. 19. The cell of embodiment 18, wherein the cell is a B cell, a myeloid cell or a hematopoietic stem cell. 20. The cell of embodiment 19, wherein the hematopoietic stem cell is a CD34+hematopoietic stem cell. 21. A method for promoting B cell survival, proliferation and / or differentiation in a subject in need thereof, the method comprising administering to the subject a vector of any one of embodiments 1 to 17 or a cell of any one of embodiments 18 to 20, and optionally measuring B cell survival, proliferation and / or differentiation in the subject or in a biological sample obtained from the subject. 22. The method of embodiment 21, further comprising the step, prior to administration, of identifying the subject as one that would benefit from receiving a therapy to promote B cell survival, proliferation and / or differentiation. 23. A method of treating X-linked agammaglobulinemia (XLA) or inhibiting or ameliorating at least one symptom thereof, in a subject, the method comprising administering to the subject a vector of any one of embodiments 1 to 17 or a cell of any one of embodiments 18 to 20, and optionally measuring an improvement in the progression of XLA or at least one symptom thereof in the subject or in a biological sample obtained from the subject. 24. The method of embodiment 23, further comprising the step, prior to administration, of identifying the subject as one that would benefit from receiving a therapy to treat X-linked agammaglobulinemia (XLA) or to inhibit or ameliorate at least one symptom thereof. 25. The method of any one of embodiments 21 to 24, wherein the cell is administered by adoptive cell transfer. 26. Use of the vector of any one of embodiments 1 to 17 or the cell of any one of embodiments 18 to 21 in the preparation of a medicament for promoting B cell survival, proliferation and / or differentiation, or for the treatment of XLA or the inhibition or amelioration of at least one symptom thereof. In order that the invention may be readily understood and put into practical effect, particular preferred embodiments will now be described by way of the following non- limiting examples. The reference in this specification to any prior publication (or information derived from it), or to any matter which is known, is not, and should not be taken as an acknowledgment or admission or any form of suggestion that that prior publication (or information derived from it) or known matter forms part of the common general knowledge in the field of endeavour to which this specification relates. EXAMPLES EXAMPLE 1 - Vectors Figure 2 and Table 2 below outline vectors described herein and used in the following examples. The nucleotide sequences of exemplary vectors pRRL-XLA-LV and TL20c- XLA-LV are provided in SEQ ID NOs:20 and 21, respectively. Table 2 In the nucleotide sequence of exemplary vector pRRL-XLA-LV as shown in SEQ ID NO:20, the modified 0.7UCOE (SEQ ID NO:2) is located from position 2396-3068, the BTK promoter (pBTK; SEQ ID NO:6) is at positions 3105-3941, the coding sequence of the codon- optimized BTK gene (coBTK; SEQ ID NO:4) is at positions 3957-5936, and the WPRE (SEQ ID NO:9) is at positons 5943-6531. In the nucleotide sequence of exemplary vector TL20c-XLA-LV as shown in SEQ ID NO:21, the modified 0.7UCOE (SEQ ID NO:2) is located from position 2390-3062, the BTK promoter (pBTK; SEQ ID NO:6) is at positions 3099-3935, the coding sequence of the codon- optimized BTK gene (coBTK; SEQ ID NO:4) is at positions 3951-5930, and the WPRE (SEQ ID NO:9) is at positions 5937-6526. The lentivirus particles were produced essentially as described previously. For pRRL-XLA LV, production was performed by transient transfection as described in Seymour et al. 2021, Molecular Therapy: Methods & Clinical Development 20: 635-651; Kerns et al. 2010, Blood 115(11):2146-2155; and International Patent Publication No. WO2018195297. For TL20c-XLA LV, production utilised a stable producer cell line as described in Throm et al. 2009, Blood 113:5104–5110, International Patent Publication No. WO2016183260, International Patent Publication No. WO2023187691. EXAMPLE 2 – Gene therapy in mouse model of XLA (mXLA mice) A mouse model of XLA (mXLA) was used to evaluate the safety and efficacy of lentiviral (LV) vectors #3365 and pRRL-XLA-LV (#3366) in mXLA primary and secondary recipient mice. Vector UCOE.pBTK.coBTK (#1419; LVXLA1) containing an unmodified 0.7UCOE was used for comparison. Recipients receiving mock-transduced mXLA (KO Mock) or wild-type hematopoietic stem and progenitor cells (HSPCs) (WT Mock) represented negative and positive controls, respectively. Parameters of interest included: restored B cell development and function, stable reconstitution of BTK expression in serial transplant, methylation status of the LV BTKp and safety, as assessed by general health screening and immunophenotyping hematopoietic tissues. The experimental timeline for the study is shown in Figure 3. Bone marrow was isolated from donor mice, then enriched for lineage-negative cells (Lin-). The isolated Lin- cells (mHSC) were transduced with or without the lentiviral (LV) constructs in SCGM media supplemented with murine SCF and TPO cytokines. The amount of lentivirus used was 10% of culture volume for vectors 3365, 3366 and 1419 and 20% of culture volume for BTKp.coBTK. The mock-transduced negative control and the WT positive control cells were cultured without virus in the same manner. 16-17 hours post LV- or mock-transduction time point, cells were washed and resuspended in saline for injection into the recipient mice. Primary recipient mice received one 9.0 Gy dose of total body irradiation. Following irradiation, the LV transduced or control mHSCs were washed then resuspended with PBS, and 2 million cells were transplanted by retro-orbital (r.o.) injection into the lethally irradiated mXLA recipients. Donor mHSCs were assayed for viral copy number (VCN) using cells cultured in methocult for 7 days after transduction and VCN determined using digital droplet PCR (ddPCR). Results of lentiviral transduction (all with protamine sulfate with Lentiboost (1:100) and dmPGE2 (20µM)) are shown in Table 3. Table 3 At 12 weeks post-transplant, lentiviral gene therapy (LV-GT) recipients were immunized with the T cell-dependent antigen NP-CGG in alum, and serum was collected prior to and 10 days after primary immunization. One month following primary challenge, mice were re-challenged with NP-CGG in phosphate-buffered saline (PBS) and serum was collected 10 days after challenge. NP-specific IgG and IgM, as quantified by enzyme-linked immunosorbent assay (ELISA). Safety parameters were determined by regular health checks and at the experimental endpoint, extensive immunophenotyping and viral copy number of hematopoietic tissues were assessed. The efficacy of the lentiviral therapy was assessed as serum antibody responses to T-dependent antigens after 12-weeks post-transplant, and at the experimental endpoint by immunophenotyping (including BTK expression) and enumerating the B cell compartment in post-mortem lymphoid tissues. To assess sustained B cell rescue and BTK expression, 106bone marrow cells (without lineage depletion) from the primary recipients was transplanted into irradiated secondary recipients to drive the cycling of mHSCs and improve the chances of detecting clonal expansion of hematopoietic cells. Secondary recipient mice were monitored for 17 weeks. The methylation status of the proviral BTK promoter from LV- treated primary recipient bone marrow was also assessed. Comparison of percent B cell rescue using alternative LV constructs in mXLA gene therapy at 10-week post-transplant peripheral blood LVs were used to express human BTK cDNA driven by the BTK promoter (BTKpBTK) and variations of the vector containing a 0.7 UCOE (1419, 3365 and 3366). LV gene therapy-treated mice were analyzed for % B cells (% B220+) by flow cytometry from peripheral blood at 10 weeks post-transplant. As shown in Figure 4, LV vectors 1419, 3365 and 3366 had improved B cell percentages compared to mXLA mock. However BTKpBTK LV, was less effective. Comparison of B cell rescue using alternative LV constructs in mXLA gene therapy at primary transplant endpoint BTKpBTK and variations of the vector containing a 0.7 UCOE (1419, 3365 and 3366) from LV gene therapy-treated mice were analyzed at 28 weeks post-transplant by flow cytometry. Figure 5 shows reconstitution of B cells numbers in spleen (%B220+cells * total splenocyte count), and percentage of B cells in bone marrow and spleen (%B220+of total lymphocyte gate). Total splenic mature B cell numbers are similar for all UCOE vectors and equivalent to WT mock control. Only BTKpBTK vector failed to rescue B cell numbers. Proportions of B cells in the spleen and bone marrow were similar for all UCOE containing vectors and similar to WT mock control levels in both bone marrow and spleen. All UCOE containing vectors restore in vivo B cell responses to T-dependent immunogen and total IgG compared to mXLA mock LV-GT-treated mXLA mice were immunized with NP-CGG in alum at 12 weeks post-transplant and boosted 1 month later. Levels of NP-specific IgG in serum was measured by ELISA and quantified relative to an IgG standard. High-affinity NP-IgG was measured from serum prior to and 10 days after primary immunization (1o). One month following primary challenge, mice were re-challenged with NP-CGG in PBS and serum was collected 10 days later (2o). Differences between experimental and mXLA mock cohorts for both the primary and secondary antibody responses are shown in Figure 6A. Total serum IgG in serum from mice receiving LV-GT was measured by ELISA at endpoint analysis (28 weeks post-transplant). Differences between experimental and mXLA mock cohorts are shown in Figure 6B. NP-specific IgG, as quantified by enzyme-linked immunosorbent assay (ELISA), revealed increased production of high-affinity NP-IgG in response to primary and secondary immunization (Figure 6A). Mice receiving LV-GT vectors showed high-affinity NP-specific IgG responses to secondary challenge equivalent to those of WT mock control animals. Total IgG, for all experimental groups, as quantified by ELISA, showed an increase over mXLA mock, and the average amount of total IgG in the 1419, 3365, 3366, and BTKpBTK groups were equivalent to WT mock control animals (Figure 6B). VCN in input cells and in bone marrow and spleen from primary recipients To determine the viral copy number (VCN) of the input cells, mouse CFU media was inoculated with 10,000 cells 16-17 hours post-transduction or mock transduction and cultured for 7 days. The colonies were then harvested (2 plates per condition for separate replicates) and gDNA extracted, then VCN determined using ddPCR (see Figure 7A). Genomic DNA (gDNA) was isolated from total bone marrow (Figure 7B) and (Figure 7C) spleen at endpoint analysis (29 weeks post-transplant), and VCN per cell was quantified by ddPCR to determine the average number of vector copies per cell in each sample. All UCOE-containing BTK constructs achieved functional rescue at clinically relevant VCN in bone marrow and SP. VCN in primary recipients was higher than input of ~1, likely reflecting positive selection for BTK expression throughout B cell development (bone marrow cells include pre-B, immature and mature recirculating B cells which require BTK for development and survival) and spleen includes multiple mature B cell populations. The progressive increase in VCN in spleen relative to bone marrow reflects reflect positive selection. Methylation of BTKpBTK and LV with unmodified and optimized UCOE Methylation of BTKp within the integrated LV sequences in bone marrow cells as quantified by bisulfite sequencing. Data are shown in Figure 8 as a percentage of methylated CpGs within the BTKp for LV with unmodified UCOE (1419), LV with optimized UCOE (3366) and LV without UCOE (BTKpBTK). Vectors containing UCOE were effective in eliminating methylation of the BTKp. BTKp sequences derived from BTKp.BTK LV was substantially methylated (41%), while minimal methylation (1.7% and 1.8%) was observed for the LV with unmodified UCOE (1419) and LV with optimized UCOE (3366) respectively. Schematic of Transplanting secondary recipients A schematic for the transplantation of secondary recipient mice is shown in Figure 9. For secondary transplants, 5 X 106bone marrow cells (without lineage depletion) from WT mock, mXLA mock, or LV-GT primary recipient mice were transplanted into irradiated mXLA secondary recipient mice with 1 secondary recipient / donor. Mice were monitored for 17 weeks and immunophenotyping and VCN analysis performed at endpoint. LV with unmodified and optimized UCOE mediate sustained BTK expression and B cell rescue in secondary mXLA recipients Hematopoietic cells were collected from the bone marrow and spleen of secondary recipients (which had received bone marrow cells from primary recipient mice of BTKpBTK, 1419, 3366 LV-GT or mock wild type and mXLA controls) and analyzed for B cell rescue and BTK expression. Figure 10 shows the percentage of B cells (B220+ of singlets) and % BTK+ B cells (B220+ BTK+) at 17 weeks post-transplant in bone marrow (A-B) and spleen (C-D) as determined by flow cytometry analysis. Optimized UCOE LV (3366) mediated sustained BTK expression and rescues B cell numbers at clinically relevant VCN in secondary recipient mice demonstrating sustained LV transduction and functional correction on long- term HSC. In contrast to UCOE LV (3366) treated mice, recipients of BTKpBTK uninsulated vector exhibited much lower BTK expression and % B cells in bone marrow and spleen tissues, despite a much higher VCN. VCN in bone marrow and spleen from secondary recipients Genomic DNA (gDNA) was isolated from total bone marrow and spleen (B) at endpoint analysis (17 weeks post-transplant), and VCN per cell was quantified by ddPCR to determine the average number of vector copies per cell in each sample (Figure 11). VCNs are similar for 1419 and 3366 and similar to the VCN in primary recipient mice. VCN for BTKpBTK was much higher, as seen historically. In summary of the mXLA in vivo study described in this Example, lentiviral GT with pRRL-XLA LV (#3366) resulted in: stable engraftment of LV-treated HSPCs; rescue of B cell development at clinically relevant VCN in bone marrow and spleen; restoration of antibody responses to T-dependent antigens; and protection from CpG methylation in primary recipients. EXAMPLE 3 – In vivo LVXLA2 gene therapy in XLA humanized mice The inventors next investigated lentiviral gene therapy in XLA humanized mice using the LVXLA2 (TL20c-XLA-LV) vector to determine if LVXLA2 transduction improved the ability of XLA HSPCs to produce mature B cells in this mouse model and to assess safety. The inventors used a humanized mouse model previously described in Seymour et al (PMID: 33718514), and tested the efficacy of LVXLA2 gene therapy in XLA patient cells for rescue of B cell development. XLA patient HSPCs were either mock-transduced or transduced with LVXLA2 in vitro, then transplanted into recipient NBSGW mice and compared in parallel to a cohort of mice receiving healthy donor (HD) HSPCs. The immunophenotype and VCN were compared between all groups. The experimental timeline of mouse humanization with LVXLA2 transduced human HSPCs and analysis is shown in Figure 12. Cryopreserved CD34+ enriched apheresis products from adult male XLA subject (G-CSF mobilized) and healthy donor (HD; G-CSF- mobilized) were thawed and prestimulated, then cultured with or without LVXLA2 at 15 MOI in the presence of Poloxamer F127 and dmPGE2. One day following transduction, the HSPCs (2.0e6 per mouse) were transplanted directly into busulfan-conditioned NBSGW mice by r.o. injection. In addition, ~150-250 cells from each sample were placed in MethoCult for colony- forming unit (CFU) assays and cultured for 1-and 2- weeks. gDNA was extracted from individual CFU colonies and VCNs were determined by digital droplet PCR (ddPCR). An MOI of 15 yielded an average VCN / colony of 1.6 for HD and 2.3 for XLA. Mice were monitored closely for health status post-transplant until the experimental endpoint (16 weeks). At experimental endpoint, extensive immunophenotyping and viral copy number of hematopoietic tissues were assessed. Human CD45+ engraftment in recipient mice at in vivo endpoint The proportions of human CD45 (hCD45) cells within the lymphocyte gate among cells collected from the bone marrow and spleen of humanized mice 16-weeks post- transplant were determined by flow cytometry. The results are shown in Figure 13. Engraftment of human CD45+cells was similar between LV-transduced and mock-transduced cells in both healthy donor and XLA patient engrafted mice implying that enforced expression of BTK using the LVXLA2 vector did not impact HSPC engraftment and that HD and XLA patient cells engrafted at similar levels. Three animals exhibited reduced engraftment due to technical problems with the initial transplantation of HPSCs. Lineage characterization of bone marrow cells at in vivo endpoint The proportions of hCD45 cells within the lymphocyte gate that are CD19+B cells, CD4 / CD8+ T cells and CD33+ myeloid cells in bone marrow of humanized mice 16- weeks post-transplant were also determined by flow cytometry. The results are shown in Figure 14. The percentage of human CD45 cells that were CD19+B cells was significantly lower in mock treated XLA- vs HD-humanize mice; consistent with the reduction of B cells in mock-XLA treated mice, the relative proportion of T cells and myeloid cells was higher in mock-XLA treated mice in comparison with mock HD humanized mice. LVXLA2 gene therapy in XLA patient cells significantly improved proportion of B cells in comparison with mock- transduced XLA engrafted mice, and accordingly lead to proportional decreases the T cells and myeloid cells to levels closer to healthy donor levels. Evidence that LVXLA2 gene therapy alleviates the pre-B cell developmental block in the bone marrow The proportion and number of B cell developmental subsets in the bone marrow of humanized mice 16-weeks post-transplant were determined by flow cytometry and are shown in Figure 15A and Figure 15B, respectively. Specifically shown in Figure 15A are the proportion of precursor cells, Pro-B cells, Pre-B cells and Immature B cells in the CD4- , CD8-, CD33- lymphocyte gate of bone marrow cells. Markers used to specify B cell subsets were:Precursor (CD45+CD19-CD34+); Pro B-Cell (CD19+CD34+); Pre-B cell (CD19+CD179a+); immature B cells (CD19+CD179a-IgM+). B cell defects observed in the bone marrow of mice receiving mock-transduced XLA HSPCs were ameliorated by LVXLA2 transduction. Specifically, LVXLA2 transduction increased the fraction of XLA CD45+cells that were CD19+to proportions towards what is seen with HD-humanized mice (Figure 15A). As expected, LVXLA2 treatment resolved the pre-B cell developmental block phenotype in XLA humanized mice, observed as a reduction in the fraction of pre-B cells and an increase in the fraction and absolute numbers of immature IgM+B cells relative to mice receiving mock- transduced XLA HSPCs. Lineage characterization of spleen cells at in vivo endpoint The proportions of hCD45 cells within the lymphocyte gate that are CD19+B cells, CD4 / CD8+ T-cells and CD33+ myeloid cells in the spleens of humanized mice 16-weeks post-transplant were determined by flow cytometry, as shown in Figure 16. Mock-treated XLA humanized mice had a smaller fraction of CD19+human leukocytes and an increase in T- cells and Myeloid cells in the spleen compared with mock-treated HD humanized mice. LVXLA2 gene therapy enabled XLA-humanized mice to generate CD19+B cells at a proportion more similar to mice humanized with HD HSPCs as well as a decrease in T cell and myeloid cells trending towards HD HSPCs. LVXLA2 gene therapy restores mature human B cell compartments in the spleen of XLA humanized mice The proportion of B cell subsets in spleen of humanized mice 16 weeks post- transplant were determined by flow cytometry, as shown in Figure 17. Specifically shown are the proportion of Pre-B, transitional and mature naïve cells in the CD19+lymphocyte gate of the spleen cells (Figure 17A) and the absolute counts of indicated B cell subsets (Figure 17B). Markers used to specify B cell subsets were: Pre-B (CD179a+in CD19+gate); transitional B (CD24+CD38+in CD19+gate); and mature naïve cells (CD24-CD38-). LVXLA2 gene therapy reduced the proportion of pre-B cells (and absolute pre-B cell numbers) localized in the spleen relative to mock-transduced XLA humanized mice consistent with the alleviation of the bone marrow pre-B cell developmental block. Importantly, LVXLA2 gene therapy improved the % and absolute count of mature naïve B cell compartment in XLA humanized mice. VCN of input LVXLA2-transduced HSPC cells and hematopoietic tissues at in vivo endpoint Shown in Figure 18A are “input” cell VCN from LVXLA2-transduced healthy donor and XLA HSPC transplanted into NBSGW mice determined by ddPCR of gDNA collected from CFU single colonies 14-days post-transduction. VCN from bone marrow (Figure 18B) and spleen (Figure 18C) of humanized mice 16-weeks post-transplant were determined by ddPCR. For the input cells, the VCN averaged 1.7 and 2.5 viral copies per colony for LVXLA2- transduced HD and XLA cells respectively (Figure 18A). After 16-weeks post-transplant, the VCN of mice receiving HD cells transduced with LVXLA2 averaged 2.92 viral copies per cell in bone marrow and 2.48 viral copies / cell in spleen, while the average VCN of mice receiving XLA cells transduced with LVXLA averaged 1.26 viral copies / cell and 1.69 viral copies / cell for bone marrow and spleen respectively (see Figure 18B and Figure 18C). The humanized model of XLA used in this Example recapitulates key features of the human disease including the pre-B cell developmental block in the bone marrow, the lack of mature peripheral B cells, the inability to flux calcium in response to BCR engagement, and the lack of serum antibodies. Using this model, the inventors have shown that HSPCs from an XLA patient can be transduced using LVXLA2 to achieve a clinically relevant VCN and demonstrate that gene therapy with LVXLA2 alleviates the B cell developmental deficits identified in XLA humanized mice. These data support the use of the optimized LVXLA2 vector for clinical studies. EXAMPLE 4 – BTK knock out and potency assays Figure 19 illustrates a schematic of the steps used by the inventors to generate and characterize a BTK knock-out B cell line. The Ramos B cell line was chosen due to its intact BCR to Calcium flux signaling axis. Ramos B cells were electroporated with TALEN mRNA then incubated with an AAV6 engineered to contain a BTK-specific donor template. The resulting cells were characterized by flow cytometry to analyze GFP positivity and subsequent BTK staining to confirm the BTK loss. Single-cell clones were next obtained by single cell sorting and culturing for several weeks. GFP positivity and the lack of BTK expression was determined for each clone by flow cytometry. We chose one BTK-KO clone (stably expressing GFP and lacking BTK) for further testing. The BTK KO clone was transduced with LVXLA2 vector at one of three MOIs to generate a range of vector copy numbers (VCN). After assessing BTK expression in the transduced clones, LVXLA2 transduced BTK-KO clone cells were stimulated with IgM, and phosphoPLCg2 was analyzed by intracellular flow cytometry. Homology directed repair was then used to knock in GFP into exon2 of the BTK gene (see Figure 20). A GFP cDNA under the control of the MND promoter was inserted to replace most of BTK exon 2 by HDR as indicated in the schematic. Alleles with successful HDR editing have disrupted (absent) BTK expression, but constitutive GFP expression. BTK KO Ramos clone 1 expresses GFP but not BTK BTK KO Ramos clone 1 cells were assessed by flow cytometry in comparison with the parental (unedited) Ramos B cell line for GFP expression and intracellular BTK within the lymphocyte gate (see Figure 21). These data demonstrate sustained loss of BTK expression following gene editing of Ramos B cells in BTK KO Ramos clone 1. BTK expression increases with lentiviral copy number in LVXLA2-transduced BTK KO Ramos clone-1 cells GFP and BTK expression was determined in LVXLA2-transduced BTK KO Ramos clone-1 cells as shown in Figure 22. These data demonstrate restoration of BTK expression following LVXLA2 treatment of BTK KO Ramos B cells. Restoration of endogenous levels of BTK is achieved with a VCN of 1-3. Intracellular phospho-PLCγ2 levels increase in response to IgM stimulation and correlate with LVXLA2 copy number in BTK KO Ramos clone1 cells BTK dependent phosphorylation of PLC^2 (in response to BCR engagement) is required for B cell activation and downstream signals that lead to NFKB activation and calcium flux. Restoration of BTK dependent PLC^2 phosphorylation therefore provides a sensitive assay for restoration of BTK dependent signaling in human B cells. The inventors investigated intracellular phospho-PLC^2 levels in response to IgM stimulation in LVXLA2- transduced BTK KO Ramos clone-1 cells (see Figure 23). These data demonstrate restoration of BTK dependent p-PLCg2 following LVXLA2 treatment of BTK KO Ramos B cells. Restoration of endogenous levels of p-PLCg2 is achieved with a VCN of 1-3. EXAMPLE 5 – In vitro immortalization (IVIM) Assay to assess safety of LVXLA2 The In Vitro Immortalization (IVIM) Assay is designed to evaluate the potential for genotoxicity by assessing the replating frequency (a measure of immortalization) of murine Lin- hematopoietic stem cells following integrating gene transfer. Lack of insertional mutants in this assay following transduction with LVXLA2 would provide strong support for safety of this LV design. A γ retroviral vector was used as a positive control as this vector backbone consistently leads to a significant increase in immortalization. Mock-transduced cells are used as negative controls. A schematic of the IVIM assay as employed herein is shown in Figure 24. Table 4 below shows the results of the VCN / dg and flow cytometry measurements for the vectors containing a reporter gene. The VCN / dg was measured on day 4 after transduction for all assays. For an IVIM assay to be valid, the mean VCN / dg must be above 1. A meta-analysis of RSF91 transductions revealed that with a starting amount of 1x105cells, the positive control vector only reliably scored positive if the VCN / dg was above 1. The mCherry or EGFP transgene expression for RSF91 was also determined on day 4 by flow cytometry. If cell proliferation is normal, day 4 is the first time point when proliferation reduces the number of vector-derived episomal elements (circular DNA) enough to yield reproducible mean copy numbers and early enough to exclude vector-induced clonal skewing influencing the overall VCN / dg and transduction efficiency. Table 4 The inclusion criterion for genotoxicity analysis in IVIM is a mean VCN / dg above one, preferably above three. Vector doses (cumulative multiplicity of infection = c.MOI) of 60 were used for the positive control RSF91 resulting in a mean VCN of 7.7 ± 1.7 copies. Based on previous experience with SIN-lentiviral vectors, the inventors used c.MOIs of 200 for the test vectors. For vector LVXLA2, this resulted in VCN / dg levels of 16.1 ± 6.8. Proliferation rate of samples at different days in the IVIM assay were determined (Figure 25). There were no differences in the proliferation behavior relative to the current mock samples (analyzed by Kruskal-Wallis test with Dunn's correction). No difference in proliferation between MOCK or RSF91 controls and meta proliferation data of non-transduced cells was observed. When cells were transduced with LVXLA2, proliferation was normal during the complete course of the assay. The data suggests no general toxic effect of the transgene and the vector supernatant. After the replating step on day 15, positive wells were scored microscopically and with a MTT clonal scoring system (see Table 5 below). C1 to C4 describe the different categories of cell growth: C1 and C2 wells were counted as positive wells; C4 were counted negative wells; C3 wells are either positive or negative, depending on the absorbance reading of the MTT measurement. If C3 was above the maximal MTT absorbance reading of the MOCK plate, the well was counted positive. If C3 was below this threshold, the well was counted as negative. Table 5 shows the number of positive wells, either microscopically identified as C1 or by MTT-assay as above threshold. The replating frequency was used to create Figure 26. For assay 220615_VD, the microscopic scoring showed no C1 or C2 wells for any of the 96-well plates; the MTT-assay results showed no wells above threshold for the positive control. Hence, the replating results for this assay are not valid and were excluded from Table 5 and Figure 26. Table 5 Replating Frequency (RF) of the control samples MOCK or RSF91 and the test vectors was compared to data of a meta-analysis for control samples (Mock-MA, RSF91- MA, lv-SF-MA) (see Figure 26). Differences in the incidence of positive and negative assays relative to Mock-MA or RSF91-MA were analyzed by Fisher's exact test with Benjamini- Hochberg correction (***P < 0.001; **P < 0.01; *P < 0.05; NS = not significant). MOCK control plates contained wells of microscopic category C3 to C4 in all assays. No C1 clones (the most obvious case of immortalization) were detected for MOCK or the test vectors. The positive control RSF91 showed C1 wells in 2 out of 9 plates. After the MTT-analysis, 4 of 9 positive control vector plates were above the Q1-quantification threshold. The incidence of plates with insertional mutants for RSF91 was significantly lower compared to the metadata of RSF91. For the test LVXLA2, no C1 wells were detectable. In the MTT-assay, three plates were above the detection limit and two above the Q1-quantification threshold. The difference in the incidence of positive plates between the metadata of the positive control and LVXLA2 was statistically significant. Hence, the LVXLA2 has a significantly lower genotoxic potential than the mutagenic positive control. Thus, the IVIM assay indicates that LVXLA2 appears safe and exhibits no significant increase in mutagenic risk in comparison to MOCK negative control treated cells. EXAMPLE 6 - SAFETY AND EFFICACY OF LVXLA2 GENE THERAPY IN A MOUSE XLA MODEL The safety and efficacy of LVXLA2 in ex vivo lentiviral gene therapy (LV-GT) of murine bone marrow HSCs was assessed in a mouse model of XLA (Btk- / - Tec- / - mice, scientific nomenclature B6:129S-Btktm1WkTectm1Welm, referred to hereafter as mXLA mice). The study used LVXLA2 (MOI = 18, using transduction enhancers protamine sulfate and poloxamer F127) to transduce lineage-depleted (linneg) bone marrow cells (murine HSC, or mHSC) obtained from mXLA mice. Mock-transduced mXLA mHSCs (mXLA Mock) or mock- transduced WT mHSCs from congenic C57Bl / 6 mice (WT Mock) were manufactured in parallel to serve as negative and positive control treatments for the gene therapy, respectively. Three experiments, each comprised of all three treatment groups, were performed using serial mHSC transplantation in lethally irradiated, recipient mXLA mice. Methodology Studies in Primary Transplant Recipient Mice mHSCs treated ex vivo with or without LVXLA2 transduction were transplanted into lethally irradiated primary mXLA mice (recipients). To differentiate donor vs. recipient cells during the in-life and post-mortem phases of the study, the donor cell strain had a different congenic CD45 marker than the recipient mice. In-life assessments included health (body weight and survival), and engraftment of hematopoietic cells in the peripheral blood (PB) in addition to antibody responses to antigenic challenge (assessed by serum ELISA). Scheduled euthanasia was performed 18- to 22-weeks post-transplant. Safety and efficacy parameters at the experimental endpoint included: CBCs, blood chemistry, histopathology, extensive immunophenotyping (including BTK expression) and viral copy number per cell (VCN) assessment of hematopoietic tissues. Additional assays of BTK function in B lymphocytes were performed, testing B cell proliferation in response to B cell receptor (BCR) stimulation. Studies in Secondary Transplant Recipient Mice Bone marrow (BM) hematopoietic cells collected from primary recipient mice were transplanted into irradiated secondary recipients to drive the cycling of mHSCs and to improve the chances of detecting clonal expansion of LV-transduced hematopoietic cells. This approach also permitted assessment of the stability of LV transduction of mHSCs and the capacity for sustained LV-mediated BTK expression and function. For secondary recipients, in- life assessments were health (body weight and survival) and engraftment of hematopoietic cells in the peripheral blood. Safety and efficacy parameters at the scheduled experimental endpoint (16-18-weeks post-transplant of the secondary recipients) included: CBCs, blood chemistry, histopathology, and extensive immunophenotyping (including BTK expression), and VCN assessment of hematopoietic tissues. For both primary and secondary recipients, safety was measured as the absence of abnormalities in overt health, blood counts, serum chemistry, tissue pathology, clonal dominance, or tumorigenesis that might be a result of the lentiviral vector. Efficacy was measured based upon sustained LV marking in engrafted hematopoietic cells, restoration of BTK expression at endogenous levels in mHSCs and across hematopoietic lineages, and improvement in immune hematopoiesis and function. Results There was no evidence of toxicity observed in LVXLA2 recipient animals. Specifically, there were no significant differences between recipients of LVXLA2-transduced mXLA mHSCs in comparison with Mock-transduced mXLA mHSCs or mock-transduced WT mHSCs recipient mice based on weight gain survival and donor chimerism or in GLP histopathology, complete blood count, or clinical chemistry studies. Primary and secondary recipients of LVXLA2-transduced mXLA mHSCs exhibited sustained viral making (VCN / cell) at clinically relevant and safe levels in BM and spleen (Figure 27). mXLA mice transplanted with LVXLA2-transduced mXLA mHSCs showed improved B cell hematopoiesis in the bone marrow (BM; Figure 27D, E) and spleen (Figure 27F, G). In the bone marrow, the XLA characteristic block in B cell development at the pre- pro B cell stage was observed in mice receiving Mock mXLA mHSCs, along with the resulting decrease in B cells at later stages of development (mature and immature B cells). These abnormalities were corrected in mice receiving LVXLA2-transduced mHSCs, leading to features nearly identical to WT Mock positive control recipients (Figure 27E). The proportion of splenic follicular mature B cells was also significantly increased by LVXLA2 treatment, compared with the mXLA Mock treatment group (Figure 27G). Recipients of LVXLA2-transduced mXLA mHSCs exhibited increased BTK expression in relevant hematopoietic cells within the bone marrow and spleen (Figure 28A- D) including mHSCs, B cells, monocytes, and neutrophils compared to mXLA Mock mHSC recipients. Notably, LVXLA2 did lead to expression of BTK in T cells, consistent with lack of endogenous BTK expression in this lineage. The amount of BTK expressed by LVXLA2 per cell, shown in Figure 28C-D as the mean fluorescence intensity of the cells, was intermediate between the mXLA Mock and WT Mock recipients. This reflects a desired amount of expression to ensure vector safety. mXLA recipients of LVXLA2 treated mHSCs had improved humoral (antibody) immune responses to T-dependent antigens vs. recipients of mXLA Mock mHSCs. B cell receptor (BCR) ligation with anti-IgM led to increased B cell proliferation in LVXLA2 recipients, averaging ~50% of live B cells proliferating (Figure 28E-F). While this did not reach the observed level of proliferation in WT Mock recipients (~88%), it was a marked increase compared to the minimal response (~2%) in mXLA Mock recipient B cells. These results demonstrate that the LVXLA2 vector is not only safe, but is also very effective. EXAMPLE 7 - SAFETY OF LVXLA2 GENE THERAPY IN HUMAN CELLS IN HUMANIZED MICE A study to evaluate the safety of LVXLA2-transduced healthy donor derived, human HSPCs in vivo was performed using a variety of assays to determine the effect on HSPC pluripotency and engraftment in humanized mice. Additionally, viral copy number (VCN) quantification, integration site mapping, and the overall health of the mice (including clinical and anatomic pathology) were assessed. Methodology Manufacturing LVXLA2-transduced PB-HSPCs for transplantation Manufacturing of LVXLA2-transduced PB-HSPCs (termed the LVXLA2 drug product, or LVXLA2 DP) was performed to closely match GMP materials and procedures that would be employed during clinical manufacturing. Human PB-HSPCs were separately obtained from two different healthy volunteers as follows. Volunteers were treated with G-CSF then subjected to peripheral blood apheresis by BloodWorks NW (Seattle, WA). The apheresis products applied to a CliniMacs column to select CD34+ cells (=PB-HSPCs). The PB-HSPCs were pre-stimulated ~24 hours in culture with SCGM and human cytokines TPO, FLT3, and SCF. Transduction enhancers (protamine sulfate and Poloxamer F127) were then added to the cell media, along with LVXLA2 (MOI=21), or without LVXLA2 as Mock transduction control DP. After LVXLA2- or mock-transduction, the HSPCs were frozen in aliquots intended for either quality control (QC) assays (~0.25 mL or other volumes), viral integration site analysis (VISA), or for transplantation into mice. After passing QA control (e.g. viability sterility, mycoplasma, endotoxin, and VCN per CFU (Figure 29A), among other indices), LVXLA2 DP and Mock DP were thawed for transplantation into mice. In vivo assessment of LVXLA2- vs Mock DPs Busulfan-conditioned immune incompetent NBSGW mice were transplanted with either LVXLA2 DP or the Mock DP in two experimental cohorts as described in Example 3. Each cohort used LVXLA2 and Mock DPs from a unique human donor. The health of the mice and the engraftment of human cells in the peripheral blood were monitored for 16- to 18- weeks, then the mice were euthanized. Post-mortem analyses assessed: the engraftment and lineage differentiation of LVXLA2-transduced vs. Mock-transduced human CD34+HSPCs by immunophenotyping of hematopoietic tissues; VCN of bone marrow and splenic leukocytes; VISA of bone marrow leukocytes; blood counts, serum chemistry, and histopathology. Vector integration site analysis (VISA) VISA was performed on gDNA extracted from a portion of the cells as previously described (Wang et al. 2014, Blood 124(6): 913-923; and Singh et al. 2017, Mol Ther Methods Clin Dev 4: 1-16), mapping the LV-genome junction sequences by alignment with the human genome. VISA analyses was performed to assess clonality and to determine: 1) the integration profile of LVXLA2 (location relative to neighboring genes, distance to transcription start site, frequency of intragenic insertions; 2) distance to reported oncogenes; and 3) clusters of integrations relative to a reference dataset. Results It was determined that VCN per positive CFU colony averaged ~2, a value within the FDA recommended upper limit of 5 (Figure 29A). The VCN of long-term engrafted human cells in the bone marrow and spleen were approximately 5 and 4, respectively (Figure 29A). There were no statistically significant differences in survival between the two treatment groups over the 16-18 week period (Figure 29B). Two mice receiving LVXLA2 DP died or were euthanized early; one was for an undetermined reason, the other was unrelated to the LVXLA2 DP therapy. Importantly, there were no health / safety impacts of the transduced LVXLA2 based on comprehensive GLP clinical pathology or histopathology assessments. There were no differences in the proportion or absolute number of human HSCs or long-term HSCs (LT-HSCs; the most primitive hematopoietic precursor cells, with self-renewing capability) that engrafted in the bone marrow (Figure 30A). Similarly, there were no differences in the proportion or absolute numbers of human CD45+ leukocytes in the bone marrow, including myeloid, T cells, and B-cells (Figure 30B). The amount of intracellular BTK per cell, measured by the mean fluorescence intensity (MFI) of BTK during immunophenotyping, was similar in bone marrow leukocytes from recipients of Mock and LVXLA2 DP. Consistent with the level of viral transduction, BTK expression was slightly increased in splenic leukocytes of LVXLA2 vs Mock DP recipients (Figure 30C). VISA of each the input LVXLA2 DPs demonstrated a highly polyclonal insertion site profile (~14,000-30,000 per sample), with none of the top 10 sequence counts accounting for more than 0.1% of the total amplicons (Figure 31). The 10 most prominent integration sites (IS) for the two DPs did not overlap with those of the other, confirming random integration of LVXLA2. VISA of BM gDNA from LVXLA2 DP recipients was analyzed in tandem with LVXLA2 XLA DP, (described in Example 8 below). Combined, LVXLA2 integrations into the healthy donor and XLA patient PB-HSPCs identified a large number of IS (~4,000-13,000 per pooled sample) that exhibited a characteristic lentiviral profile (Wu and Burgess, 2004, Cell Mol Life Sci. 61(19-20):2588-96; Demeulemeester et al, 2015, Bioessays 37(11):1202-14) with 83.8% insertion sites in gene coding regions (79.1% within intronic regions), a low integration frequency upstream of transcription start sites (3.2% of insertion sites within 10 kb) and distribution across all chromosomes. Insertion site analysis of the examined samples demonstrated a polyclonal or oligoclonal integration profile for LVXLA2 with low relative sequence counts for the 10 most prominent insertion sites (Figure 31). Common integration site (CIS) analysis revealed a diverse composition of clusters located at different chromosomes and of large dimensions, not indicating presence of IS hotspot(s). IS data revealed that 16.42% of all uniquely mappable IS were close to predefined, well characterized genes listed in the Cancer Gene Census database, with an overall low occurrence. Overall, S- EPTS / LM-PCR analysis resulted high sequencing depth and identification of large numbers of IS. LVXLA2-transduced human CD34+ cells samples showed a polyclonal IS integration profile and no dominant IS (>30 %) indicating no evidence for clonal expansion and raising no safety concerns. EXAMPLE 8 - SAFETY AND EFFICACY OF LVXLA2 GENE THERAPY IN HUMAN XLA CELLS IN HUMANIZED MICE In this study, clinical scale LVXLA2 transduction of human peripheral blood hematopoietic stem and progenitor cells (PB-HSPCs) collected from XLA patients (=LVXLA2 XLA DP) was modelled. The degree of viral marking and the genomic locations of viral integrants, and in vivo engraftment, multilineage differentiation, and safety, was then determined. The primary objective was to evaluate the safety of LVXLA2-XLA DP, relative to XLA HSPCs that were handled identically but without LV transduction (Mock XLA DP). A variety of assays were used to determine the effect of the transduction on HSPC pluripotency and engraftment, viral copy number (VCN) quantification and integration site mapping, and the overall health of the mice. The secondary objective of this study was to assess the efficacy of LVXLA2 XLA DP in overcoming the characteristic developmental block in B cell development found in XLA patients by immunophenotyping B cell subsets in the bone marrow and spleen of the recipient humanized mice. Methodology Manufacturing LVXLA2-transduced XLA PB-HSPCs for transplantation Mobilized XLA patients were apheresed at BloodWorks NW (Seattle, WA), and PB-HSPCs were affinity purified from the apheresis products as described above. Cells were cultured with or without LVXLA2 (=LVXLA2 XLA DP or Mock XLA DP), using the approximate clinical transduction scale and GMP materials that would be employed during clinical manufacturing and as described in Eaxmple 7. For cohort 2, the PB-HSPCs were frozen and stored prior to culturing for LV or mock transduction. HSPCs from healthy / unaffected donors were also handled identically but without LV for a positive control (=Mock HD DP). After LVXLA2- or mock-transduction, the HSPCs were either transplanted directly into NBSGW mice (cohort 2), or frozen in aliquots for QC assays (as described for the previous study) or for transplantation into mice (cohort 1). VISA analysis was performed on an aliquot of the transduced cells, after further expansion in liquid media to assess the spectrum of viral integration sites. In vivo comparisons of LVXLA2 XLA, Mock XLA, and Mock WT DPs For HSPCs frozen after transduction, once QA was performed and passed, cell aliquots were thawed and transplanted by injection into the retro-orbital (r.o.) venous sinus of busulfan-conditioned NBSGW mice (NOD-B6-scid-Il2rgNULLKitW41). If HSPCs were not frozen post manufacturing, the HSPCs were washed and directly transplanted into busulfan- conditioned NBSGW mice by retro-orbital (r.o.) injection into the venous sinus. The health of the mice and the engraftment of human cells in the peripheral blood were monitored regularly; the mice were euthanized 16- to 18-weeks post-transplant. Post-mortem analysis were further assessed the engraftment and B lineage differentiation of LVXLA2-transduced HSPCs compared to Mock-transduced XLA- and unaffected-HSPCs by immunophenotyping of hematopoietic tissues. VCN determination and VISA was performed on bone marrow and / or splenic hematopoietic cells. Blood counts, serum chemistry, general health, histopathology, and survival were measured as safety assessments of the gene therapy. Results The VCN of input cells was determined to be 2.5 viral copies per transduced CFU colony, below the FDA recommended upper limit of 5 (Figure 32A). The average VCN of bone marrow and splenic leukocytes extracted from LVXLA2 XLA DP engrafted recipient mice was 4.4 and 5.3, respectively (Figure 32A). There were no statistically significant differences in survival between the three treatment groups over the 17- to 18-week period (Figure 32B). Four mice in total died prior to the experimental endpoint (2 LVXLA2 XLA DP, 1 Mock XLA, and 1 Mock HD recipients) of either undetermined reasons, or not related to the vector. Importantly, there were no health / safety impacts of the transduced LVXLA2 based on comprehensive GLP clinical pathology or histopathology assessments. Engraftment of long-term hematopoietic stem cell (LT-HSCs) and human hematopoietic cells (hCD45+) in the bone marrow were similar between all three groups, suggesting no impact of the LVXLA2 transduction on the engraftment of LT-HSCs (Figure 33A). This is a desirable outcome for durable / long lasting correction of the XLA phenotype. Of note, the proportion of CD34+HSCs was lower for the LVXLA2 vs the Mock XLA groups, reaching levels similar to that of Mock HDs. The increase in CD34+cells in untreated XLA subjects reflects the block in early B cell development with an increased proportion of B lineage progenitors expressing CD34. Restoration of HSCs to the normal range demonstrates rescue of this defect following LVXLA2 treatment. Bone marrow myeloid and T cells were present in similar numbers between the three treatment groups. However, there were fewer bone marrow B cells in recipients of the Mock XLA DP vs. the LVXLA2 XLA DP treatment group (Figure 33A), consistent with improved B cell hematopoiesis post-LVXLA2 treatment. Increased numbers of B cells for LVXLA2 XLA DP vs. Mock XLA DP recipients were also observed in the spleen (Figure 33B). Splenic engraftment of human CD45+hematopoietic cells, myeloid, and T cells were statistically similar between the LVXLA2 XLA and Mock XLA DP treatment groups (Figure 33B). Immunophenotyping to define developmental B cells subsets within the bone marrow revealed that Mock XLA recipient mice exhibited the block in B cell development at the Pro-B cell stage that is a characteristic of XLA patients (Figure 34A). This Pro- to Pre-B cell developmental block was not present in the LVXLA2 XLA DP recipients, consistent with correction of the BTK signaling defect in XLA. Note the lower numbers of Pro-B cells, along with the higher numbers of Pre-B cells and immature B cells in this group vs. the Mock XLA group. B cell development in the spleen was also improved with LVXLA2 transduction of the XLA patient cells. Figure 34B shows the number and proportion of mature B cells in the spleens of mice receiving LVXLA2 XLA DP were substantially greater than recipients of the Mock XLA DP. The results of the VISA analysis for input cells and BM cells at the experimental endpoint is described above in Example 7. Overall, this study clearly demonstrates the desirable safety profile and therapeutic efficacy of LVXLA2. EXAMPLE 9 - DETERMINING CHIMERISM OF BTK+MURINE HSCS REQUIRED FOR OPTIMAL B CELL HEMATOPOIESIS AND FUNCTION XLA is a promising candidate disease for a gene replacement therapy in part because gene-corrected BTK+B cells are predicted to exhibit a selective advantage during early B cell development. Thus, partial correction of the predecessor stem cells will likely be sufficient to cure the disease phenotypes. This concept is supported by previous studies of non-random X-inactivation in female carrier mice and humans, and by using limiting murine hematopoietic stem cell (mHSC) transfer experiments in BTK mutant XID mice (an imperfect model of XLA). The primary objective of this study was to estimate the proportion of WT murine hematopoietic stem cells needed to reconstitute B cell numbers and antibody responses in mXLA mice, testing a range of doses of BTK-sufficient bone marrow (WT BM; containing mHSCs) in a mixture with mXLA BM, in a chimeric transplant experiment. Methodology Bone marrow cells obtained from WT or mXLA mice (donors) were mixed in 6 different proportions (Table 6), and then transplanted into lethally-irradiated mXLA mice recipients. To allow identification of WT vs. mXLA derived cells during immunophenotyping, mXLA recipients had the same congenic CD45 marker as the mXLA donor, but a different congenic CD45 marker than WT donors. Experimental controls were transplanted with WT BM cells only (positive control) or mXLA BM cells only (negative control). Table 6. Percentages of WT and mXLA bone marrow in the experimental groups *CD45.2; †CD45.1 During the in-life phase of the study, peripheral blood samples were obtained ~10 weeks post-transplant to monitor engraftment and hematopoiesis. The ability of B cells to respond to T-dependent antigens was assessed by immune challenges with NP-CGG 12- and 17-weeks post-transplant (primary and secondary immunizations, respectively), followed 1-week post each immunization by collection of blood samples for serum ELISAs. Mice were sacrificed 20-weeks post-transplant, and spleen and bone marrow cells were collected and immunophenotyped by flow cytometry to detect immune cell subsets and the congenic markers. Results Secretion of class-switched IgG antibodies specific for the T-dependent antigen was evident in mice receiving as little as 1 to 5% of WT bone marrow. Mice receiving 10-20% of WT bone marrow had similar serum levels of the specific antibody as those receiving only WT cells (Figure 35A). These results indicate functional restoration of B cell immunity required a chimerism of ≤ ~10% WT BM. The proportion of BTK+splenic B cells increased almost linearly with the WT chimerism of the graft, until the WT chimerism reached about 20% (Figure 35B). At this point (20% WT chimerism) approximately 80% of the splenic B cells are BTK+. The % BTK+B cells does not appreciably increase at higher WT chimerism. Note that chimerism is based on the % of WT BM neutrophils. Neutrophil chimerism is a standard clinical marker of HSC engraftment due to their short half-life. These results indicate that restoration a chimerism of ≤ ~20% WT BM is sufficient for nearly complete rescue of BTK+B cell populations within the peripheral immune system. The disclosure of every patent, patent application, and publication cited herein is hereby incorporated herein by reference in its entirety. The citation of any reference herein should not be construed as an admission that such reference is available as “Prior Art” to the instant application. Throughout the specification the aim has been to describe the preferred embodiments of the invention without limiting the invention to any one embodiment or specific collection of features. Those of skill in the art will therefore appreciate that, in light of the instant disclosure, various modifications and changes can be made in the particular embodiments exemplified without departing from the scope of the present invention. All such modifications and changes are intended to be included within the scope of the appended claims. Sequences disclosed herein Unmodified 0.7 UCOE Fwd (SEQ ID NO:1) ccgcaaacacccgaatcaacttctagtcaaattattgttcacgccgcaatgacccacccctggcccgcgtctgtggaactgac ccctggtgtacaggagagttcgctgctgaaagtggtcccaaaggggtactagtttttaagctcccaactccccctcccccagc gtctggaggattccacaccctcgcaccgcaggggcgaggaagtgggcggagtccggttttggcgccagccgctgaggctgcca agcagaaaagccaccgctgaggagactccggtcactgtcctcgccccgcctcccccttccctccccttggggaccaccgggcg ccacgccgcgaacggtaagtgccgcggtcgtcggcgcctccgccctccccctagggccccaattcccagcgggcgcggcgcgc ggcccctccccccgccgggcgcgcgcccgctgccccgcccttcgtggccgcccggcgtgggcggtgccacccctccccccagc ggccccgcgcgcagctcccggctccctcccccttcggatgtggcttgagctgtaggcgcggagggccggagacgctgcagacc cgcgacccggagcagctcggaggcggtgaagtcggtggctttccttctctctagctctcgctcgctggtggtgcttcagatgc cacacgcgg Modified 0.7 UCOE Fwd (SEQ ID NO:2) Ccggaaacacccgaatcaacttctagtcaaattattgttcacgccgcaatgacccacccctggcccgcgtctgtggaactgac ccctggtgtacaggagagttcgctgctgaaagtggtcccaaaggggtactagtttttaagctcccaactccccctcccccagc gtctggaggattccacaccctcgcaccgcaggggcgaggaagtgggcggagtccggttttggcgccagccgctgaggctgcca agcagaaaagccaccgctgaggagactccggtcactgtcctcgccccgcctcccccttccctccccttggggaccaccgggcg ccacgccgcgaacgcgaactgccgcggtccgcgccgcctccgccctcccccttgggccccaattcccagcgggcgcggcgcgc ggcccctccccccgccgggcgcgcgcccgctgccccgcccttcgtggccgcccggcgtgggcggtgccacccctccccccggc ggccccgcgcgcagctcccggctccctcccccttcggatgtggcttgagctgtaggcgcggagggccggagacgctgcagacc cgcgacccggagcagctcggaggcggtgaacgcgctggctttccttctctctagctctcgctcgctggtggtgcttcagatgc cacacgcgt BTK polypeptide sequence (NCBI Ref: NC_000023.11) (SEQ ID NO:3) MAAVILESIFLKRSQQKKKTSPLNFKKRLFLLTVHKLSYYEYDFERGRRGSKKGSIDVEKITCVETVVPEKNPPPERQIPRRG EESSEMEQISIIERFPYPFQVVYDEGPLYVFSPTEELRKRWIHQLKNVIRYNSDLVQKYHPCFWIDGQYLCCSQTAKNAMGCQ ILENRNGSLKPGSSHRKTKKPLPPTPEEDQILKKPLPPEPAAAPVSTSELKKVVALYDYMPMNANDLQLRKGDEYFILEESNL PWWRARDKNGQEGYIPSNYVTEAEDSIEMYEWYSKHMTRSQAEQLLKQEGKEGGFIVRDSSKAGKYTVSVFAKSTGDPQGVIR HYVVCSTPQSQYYLAEKHLFSTIPELINYHQHNSAGLISRLKYPVSQQNKNAPSTAGLGYGSWEIDPKDLTFLKELGTGQFGV VKYGKWRGQYDVAIKMIKEGSMSEDEFIEEAKVMMNLSHEKLVQLYGVCTKQRPIFIITEYMANGCLLNYLREMRHRFQTQQL LEMCKDVCEAMEYLESKQFLHRDLAARNCLVNDQGVVKVSDFGLSRYVLDDEYTSSVGSKFPVRWSPPEVLMYSKFSSKSDIW AFGVLMWEIYSLGKMPYERFTNSETAEHIAQGLRLYRPHLASEKVYTIMYSCWHEKADERPTFKILLSNILDVMDEES Codon optimized BTK nucleotide sequence (coBTK) (SEQ ID NO:4) atggccgctgtgatcctggagagcattttcctgaagaggtcccagcagaaaaagaaaacctctcccctgaactttaagaaaag actgttcctgctgacagtgcacaagctgtcttactatgagtacgactttgagcggggccgccgaggatcaaaaaaggggagca tcgatgtggagaagattacatgcgtggagaccgtggtccctgaaaagaatccaccccctgagaggcagatcccaagacggggc gaggagtcctctgagatggagcagattagtatcattgagcgcttcccctatccttttcaggtggtgtacgacgagggaccact gtatgtgttctcacccacagaggagctgagaaagaggtggattcaccagctgaagaacgtgattagatacaatagcgatctgg tgcagaagtatcacccttgtttttggatcgacgggcagtacctgtgctgttcccagacagctaagaacgctatgggatgccag attctggaaaatcggaacggatctctgaaaccagggagttcacaccgcaagaccaaaaagcccctgcctccaacacccgagga ggatcagatcctgaaaaagcctctgccacccgagcctgctgcagccccagtcagcacttccgaactgaaaaaggtggtggctc tgtatgactacatgcccatgaatgctaacgatctgcagctgagaaagggcgacgagtatttcattctggaagagtctaatctg ccttggtggagggccagagataagaacggacaggaggggtacatcccatctaattatgtgaccgaggctgaggactctattga gatgtacgagtggtatagcaagcacatgacacggtcccaggctgagcagctgctgaagcaggagggcaaagagggagggttta tcgtgcgcgattctagtaaggccggcaaatacactgtgtcagtgttcgctaagagcaccggagacccccagggcgtgatcaga cactatgtggtgtgttccacacctcagtctcagtactatctggctgagaagcacctgtttagtacaatcccagagctgattaa ctaccaccagcacaattctgccggcctgatcagcaggctgaagtatcccgtctcccagcagaacaaaaatgctccttctaccg ctggactggggtacggcagttgggagattgatccaaaggacctgacattcctgaaggagctgggaactgggcagtttggcgtg gtgaagtatggaaaatggagagggcagtacgatgtggccatcaagatgatcaaggagggctcaatgagcgaggacgagttcat cgaggaggctaaggtcatgatgaacctgtcccacgagaaactggtgcagctgtatggagtgtgcaccaagcagcggcccattt ttatcattacagagtacatggctaatgggtgtctgctgaactatctgcgcgagatgagacacagattccagacacagcagctg ctggaaatgtgcaaggatgtgtgtgaggctatggagtacctggagtctaagcagtttctgcaccgggacctggctgctcgcaa ttgcctggtgaacgatcagggcgtggtgaaggtgagtgacttcggactgtcaaggtatgtgctggatgacgagtacaccagct ccgtgggctctaagtttcctgtgagatggtctccacccgaggtgctgatgtatagcaagttctcctctaagagcgatatctgg gcctttggcgtgctgatgtgggaaatctacagcctgggcaagatgccttacgagcggttcacaaattccgagacagctgagca catcgcccagggcctgcgcctgtaccggccacatctggcctctgagaaggtgtacaccatcatgtacagctgttggcacgaga aggccgacgagagacccacattcaagatcctgctgtccaacattctagatgtgatggacgaggagagctga Codon optimized BTK nucleotide sequence (coBTK) (SEQ ID NO:5) atggccgccgtgatcctggaaagcatcttcctgaagcggagccagcagaagaagaaaaccagccccctgaacttcaagaagcg gctgttcctgctgaccgtgcacaagctgtcctactacgagtacgacttcgagcggggcagacggggcagcaagaagggcagca tcgacgtcgagaagatcacctgcgtggagaccgtggtgcccgagaagaacccccctcccgagcggcagatccccagacggggc gaggaaagcagcgagatggaacagatcagcatcatcgagcggttcccttacccattccaagtggtgtacgacgagggccccct gtacgtgttcagccccaccgaggaactgcggaagcggtggattcaccagctgaagaacgtgatccggtacaacagcgacctgg tgcagaagtaccacccctgcttttggatcgacggccagtacctgtgctgcagccagaccgccaagaacgctatgggctgccag attctggaaaaccggaacggcagcctgaagcccggcagcagccacagaaagaccaagaagcccctgccccccacccccgaaga ggaccagatcctgaagaagcctctgcctcccgagcccgccgctgcacctgtgagcaccagcgagctgaagaaagtggtggccc tgtacgactacatgcccatgaacgccaacgacctgcagctgcggaagggcgacgagtacttcatcctggaagaaagcaacctg ccctggtggcgggccagggacaagaacggccaggaaggctacatccccagcaactacgtgaccgaggccgaggactccatcga gatgtacgagtggtacagcaagcacatgaccagaagccaggccgaacagctgctgaagcaggaaggcaaagagggcggcttca tcgtccgggacagcagcaaggccggcaagtacaccgtgagcgtgttcgccaagagcaccggcgacccccagggcgtgatccgg cactacgtggtgtgcagcaccccccagagccagtactacctggccgagaagcacctgttcagcaccatccccgagctgatcaa ctatcaccagcacaacagcgctggactgatttctcggctgaagtaccccgtgtcccagcagaacaaaaacgcccccagcacag ccggcctgggctacggcagctgggagatcgaccccaaggacctgaccttcctgaaagagctgggcaccggccagttcggcgtg gtgaagtacggcaagtggaggggccagtacgacgtggccatcaagatgatcaaggaaggcagcatgagcgaggacgagttcat cgaggaagccaaagtgatgatgaacctgagccacgagaagctggtgcagctgtacggcgtgtgcaccaagcagcggcccatct tcatcatcaccgagtacatggccaacggctgcctgctgaactacctgcgggagatgcggcacaggttccagacacagcagctg ctcgaaatgtgcaaggacgtgtgcgaggctatggaatacctggaatccaagcagttcctgcaccgggacctggccgccagaaa ctgcctggtgaacgaccagggggtggtgaaggtgtccgacttcggcctgagcagatacgtgctggacgacgagtacaccagca gcgtgggcagcaagttccccgtgcggtggagcccccctgaggtgctgatgtacagcaagttcagcagcaagagcgacatctgg gccttcggcgtgctgatgtgggagatctacagcctgggcaagatgccctacgagcggttcaccaacagcgagaccgccgagca catcgcccagggcctgcggctgtacaggccccacctggccagcgagaaggtgtacaccatcatgtacagctgctggcacgaga aggccgacgagaggcccaccttcaagatcctgctgtccaacatcctggacgtgatggacgaggaaagctga BTK promoter (BTKp) (SEQ ID NO:6) tgcatttcctaggagaatccctgggggaatcattgcagttggagcataatgtagggggcccctgagaaaacctccaggcttca agtgacatacctagtctgctttaccggtttacaggactcaagagaaaggtggacattgagagttaatccctgaggccaaatct taaatggagaaagtcaacatccacagaaaatggggaagggcacaagtatttctgtgggcttatattccgacatttttatctgt aggggaaaaatgctttcttagaaaatgactcagcacggggaagtcttgtctctacctctgtcttgttttgtcctttggggtcc cttcactatcaagttcaactgtgtgtccctgagactcctctgccccggaggacaggagactcgaaaaacgctcttcctggcca gtctctttgctctgtgtctgccagcccccagcatctctcctctttcctgtaagcccctctccctgtgctgactgtcttcatag tactttaggtatgttgtccctttacctctgggaggatagcttgatgacctgtctgctcaggccagccccatctagagtctcag tggccccagtcatgttgagaaaggttctttcaaagatagactcaagatagtagtgtcagaggtcccaagcaaatgaagggcgg ggacagttgagggggtggaatagggacggcagcagggaaccagatagcatgctgctgagaagaaaaaaagacattggtttagg tcaggaagcaaaaaaagggaactgagtggctgtgaaagggtggggtttgctcagactgtccttcctctctggactgtaagaat tagtctc BTKp.coBTK (SEQ ID NO:7) tgcatttcctaggagaatccctgggggaatcattgcagttggagcataatgtagggggcccctgagaaaacctccaggcttca agtgacatacctagtctgctttaccggtttacaggactcaagagaaaggtggacattgagagttaatccctgaggccaaatct taaatggagaaagtcaacatccacagaaaatggggaagggcacaagtatttctgtgggcttatattccgacatttttatctgt aggggaaaaatgctttcttagaaaatgactcagcacggggaagtcttgtctctacctctgtcttgttttgtcctttggggtcc cttcactatcaagttcaactgtgtgtccctgagactcctctgccccggaggacaggagactcgaaaaacgctcttcctggcca gtctctttgctctgtgtctgccagcccccagcatctctcctctttcctgtaagcccctctccctgtgctgactgtcttcatag tactttaggtatgttgtccctttacctctgggaggatagcttgatgacctgtctgctcaggccagccccatctagagtctcag tggccccagtcatgttgagaaaggttctttcaaagatagactcaagatagtagtgtcagaggtcccaagcaaatgaagggcgg ggacagttgagggggtggaatagggacggcagcagggaaccagatagcatgctgctgagaagaaaaaaagacattggtttagg tcaggaagcaaaaaaagggaactgagtggctgtgaaagggtggggtttgctcagactgtccttcctctctggactgtaagaat tagtctcgagaaagaagccaccatggccgctgtgatcctggagagcattttcctgaagaggtcccagcagaaaaagaaaacct ctcccctgaactttaagaaaagactgttcctgctgacagtgcacaagctgtcttactatgagtacgactttgagcggggccgc cgaggatcaaaaaaggggagcatcgatgtggagaagattacatgcgtggagaccgtggtccctgaaaagaatccaccccctga gaggcagatcccaagacggggcgaggagtcctctgagatggagcagattagtatcattgagcgcttcccctatccttttcagg tggtgtacgacgagggaccactgtatgtgttctcacccacagaggagctgagaaagaggtggattcaccagctgaagaacgtg attagatacaatagcgatctggtgcagaagtatcacccttgtttttggatcgacgggcagtacctgtgctgttcccagacagc taagaacgctatgggatgccagattctggaaaatcggaacggatctctgaaaccagggagttcacaccgcaagaccaaaaagc ccctgcctccaacacccgaggaggatcagatcctgaaaaagcctctgccacccgagcctgctgcagccccagtcagcacttcc gaactgaaaaaggtggtggctctgtatgactacatgcccatgaatgctaacgatctgcagctgagaaagggcgacgagtattt cattctggaagagtctaatctgccttggtggagggccagagataagaacggacaggaggggtacatcccatctaattatgtga ccgaggctgaggactctattgagatgtacgagtggtatagcaagcacatgacacggtcccaggctgagcagctgctgaagcag gagggcaaagagggagggtttatcgtgcgcgattctagtaaggccggcaaatacactgtgtcagtgttcgctaagagcaccgg agacccccagggcgtgatcagacactatgtggtgtgttccacacctcagtctcagtactatctggctgagaagcacctgttta gtacaatcccagagctgattaactaccaccagcacaattctgccggcctgatcagcaggctgaagtatcccgtctcccagcag aacaaaaatgctccttctaccgctggactggggtacggcagttgggagattgatccaaaggacctgacattcctgaaggagct gggaactgggcagtttggcgtggtgaagtatggaaaatggagagggcagtacgatgtggccatcaagatgatcaaggagggct caatgagcgaggacgagttcatcgaggaggctaaggtcatgatgaacctgtcccacgagaaactggtgcagctgtatggagtg tgcaccaagcagcggcccatttttatcattacagagtacatggctaatgggtgtctgctgaactatctgcgcgagatgagaca cagattccagacacagcagctgctggaaatgtgcaaggatgtgtgtgaggctatggagtacctggagtctaagcagtttctgc accgggacctggctgctcgcaattgcctggtgaacgatcagggcgtggtgaaggtgagtgacttcggactgtcaaggtatgtg ctggatgacgagtacaccagctccgtgggctctaagtttcctgtgagatggtctccacccgaggtgctgatgtatagcaagtt ctcctctaagagcgatatctgggcctttggcgtgctgatgtgggaaatctacagcctgggcaagatgccttacgagcggttca caaattccgagacagctgagcacatcgcccagggcctgcgcctgtaccggccacatctggcctctgagaaggtgtacaccatc atgtacagctgttggcacgagaaggccgacgagagacccacattcaagatcctgctgtccaacattctagatgtgatggacga ggagagctga 0.7UCOE.BTKp.coBTK (SEQ ID NO:8) ccggaaacacccgaatcaacttctagtcaaattattgttcacgccgcaatgacccacccctggcccgcgtctgtggaactgac ccctggtgtacaggagagttcgctgctgaaagtggtcccaaaggggtactagtttttaagctcccaactccccctcccccagc gtctggaggattccacaccctcgcaccgcaggggcgaggaagtgggcggagtccggttttggcgccagccgctgaggctgcca agcagaaaagccaccgctgaggagactccggtcactgtcctcgccccgcctcccccttccctccccttggggaccaccgggcg ccacgccgcgaacgcgaactgccgcggtccgcgccgcctccgccctcccccttgggccccaattcccagcgggcgcggcgcgc ggcccctccccccgccgggcgcgcgcccgctgccccgcccttcgtggccgcccggcgtgggcggtgccacccctccccccggc ggccccgcgcgcagctcccggctccctcccccttcggatgtggcttgagctgtaggcgcggagggccggagacgctgcagacc cgcgacccggagcagctcggaggcggtgaacgcgctggctttccttctctctagctctcgctcgctggtggtgcttcagatgc cacacgcgtccggggctagctacgacgcgttccggaattcgcccttgcatttcctaggagaatccctgggggaatcattgcag ttggagcataatgtagggggcccctgagaaaacctccaggcttcaagtgacatacctagtctgctttaccggtttacaggact caagagaaaggtggacattgagagttaatccctgaggccaaatcttaaatggagaaagtcaacatccacagaaaatggggaag ggcacaagtatttctgtgggcttatattccgacatttttatctgtaggggaaaaatgctttcttagaaaatgactcagcacgg ggaagtcttgtctctacctctgtcttgttttgtcctttggggtcccttcactatcaagttcaactgtgtgtccctgagactcc tctgccccggaggacaggagactcgaaaaacgctcttcctggccagtctctttgctctgtgtctgccagcccccagcatctct cctctttcctgtaagcccctctccctgtgctgactgtcttcatagtactttaggtatgttgtccctttacctctgggaggata gcttgatgacctgtctgctcaggccagccccatctagagtctcagtggccccagtcatgttgagaaaggttctttcaaagata gactcaagatagtagtgtcagaggtcccaagcaaatgaagggcggggacagttgagggggtggaatagggacggcagcaggga accagatagcatgctgctgagaagaaaaaaagacattggtttaggtcaggaagcaaaaaaagggaactgagtggctgtgaaag ggtggggtttgctcagactgtccttcctctctggactgtaagaattagtctcgagaaagaagccaccatggccgctgtgatcc tggagagcattttcctgaagaggtcccagcagaaaaagaaaacctctcccctgaactttaagaaaagactgttcctgctgaca gtgcacaagctgtcttactatgagtacgactttgagcggggccgccgaggatcaaaaaaggggagcatcgatgtggagaagat tacatgcgtggagaccgtggtccctgaaaagaatccaccccctgagaggcagatcccaagacggggcgaggagtcctctgaga tggagcagattagtatcattgagcgcttcccctatccttttcaggtggtgtacgacgagggaccactgtatgtgttctcaccc acagaggagctgagaaagaggtggattcaccagctgaagaacgtgattagatacaatagcgatctggtgcagaagtatcaccc ttgtttttggatcgacgggcagtacctgtgctgttcccagacagctaagaacgctatgggatgccagattctggaaaatcgga acggatctctgaaaccagggagttcacaccgcaagaccaaaaagcccctgcctccaacacccgaggaggatcagatcctgaaa aagcctctgccacccgagcctgctgcagccccagtcagcacttccgaactgaaaaaggtggtggctctgtatgactacatgcc catgaatgctaacgatctgcagctgagaaagggcgacgagtatttcattctggaagagtctaatctgccttggtggagggcca gagataagaacggacaggaggggtacatcccatctaattatgtgaccgaggctgaggactctattgagatgtacgagtggtat agcaagcacatgacacggtcccaggctgagcagctgctgaagcaggagggcaaagagggagggtttatcgtgcgcgattctag taaggccggcaaatacactgtgtcagtgttcgctaagagcaccggagacccccagggcgtgatcagacactatgtggtgtgtt ccacacctcagtctcagtactatctggctgagaagcacctgtttagtacaatcccagagctgattaactaccaccagcacaat tctgccggcctgatcagcaggctgaagtatcccgtctcccagcagaacaaaaatgctccttctaccgctggactggggtacgg cagttgggagattgatccaaaggacctgacattcctgaaggagctgggaactgggcagtttggcgtggtgaagtatggaaaat ggagagggcagtacgatgtggccatcaagatgatcaaggagggctcaatgagcgaggacgagttcatcgaggaggctaaggtc atgatgaacctgtcccacgagaaactggtgcagctgtatggagtgtgcaccaagcagcggcccatttttatcattacagagta catggctaatgggtgtctgctgaactatctgcgcgagatgagacacagattccagacacagcagctgctggaaatgtgcaagg atgtgtgtgaggctatggagtacctggagtctaagcagtttctgcaccgggacctggctgctcgcaattgcctggtgaacgat cagggcgtggtgaaggtgagtgacttcggactgtcaaggtatgtgctggatgacgagtacaccagctccgtgggctctaagtt tcctgtgagatggtctccacccgaggtgctgatgtatagcaagttctcctctaagagcgatatctgggcctttggcgtgctga tgtgggaaatctacagcctgggcaagatgccttacgagcggttcacaaattccgagacagctgagcacatcgcccagggcctg cgcctgtaccggccacatctggcctctgagaaggtgtacaccatcatgtacagctgttggcacgagaaggccgacgagagacc cacattcaagatcctgctgtccaacattctagatgtgatggacgaggagagctga wild-type WPRE (SEQ ID NO:9) atcaacctctggattacaaaatttgtgaaagattgactgatattcttaactatgttgctccttttacgctgtgtggatatgct gctttaatgcctctgtatcatgctattgcttcccgtacggctttcgttttctcctccttgtataaatcctggttgctgtctct ttatgaggagttgtggcccgttgtccgtcaacgtggcgtggtgtgctctgtgtttgctgacgcaacccccactggctggggca ttgccaccacctgtcaactcctttctgggactttcgctttccccctcccgatcgccacggcagaactcatcgccgcctgcctt gcccgctgctggacaggggctaggttgctgggcactgataattccgtggtgttgtcggggaagctgacgtcctttccatggct gctcgcctgtgttgccaactggatcctgcgcgggacgtccttctgctacgtcccttcggctctcaatccagcggacctccctt cccgaggccttctgccggttctgcggcctctcccgcgtcttcgctttcggcctccgacgagtcggatctccctttgggccgcc tccccgcctg cPPT (SEQ ID NO:12) aattccctacaatccccaaagtcaaggagtagtagaatctatgaataaagaattaaagaaaattataggacaggtaagagatc aggctgaacatcttaagacagcagtacaaatggcagtattcatccacaattttaaaagaaaaggggggattggggggtacagt gcaggggaaagaatagtagacataatagcaacagacatacaaactaaagaattacaaaaacaaattacaaaaattcaaaattt tcgggtttattacagggacagcagaaatccactttggaaaggaccagcaaagctcctctggaaaggtgaaggggcagtagtaa tacaagataatagtgacataaaagtagtgccaagaagaaaagcaaagatcattagggattatggaaaacagatggcaggtgat gattgtgtggcaagtagacaggatgaggattagaacatggaaaagtttagtaaaacaccata ^U3 (SEQ ID NO:13) ctggaagggctaattcactcccaaagaagacaagatagatctgctttttgcctgtactgggtctctctggttagaccagatct gagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagctt RU5 (SEQ ID NO:14) ggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttg ccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtg gaaaatctctagcag Gag (SEQ ID NO:15) atgggtgcgagagcgtcagtattaagcgggggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaagaaa aaatataaattaaaacatatagtatgggcaagcagggagctagaacgattcgcagttaatcctggcctgttagaaacatcaga aggctgtagacaaatactgggacagctacaaccatcccttcagacaggatcagaagaacttagatcattatataatacagtag caaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttagacaagatagaggaagagcaaaacaaa agtaagaccaccgcacagcaagcggccgctgat 3' LTR (^U3) (SEQ ID NO:16) tggaagggctaattcactcccaacgaagacaagatctgctttttgcttgtactgggtctctctggttagaccagatctgagcc tgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccg tctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaatctctagca rabbit ^-globin polyA (SEQ ID NO:17) gatctttttccctctgccaaaaattatggggacatcatgaagccccttgagcatctgacttctggctaataaaggaaatttat tttcattgcaatagtgtgttggaattttttgtgtctctcactcggaaggacatatgggagggcaaatcatttaaaacatcaga atgagtatttggtttagagtttggcaacatatgcccatatgctggctgccatgaacaaaggttggctataaagaggtcatcag tatatgaaacagccccctgctgtccattccttattccatagaaaagccttgacttgaggttagattttttttatattttgttt tgtgttatttttttctttaacatccctaaaattttccttacatgttttactagccagatttttcctcctctcctgactactcc cagtcatagctgtccctcttctcttatggagatc SV40 ori / polyA (SEQ ID NO:18) tggctctagctatcccgcccctaactccgcccagttccgcccattctccgccccatggctgactaattttttttatttatgca gaggccgaggccgcctcggcctctgagctattccagaagtagtgaggaggcttttttggaggcctaggcttttgcgtcgagac gtacccaattcgccctatagtgagtcgtattacgcgcgctc SV40 ori Vector pRRL-XLA LV (SEQ ID NO:20) tcgcgcgtttcggtgatgacggtgaaaacctctgacacatgcagctcccggagacggtcacagcttgtctgtaagcggatgcc gggagcagacaagcccgtcagggcgcgtcagcgggtgttggcgggtgtcggggctggcttaactatgcggcatcagagcagat tgtactgagagtgcaccatatgcggtgtgaaataccgcacagatgcgtaaggagaaaataccgcatcaggcgccattcgccat tcaggctgcgcaactgttgggaagggcgatcggtgcgggcctcttcgctattacgccagctggcgaaagggggatgtgctgca aggcgattaagttgggtaacgccagggttttcccagtcacgacgttgtaaaacgacggccagagaattcgagctcggtacctc gcgaatgcatctagatttaatgtagtcttatgcaatactcttgtagtcttgcaacatggtaacgatgagttagcaacatgcct tacaaggagagaaaaagcaccgtgcatgccgattggtggaagtaaggtggtacgatcgtgccttattaggaaggcaacagacg ggtctgacatggattggacgaaccactgaattgccgcattgcagagatattgtatttaagtgcctagctcgatacataaacgg gtctctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgc cttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtgg aaaatctctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccagaggagctctctcgacgcaggactcggctt gctgaagcgcgcacggcaagaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcggaggctagaaggagag agatgggtgcgagagcgtcagtattaagcgggggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaaga aaaaatataaattaaaacatatagtatgggcaagcagggagctagaacgattcgcagttaatcctggcctgttagaaacatca gaaggctgtagacaaatactgggacagctacaaccatcccttcagacaggatcagaagaacttagatcattatataatacagt agcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttagacaagatagaggaagagcaaaaca aaagtaagaccaccgcacagcaagcggccgctgatcttcagacctggaggaggagatatgagggacaattggagaagtgaatt atataaatataaagtagtaaaaattgaaccattaggagtagcacccaccaaggcaaagagaagagtggtgcagagagaaaaaa gagcagtgggaataggagctttgttccttgggttcttgggagcagcaggaagcactatgggcgcagcgtcaatgacgctgacg gtacaggccagacaattattgtctggtatagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagcatctgtt gcaactcacagtctggggcatcaagcagctccaggcaagaatcctggctgtggaaagatacctaaaggatcaacagctcctgg ggatttggggttgctctggaaaactcatttgcaccactgctgtgccttggaatgctagttggagtaataaatctctggaacag atttggaatcacacgacctggatggagtgggacagagaaattaacaattacacaagcttaatacactccttaattgaagaatc gcaaaaccagcaagaaaagaatgaacaagaattattggaattagataaatgggcaagtttgtggaattggtttaacataacaa attggctgtggtatataaaattattcataatgatagtaggaggcttggtaggtttaagaatagtttttgctgtactttctata gtgaatagagttaggcagggatattcaccattatcgtttcagacccacctcccaaccccgaggggacccgacaggcccgaagg aatagaagaagaaggtggagagagagacagagacagatccattcgattagtgaacggatctcgacggtatcggttaactttta aaagaaaaggggggattggggggtacagtgcaggggaaagaatagtagacataatagcaacagacatacaaactaaagaatta caaaaacaaattacaaaaattcaaaattttatcgatcacgagactagcctcgaaattcgagctagtcccggccggaaacaccc gaatcaacttctagtcaaattattgttcacgccgcaatgacccacccctggcccgcgtctgtggaactgacccctggtgtaca ggagagttcgctgctgaaagtggtcccaaaggggtactagtttttaagctcccaactccccctcccccagcgtctggaggatt ccacaccctcgcaccgcaggggcgaggaagtgggcggagtccggttttggcgccagccgctgaggctgccaagcagaaaagcc accgctgaggagactccggtcactgtcctcgccccgcctcccccttccctccccttggggaccaccgggcgccacgccgcgaa cgcgaactgccgcggtccgcgccgcctccgccctcccccttgggccccaattcccagcgggcgcggcgcgcggcccctccccc cgccgggcgcgcgcccgctgccccgcccttcgtggccgcccggcgtgggcggtgccacccctccccccggcggccccgcgcgc agctcccggctccctcccccttcggatgtggcttgagctgtaggcgcggagggccggagacgctgcagacccgcgacccggag cagctcggaggcggtgaacgcgctggctttccttctctctagctctcgctcgctggtggtgcttcagatgccacacgcgtccg gggctagctacgacgcgttccggaattcgcccttgcatttcctaggagaatccctgggggaatcattgcagttggagcataat gtagggggcccctgagaaaacctccaggcttcaagtgacatacctagtctgctttaccggtttacaggactcaagagaaaggt ggacattgagagttaatccctgaggccaaatcttaaatggagaaagtcaacatccacagaaaatggggaagggcacaagtatt tctgtgggcttatattccgacatttttatctgtaggggaaaaatgctttcttagaaaatgactcagcacggggaagtcttgtc tctacctctgtcttgttttgtcctttggggtcccttcactatcaagttcaactgtgtgtccctgagactcctctgccccggag gacaggagactcgaaaaacgctcttcctggccagtctctttgctctgtgtctgccagcccccagcatctctcctctttcctgt aagcccctctccctgtgctgactgtcttcatagtactttaggtatgttgtccctttacctctgggaggatagcttgatgacct gtctgctcaggccagccccatctagagtctcagtggccccagtcatgttgagaaaggttctttcaaagatagactcaagatag tagtgtcagaggtcccaagcaaatgaagggcggggacagttgagggggtggaatagggacggcagcagggaaccagatagcat gctgctgagaagaaaaaaagacattggtttaggtcaggaagcaaaaaaagggaactgagtggctgtgaaagggtggggtttgc tcagactgtccttcctctctggactgtaagaattagtctcgagaaagaagccaccatggccgctgtgatcctggagagcattt tcctgaagaggtcccagcagaaaaagaaaacctctcccctgaactttaagaaaagactgttcctgctgacagtgcacaagctg tcttactatgagtacgactttgagcggggccgccgaggatcaaaaaaggggagcatcgatgtggagaagattacatgcgtgga gaccgtggtccctgaaaagaatccaccccctgagaggcagatcccaagacggggcgaggagtcctctgagatggagcagatta gtatcattgagcgcttcccctatccttttcaggtggtgtacgacgagggaccactgtatgtgttctcacccacagaggagctg agaaagaggtggattcaccagctgaagaacgtgattagatacaatagcgatctggtgcagaagtatcacccttgtttttggat cgacgggcagtacctgtgctgttcccagacagctaagaacgctatgggatgccagattctggaaaatcggaacggatctctga aaccagggagttcacaccgcaagaccaaaaagcccctgcctccaacacccgaggaggatcagatcctgaaaaagcctctgcca cccgagcctgctgcagccccagtcagcacttccgaactgaaaaaggtggtggctctgtatgactacatgcccatgaatgctaa cgatctgcagctgagaaagggcgacgagtatttcattctggaagagtctaatctgccttggtggagggccagagataagaacg gacaggaggggtacatcccatctaattatgtgaccgaggctgaggactctattgagatgtacgagtggtatagcaagcacatg acacggtcccaggctgagcagctgctgaagcaggagggcaaagagggagggtttatcgtgcgcgattctagtaaggccggcaa atacactgtgtcagtgttcgctaagagcaccggagacccccagggcgtgatcagacactatgtggtgtgttccacacctcagt ctcagtactatctggctgagaagcacctgtttagtacaatcccagagctgattaactaccaccagcacaattctgccggcctg atcagcaggctgaagtatcccgtctcccagcagaacaaaaatgctccttctaccgctggactggggtacggcagttgggagat tgatccaaaggacctgacattcctgaaggagctgggaactgggcagtttggcgtggtgaagtatggaaaatggagagggcagt acgatgtggccatcaagatgatcaaggagggctcaatgagcgaggacgagttcatcgaggaggctaaggtcatgatgaacctg tcccacgagaaactggtgcagctgtatggagtgtgcaccaagcagcggcccatttttatcattacagagtacatggctaatgg gtgtctgctgaactatctgcgcgagatgagacacagattccagacacagcagctgctggaaatgtgcaaggatgtgtgtgagg ctatggagtacctggagtctaagcagtttctgcaccgggacctggctgctcgcaattgcctggtgaacgatcagggcgtggtg aaggtgagtgacttcggactgtcaaggtatgtgctggatgacgagtacaccagctccgtgggctctaagtttcctgtgagatg gtctccacccgaggtgctgatgtatagcaagttctcctctaagagcgatatctgggcctttggcgtgctgatgtgggaaatct acagcctgggcaagatgccttacgagcggttcacaaattccgagacagctgagcacatcgcccagggcctgcgcctgtaccgg ccacatctggcctctgagaaggtgtacaccatcatgtacagctgttggcacgagaaggccgacgagagacccacattcaagat cctgctgtccaacattctagatgtgatggacgaggagagctgagtcgacaatcaacctctggattacaaaatttgtgaaagat tgactggtattcttaactatgttgctccttttacgctatgtggatacgctgctttaatgcctttgtatcatgctattgcttcc cgtatggctttcattttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacg tggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactt tcgctttccccctccctattgccacggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggc actgacaattccgtggtgttgtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgg gacgtccttctgctacgtcccttcggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcggcctcttc cgcgtcttcgccttcgccctcagacgagtcggatctccctttgggccgcctccccgcctggaattcgagctcggtacctttaa gaccaatgacttacaaggcagctgtagatcttagccactttttaaaagaaaaggggggactggaagggctaattcactcccaa cgaagacaagatctgctttttgcttgtactgggtctctctggttagaccagatctgagcctgggagctctctggctaactagg gaacccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaact agagatccctcagacccttttagtcagtgtggaaaatctctagcagtagtagttcatgtcatcttattattcagtatttataa cttgcaaagaaatgaatatcagagagtgagaggaacttgtttattgcagcttataatggttacaaataaagcaatagcatcac aaatttcacaaataaagcatttttttcactgcattctagttgtggtttgtccaaactcatcaatgtatcttatcatgtctggc tctagctatcccgcccctaactccgcccatcccgcccctaactccgcccagttccgcccattctccgccccatggctgactaa ttttttttatttatgcagaggccgaggccgcctcggcctctgagctattccagaagtagtgaggaggcttttttggaggccta gggacgtacccaattcgccctatagtgagtcgtattacgcgcgctcatcggatcccgggcccgtcgactgcagaggcctgcat gcaagcttggtgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgctcacaattccacacaacatacgagccgg aagcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaattgcgttgcgctcactgcccgctttccagt cgggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgct tcctcgctcactgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacggttatc cacagaatcaggggataacgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgc tggcgtttttccataggctccgcccccctgacgagcatcacaaaaatcgacgctcaagtcagaggtggcgaaacccgacagga ctataaagataccaggcgtttccccctggaagctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtc cgcctttctcccttcgggaagcgtggcgctttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgttcgctcca agctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgccttatccggtaactatcgtcttgagtccaacccggta agacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgaggtatgtaggcggtgctacagagttctt gaagtggtggcctaactacggctacactagaagaacagtatttggtatctgcgctctgctgaagccagttaccttcggaaaaa gagttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttttttgtttgcaagcagcagattacgcgcaga aaaaaaggatctcaagaagatcctttgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggatttt ggtcatgagattatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaagcccaatctgaataa tgttacaaccaattaaccaattctgattagaaaaactcatcgagcatcaaatgaaactgcaatttattcatatcaggattatc aataccatatttttgaaaaagccgtttctgtaatgaaggagaaaactcaccgaggcagttccataggatggcaagatcctggt atcggtctgcgattccgactcgtccaacatcaatacaacctattaatttcccctcgtcaaaaataaggttatcaagtgagaaa tcaccatgagtgacgactgaatccggtgagaatggcaaaagtttgtgcatttctttccagacttgttcaacaggccagccatt acgctcgtcatcaaaatcactcgcatcaaccaaaccgttattcattcgtgattgcgcctgagcgagacgaaatacgcgatcgc tgttaaaaggacaattacaaacaggaatcgaatgcaaccggcgcaggaacactgccagcgcatcaacaatattttcacctgaa tcaggatattcttctaatacctggaatgctgtttttccggggatcgcagtggtgagtaaccacgcatcatcaggagtacggat aaaatgcttgatggtcggaagaggcataaattccgtcagccagtttagtctgaccatctcatctgtaacatcattggcaacgc tacctttgccatgtttcagaaacaactctggcgcatcgggcttcccatacaagcgatagattgtcgcacctgattgcccgaca ttatcgcgagcccatttatacccatataaatcagcatccatgttggaatttaatcgcggcctcgacgtttcccgttgaatatg gctcataacaccccttgtattactgtttatgtaagcagacagttttattgttcatgatgatatatttttatcttgtgcaatgt aacatcagagattttgagacacgggccagagctgca Vector TL20c-XLA LV (SEQ ID NO:21) ggccgcctcggccaaacagcccttgagtttaccactccctatcagtgatagagaaaagtgaaagtcgagtttaccactcccta tcagtgatagagaaaagtgaaagtcgagtttaccactccctatcagtgatagagaaaagtgaaagtcgagtttaccactccct atcagtgatagagaaaagtgaaagtcgagtttaccagtccctatcagtgatagagaaaagtgaaagtcgagtttaccactccc tatcagtgatagagaaaagtgaaagtcgagtttaccactccctatcagtgatagagaaaagtgaaagtcgagctcgccatggg aggcgtggcctgggcgggactggggagtggcgagccctcagatcctgcatataagcagctgctttttgcctgtactgggtctc tctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctcaataaagcttgccttga gtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttttagtcagtgtggaaaat ctctagcagtggcgcccgaacagggacttgaaagcgaaagggaaaccagaggagctctctcgacgcaggactcggcttgctga agcgcgcacggcaagaggcgaggggcggcgactggtgagtacgccaaaaattttgactagcggaggctagaaggagagagatg ggtgcgagagcgtcagtattaagcgggggagaattagatcgcgatgggaaaaaattcggttaaggccagggggaaagaaaaaa tataaattaaaacatatagtatgggcaagcagggagctagaacgattcgcagttaatactggcctgttagaaacatcagaagg ctgtagacaaatactgggacagctacaaccatcccttcagacaggatcagaagaacttagatcattatataatacagtagcaa ccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttagacaagatagaggaagagcaaaacaaaagt aagaaaaaagcacagcaagcagcaggatcttcagacctggaaattccctacaatccccaaagtcaaggagtagtagaatctat gaataaagaattaaagaaaattataggacaggtaagagatcaggctgaacatcttaagacagcagtacaaatggcagtattca tccacaattttaaaagaaaaggggggattggggggtacagtgcaggggaaagaatagtagacataatagcaacagacatacaa actaaagaattacaaaaacaaattacaaaaattcaaaattttcgggtttattacagggacagcagaaatccactttggaaagg accagcaaagctcctctggaaaggtgaaggggcagtagtaatacaagataatagtgacataaaagtagtgccaagaagaaaag caaagatcattagggattatggaaaacagatggcaggtgatgattgtgtggcaagtagacaggatgaggattagaacatggaa aagtttagtaaaacaccataaggaggagatatgagggacaattggagaagtgaattatataaatataaagtagtaaaaattga accattaggagtagcacccaccaaggcaaagagaagagtggtgcagagagaaaaaagagcagtgggaataggagctttgttcc ttgggttcttgggagcagcaggaagcactatgggcgcagcgtcaatgacgctgacggtacaggccagacaattattgtctggt atagtgcagcagcagaacaatttgctgagggctattgaggcgcaacagcatctgttgcaactcacagtctggggcatcaagca gctccaggcaagaatcctggctgtggaaagatacctaaaggatcaacagctcctggggatttggggttgctctggaaaactca tttgcaccactgctgtgccttggaatgctagttggagtaataaatctctggaacagatttggaatcacacgacctggatggag tgggacagagaaattaacaattacacaagcttaatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaaca agaattattggaattagataaatgggcaagtttgtggaattggtttaacataacaaattggctgtggtatataaaattattca taatgatagtaggaggcttggtaggtttaagaatagtttttgctgtactttctatagtgaatagagttaggcagggatattca ccattatcgtttcagacccacctcccaaccccgaggggaccgagctcaagcttcgaagcgatcgcccggaaacacccgaatca acttctagtcaaattattgttcacgccgcaatgacccacccctggcccgcgtctgtggaactgacccctggtgtacaggagag ttcgctgctgaaagtggtcccaaaggggtactagtttttaagctcccaactccccctcccccagcgtctggaggattccacac cctcgcaccgcaggggcgaggaagtgggcggagtccggttttggcgccagccgctgaggctgccaagcagaaaagccaccgct gaggagactccggtcactgtcctcgccccgcctcccccttccctccccttggggaccaccgggcgccacgccgcgaacgcgaa ctgccgcggtccgcgccgcctccgccctcccccttgggccccaattcccagcgggcgcggcgcgcggcccctccccccgccgg gcgcgcgcccgctgccccgcccttcgtggccgcccggcgtgggcggtgccacccctccccccggcggccccgcgcgcagctcc cggctccctcccccttcggatgtggcttgagctgtaggcgcggagggccggagacgctgcagacccgcgacccggagcagctc ggaggcggtgaacgcgctggctttccttctctctagctctcgctcgctggtggtgcttcagatgccacacgcgtccggggcta gctacgacgcgttccggaattcgcccttgcatttcctaggagaatccctgggggaatcattgcagttggagcataatgtaggg ggcccctgagaaaacctccaggcttcaagtgacatacctagtctgctttaccggtttacaggactcaagagaaaggtggacat tgagagttaatccctgaggccaaatcttaaatggagaaagtcaacatccacagaaaatggggaagggcacaagtatttctgtg ggcttatattccgacatttttatctgtaggggaaaaatgctttcttagaaaatgactcagcacggggaagtcttgtctctacc tctgtcttgttttgtcctttggggtcccttcactatcaagttcaactgtgtgtccctgagactcctctgccccggaggacagg agactcgaaaaacgctcttcctggccagtctctttgctctgtgtctgccagcccccagcatctctcctctttcctgtaagccc ctctccctgtgctgactgtcttcatagtactttaggtatgttgtccctttacctctgggaggatagcttgatgacctgtctgc tcaggccagccccatctagagtctcagtggccccagtcatgttgagaaaggttctttcaaagatagactcaagatagtagtgt cagaggtcccaagcaaatgaagggcggggacagttgagggggtggaatagggacggcagcagggaaccagatagcatgctgct gagaagaaaaaaagacattggtttaggtcaggaagcaaaaaaagggaactgagtggctgtgaaagggtggggtttgctcagac tgtccttcctctctggactgtaagaattagtctcgagaaagaagccaccatggccgctgtgatcctggagagcattttcctga agaggtcccagcagaaaaagaaaacctctcccctgaactttaagaaaagactgttcctgctgacagtgcacaagctgtcttac tatgagtacgactttgagcggggccgccgaggatcaaaaaaggggagcatcgatgtggagaagattacatgcgtggagaccgt ggtccctgaaaagaatccaccccctgagaggcagatcccaagacggggcgaggagtcctctgagatggagcagattagtatca ttgagcgcttcccctatccttttcaggtggtgtacgacgagggaccactgtatgtgttctcacccacagaggagctgagaaag aggtggattcaccagctgaagaacgtgattagatacaatagcgatctggtgcagaagtatcacccttgtttttggatcgacgg gcagtacctgtgctgttcccagacagctaagaacgctatgggatgccagattctggaaaatcggaacggatctctgaaaccag ggagttcacaccgcaagaccaaaaagcccctgcctccaacacccgaggaggatcagatcctgaaaaagcctctgccacccgag cctgctgcagccccagtcagcacttccgaactgaaaaaggtggtggctctgtatgactacatgcccatgaatgctaacgatct gcagctgagaaagggcgacgagtatttcattctggaagagtctaatctgccttggtggagggccagagataagaacggacagg aggggtacatcccatctaattatgtgaccgaggctgaggactctattgagatgtacgagtggtatagcaagcacatgacacgg tcccaggctgagcagctgctgaagcaggagggcaaagagggagggtttatcgtgcgcgattctagtaaggccggcaaatacac tgtgtcagtgttcgctaagagcaccggagacccccagggcgtgatcagacactatgtggtgtgttccacacctcagtctcagt actatctggctgagaagcacctgtttagtacaatcccagagctgattaactaccaccagcacaattctgccggcctgatcagc aggctgaagtatcccgtctcccagcagaacaaaaatgctccttctaccgctggactggggtacggcagttgggagattgatcc aaaggacctgacattcctgaaggagctgggaactgggcagtttggcgtggtgaagtatggaaaatggagagggcagtacgatg tggccatcaagatgatcaaggagggctcaatgagcgaggacgagttcatcgaggaggctaaggtcatgatgaacctgtcccac gagaaactggtgcagctgtatggagtgtgcaccaagcagcggcccatttttatcattacagagtacatggctaatgggtgtct gctgaactatctgcgcgagatgagacacagattccagacacagcagctgctggaaatgtgcaaggatgtgtgtgaggctatgg agtacctggagtctaagcagtttctgcaccgggacctggctgctcgcaattgcctggtgaacgatcagggcgtggtgaaggtg agtgacttcggactgtcaaggtatgtgctggatgacgagtacaccagctccgtgggctctaagtttcctgtgagatggtctcc acccgaggtgctgatgtatagcaagttctcctctaagagcgatatctgggcctttggcgtgctgatgtgggaaatctacagcc tgggcaagatgccttacgagcggttcacaaattccgagacagctgagcacatcgcccagggcctgcgcctgtaccggccacat ctggcctctgagaaggtgtacaccatcatgtacagctgttggcacgagaaggccgacgagagacccacattcaagatcctgct gtccaacattctagatgtgatggacgaggagagctgagtcgacaatcaacctctggattacaaaatttgtgaaagattgactg gtattcttaactatgttgctccttttacgctatgtggatacgctgctttaatgcctttgtatcatgctattgcttcccgtatg gctttcattttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacgtggcgt ggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactttcgctt tccccctccctattgccacggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggcactgac aattccgtggtgttgtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgggacgtc cttctgctacgtcccttcggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgtc ttcgccttcgccctcagacgagtcggatctccctttgggccgcctccccgcacgtacgaccggtgcggccgcatcgatgccgt agtacctttaagaccaatgacttacaaggcagctgtagatcttagccactttttaaaagaaaaggggggactggaagggctaa ttcactcccaaagaagacaagatagatctgctttttgcctgtactgggtctctctggttagaccagatctgagcctgggagct ctctggctaactagggaacccactgcttaagcctcaataaagcttcagctgctcgagctagcagatctttttccctctgccaa aaattatggggacatcatgaagccccttgagcatctgacttctggctaataaaggaaatttattttcattgcaatagtgtgtt ggaattttttgtgtctctcactcggaaggacatatgggagggcaaatcatttaaaacatcagaatgagtatttggtttagagt ttggcaacatatgcccatatgctggctgccatgaacaaaggttggctataaagaggtcatcagtatatgaaacagccccctgc tgtccattccttattccatagaaaagccttgacttgaggttagattttttttatattttgttttgtgttatttttttctttaa catccctaaaattttccttacatgttttactagccagatttttcctcctctcctgactactcccagtcatagctgtccctctt ctcttatggagatccctcgacctgcagcccaagcttggcgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgc tcacaattccacacaacatacgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaatt gcgttgcgctcactgcccgctttccagtcgggaaacctgtcgtgccagcggatccgcatctcaattagtcagcaaccatagtc ccgcccctaactccgcccatcccgcccctaactccgcccagttccgcccattctccgccccatggctgactaattttttttat ttatgcagaggccgaggccgcctcggcctctgagctattccagaagtagtgaggaggcttttttggaggcctaggcttttgca aaaagctgtcgactgcagaggcctgcatgcaagcttggcgtaatcatggtcatagctgtttcctgtgtgaaattgttatccgc tcacaattccacacaacatacgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaatt gcgttgcgctcactgcccgctttccagtcgggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggagagg cggtttgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgctcggtcgttcggctgcggcgagcggtatcag ctcactcaaaggcggtaatacggttatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaaggccagcaaaag gccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgcccccctgacgagcatcacaaaaatcgacgctc aagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctggaagctccctcgtgcgctctcctgttc cgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctcatagctcacgctgtaggtat ctcagttcggtgtaggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgccttatccgg taactatcgtcttgagtccaacccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcga ggtatgtaggcggtgctacagagttcttgaagtggtggcctaactacggctacactagaagaacagtatttggtatctgcgct ctgctgaagccagttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttttt tgtttgcaagcagcagattacgcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctgacgctcagt ggaacgaaaactcacgttaagggattttggtcatgagattatcaaaaaggatcttcacctagatccttttaaattaaaaatga agttttaaatcaatctaaagtatatatgagtaaacttggtctgacagttaccaatgcttaatcagtgaggcacctatctcagc gatctgtctatttcgttcatccatagttgcctgactccccgtcgtgtagataactacgatacgggagggcttaccatctggcc ccagtgctgcaatgataccgcgagacccacgctcaccggctccagatttatcagcaataaaccagccagccggaagggccgag cgcagaagtggtcctgcaactttatccgcctccatccagtctattaattgttgccgggaagctagagtaagtagttcgccagt taatagtttgcgcaacgttgttgccattgctacaggcatcgtggtgtcacgctcgtcgtttggtatggcttcattcagctccg gttcccaacgatcaaggcgagttacatgatcccccatgttgtgcaaaaaagcggttagctccttcggtcctccgatcgttgtc agaagtaagttggccgcagtgttatcactcatggttatggcagcactgcataattctcttactgtcatgccatccgtaagatg cttttctgtgactggtgagtactcaaccaagtcattctgagaatagtgtatgcggcgaccgagttgctcttgcccggcgtcaa tacgggataataccgcgccacatagcagaactttaaaagtgctcatcattggaaaacgttcttcggggcgaaaactctcaagg atcttaccgctgttgagatccagttcgatgtaacccactcgtgcacccaactgatcttcagcatcttttactttcaccagcgt ttctgggtgagcaaaaacaggaaggcaaaatgccgcaaaaaagggaataagggcgacacggaaatgttgaatactcatactct tcctttttcaatattattgaagcatttatcagggttattgtctcatgagcggatacatatttgaatgtatttagaaaaataaa caaataggggttccgcgcacatttccccgaaaagtgccacctgacgtctaagaaaccattattatcatgacattaacctataa aaataggcgtatcacgaggccctttcgtctcgcgcgtttcggtgatgacggtgaaaacctctgacacatgcagctcccggaga cggtcacagcttgtctgtaagcggatgccgggagcagacaagcccgtcagggcgcgtcagcgggtgttggcgggtgtcggggc tggcttaactatgcggcatcagagcagattgtactgagagtgcaccatatgcggtgtgaaataccgcacagatgcgtaaggag aaaataccgcatcaggcgccattcgccattcaggctgcgcaactgttgggaagggcgatcggtgcgggcctcttcgctattac gccagctggcgaaagggggatgtgctgcaaggcgattaagttgggtaacgccagggttttcccagtcacgacgttgtaaaacg acggccagtgaattc CBX3 promoter (SEQ ID NO:22) ccggaaacacccgaatcaacttctagtcaaattattgttcacgccgcaatgacccacccctggcccgcgtctgtggaactgac ccctggtgtacaggagagttcgctgctgaaagtggtcccaaaggggtactagtttttaagctcccaactccccctcccccagc gtctggaggattccacaccctcgcaccgcaggggcgaggaagtgggcggagtccggttttg

Claims

WHAT IS CLAIMED IS:

1. A vector suitable for the treatment of X-linked agammaglobulinemia (XLA) or for inhibiting or ameliorating at least one symptom of XLA, the vector comprising: (a) a transgene expression cassette comprising a nucleotide sequence encoding Bruton's tyrosine kinase (BTK) operably linked to a promoter; (b) at least one regulatory or enhancer element; (c) a ubiquitous chromatin opening element (UCOE), wherein the UCOE comprises a nucleotide sequence at least about 90% identical to the sequence set forth in SEQ ID NO:1, and wherein the sequence of the UCOE comprises the following nucleotide substitutions relative to SEQ ID NO:1, at positions corresponding to those of SEQ ID NO:1: (i) a G residue at position 4; (ii) the sequence CGAAC at positions 347-351; (iii) the sequence CGCGC at positions 362-366; (iv) a T residue at position 385; (v) a G residue at position 496; (vi) the sequence CGCGC at positions 612-616; and (vii) a T residue at position 673.

2. The vector of claim 1, wherein the nucleotide sequence encoding BTK is codon optimized for expression in humans.

3. The vector of claim 1 or 2, wherein the nucleotide sequence encoding BTK comprises the sequence set forth in SEQ ID NO:4 or SEQ ID NO:

5.

4. The vector of any one of claims 1 to 3, wherein the promoter in the transgene expression cassette is a B cell specific promoter.

5. The vector of any one of claims 1 to 4, wherein the promoter in the transgene expression cassette is the BTK promoter, optionally comprising the sequence set forth in SEQ ID NO:

6.

6. The vector of any one of claims 1 to 5, wherein the transgene expression cassette comprises the sequence set forth in SEQ ID NO:

7.

7. The vector of any one of claims 1 to 6, wherein the UCOE is in the forward orientation in the vector with respect to the transgene expression cassette.

8. The vector of any one of claims 1 to 7, wherein the UCOE and the transgene expression cassette comprise the sequence set forth in SEQ ID NO:

8.

9. The vector of any one of claims 1 to 8, wherein the UCOE comprises the nucleotide sequence of SEQ ID NO:2, or a sequence at least about 90% identical thereto, preferably at least about 95%, 96%, 97%, 98%, 98.5%, 99% or 99.5% identical thereto.

10. The vector of any one of claims 1 to 9, wherein the UCOE is at least about 0.7kb in length.

11. The vector of any one of claims 1 to 10, wherein the UCOE comprises or consists of the nucleotide sequence of SEQ ID NO:

2.

12. The vector of any one of claim 1 to 11, wherein the at least one regulatory element is an insulator, a Woodchuck Hepatitis Virus (WHV) Posttranscriptional Regulatory Element (WPRE), or a combination thereof, optionally wherein the regulatory element is located downstream of the transgene expression cassette.

13. The vector of any one of claims 1 to 12, wherein the vector is a viral vector.

14. The vector of claim 13, wherein the vector is a retroviral vector.

15. The vector of claim 14, wherein the vector is a lentiviral vector.

16. The vector of claim 15, comprising a sequence set forth in SEQ ID NO:20 or a sequence having at least about 90% sequence identity thereto, comprising the UCOE from position 2396 to 3068 of SEQ ID NO:

20.

17. The vector of claim 15, comprising a sequence set forth in SEQ ID NO:21 or a sequence having at least about 90% sequence identity thereto, comprising the UCOE from position 2390 to 3062 of SEQ ID NO:

21.

18. A cell comprising, or transduced with, the vector of any one of claims 1 to 17.

19. The cell of claim 18, wherein the cell is a B cell, a myeloid cell or a hematopoietic stem cell.

20. The cell of claim 19, wherein the hematopoietic stem cell is a CD34+hematopoietic stem cell.

21. A method for promoting B cell survival, proliferation and / or differentiation in a subject in need thereof, the method comprising administering to the subject a vector of any one of claims 1 to 17 or a cell of any one of claims 18 to 20, and optionally measuring B cell survival, proliferation and / or differentiation in the subject or in a biological sample obtained from the subject.

22. The method of claim 21, further comprising the step, prior to administration, of identifying the subject as one that would benefit from receiving a therapy to promote B cell survival, proliferation and / or differentiation.

23. A method of treating X-linked agammaglobulinemia (XLA) or inhibiting or ameliorating at least one symptom thereof, in a subject, the method comprising administering to the subject a vector of any one of claims 1 to 17 or a cell of any one of claims 18 to 20, and optionally measuring an improvement in the progression of XLA or at least one symptom thereof in the subject or in a biological sample obtained from the subject.

24. The method of claim 23, further comprising the step, prior to administration, of identifying the subject as one that would benefit from receiving a therapy to treat X- linked agammaglobulinemia (XLA) or to inhibit or ameliorate at least one symptom thereof.

25. The method of any one of claims 21 to 24, wherein the cell is administered by adoptive cell transfer.

26. Use of the vector of any one of claims 1 to 17 or the cell of any one of claims 18 to 21 in the preparation of a medicament for promoting B cell survival, proliferation and / or differentiation, or for the treatment of XLA or the inhibition or amelioration of at least one symptom thereof.